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No theory is worth anything if it can't be applied effectively, so this month and next, I'll build a small but nontrivial application that demonstrates the object-oriented user-interface principles I've covered over the last few months: an RPN (Reverse Polish Notation) calculator that can work as either a tape calculator or a keypad calculator. (One of the main things I want to demonstrate is the flexibility of the UI; thus the two presentations.) Since this application is a real one, I need to do some groundwork before covering the calculator itself.
Consequently, this month's Java Toolbox takes the name of the column literally and presents the set of generic tools we'll need to build the calculator -- little tools of general utility that should be of use for more than the current application. Specifically, I'll cover the following:
|An implementation of an "assertion" that lets you implement pre- and postconditions.
|A class that aids in putting together automated unit tests.
|The Java equivalent of /dev/null, it throws away characters.
|A "singleton" wrapper around standard input and output that makes it easy to access these two streams.
|A convenient class for logging diagnostic messages.
|A utility containing various methods for aligning text.
|A JTextArea that has scroll bars around it.
Most of those classes are unrelated to one another, and none of them is significant enough to merit a article devoted exclusively to it, so lumping them all together in one place seems sensible. We've taken a quick look at a few of the tools in the past, but a review should prove helpful. Now, on to the tools.
TEXTBOX: TEXTBOX_HEAD: Build user interfaces for object-oriented systems: Read the whole series!
- Part 1. What is an object? The theory behind building object-oriented user interfaces
- Part 5. Useful stuff
Part 6. An RPN-calculator application demonstrates object-oriented UI principles
Bertrand Meyer, the author of the Eiffel language, came up with the notion of pre- and postconditions, commonly called assertions. Eiffel, described in Object-Oriented Software Construction (see Resources) supports those concepts directly within the syntax of the language. Java, unfortunately, provides no such support.
The basic idea of a precondition or postcondition is that every method expects the world (the object that it's working on, the global state of the program, the arguments to the method, and so on) to be in a particular state when the method is called, and that every method will leave the world in a particular state. An assertion is a body of code that guarantees that those conditions hold and typically terminates the program with an exception toss if they do not. Assertions are typically implemented as debug-time tests and removed from the code entirely when the code goes into production. (The removal can be risky since you are modifying tested code, but on the other hand, the assertions can slow down the execution of a program noticeably.)
I've implemented assertions using the two classes defined in Lists 1 and 2. The first version, in the
com.holub.tools package, is made up of empty methods. Import this version into your code when you've finished debugging. The second version, in the
com.holub.tools.debug package (List 2), actually does something. Import this version when you're still working on the code.
When you use the class like this:
Assert.is_true( condition );
the program terminates by throwing an
Assert.Failed object if the condition is not true. A second version lets you add a reasonable error message to the thrown object:
Assert.is_true( condition, "Error that caused the problem" );
I've also provided
is_false() variants as a programming convenience.
The main problem with my strategy of using an empty method to eliminate the overhead of the assertion is that the resulting efficiency is somewhat JVM dependent. Hotspot, for example, does everything right. It notices that the method is empty, so expands it inline, effectively removing the call from the
.class file. Hotspot even removes the argument to the assertion (effectively dead code). The last behavior can be important if you use string concatenation in an argument to
is_true() or its equivalent. You don't want to go through the considerable overhead of building a string with a series of concatenations if the string isn't used for anything. Earlier JVMs were not as good about this optimization: they would correctly eliminate the call itself but typically wouldn't eliminate the argument evaluation.
Finally, note that method calls in assertions are risky. Hotspot will evidently inline them up to a point, but it won't chase down the call graph indefinitely. Generally, method calls should appear in assertions only if they do nothing other than return constant values. If you want to test a method's return value, call the method first, then test the resulting value in a subsequent assertion.
Of course, methods that have side effects, which modify the state of the program, should never be invoked from within an assertion because they'll go away when you move from the debugging to the production version of the code.
Let's look at List 1:
Next, we take a look at List 2:
Help in unit testing
The next helper class of interest is the
Tester, which makes it easier to do unit testing (the standalone testing of a single class).
Test shows a typical use of this class. I typically put my unit tests in a
Test inner class of the class I'm testing. (I use an inner class so that the code that makes up the test won't be part of the class file that represents the class I'm testing. Inner classes create their own class files, which in this case do not have to be shipped with the production code.)
I have several criteria for what constitutes proper test behavior. A formal test must:
- Not print anything unless something is wrong
- Have a unique ID
- Test reasonable boundary conditions and "impossible" inputs
The first item is particularly important because, in an automated testing environment, you don't want to clutter up reports with worthless information. A test typically tries to find out when something goes wrong, so that's the only time that output should be generated. On the other hand, I occasionally do get paranoid, so I've provided a hook that modifies the behavior of my class to print both success and failure messages.
The tester is initialized with a normal constructor call that has passed two arguments: first, a Boolean flag that indicates whether the output is verbose (success messages are printed in addition to failures) or not (only failure messages are printed); second, a
PrintWriter that represents the stream to which output is sent. Thereafter, you run a test by calling the
check() method, which has several overloads.
The arguments are:
- The test ID
- The expected output of a method call
- The method call to test
The overloads change the types of the second and third arguments to handle various possible return values. Supported return values are
double (which will handle
float via the normal type conversion),
long (which will also handle
int, and so on, via the normal type conversions), and
boolean. There is only one requirement: the second argument's type must be the same as the return type of the method call in the third argument.
The final method of interest in the
Tester is the
exit() method, which causes the program to exist with the current error count as its exit status. This mechanism lets you write shell scripts that run tests and detect when something goes wrong.
The code, which is largely self-explanatory, is in List 4:


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JET SCOTT VOLUME 1 Sheldon Stark (W) and Jerry Robinson (A) Scheduled to arrive on Feb 10 Full Color. 216 pages HC, 12" x 9" In the 1950s, when the world was faced with strange or anomalous threats, there was one man who was called on to set the situation right-Jet Scott of the Office of Scientifact! Whether it was tracking down deadly Banthrax germs, uncovering the source of strange ocean creatures, or discovering the cause of spontaneously combusting pipelines in Saudi Arabia, Jet Scott was the adventurer who could get to the truth! Jerry Robinson is one of the best known of all the artists who have drawn Batman over the years and is often credited with creating the Joker, one of Comics' greatest villains! Dark Horse Comics is proud to collect the Jet Scott comic strips, for the first time ever, which were originally published from 1953 to 1955 by the New York Herald Tribune!
Jet Scott Vol. 01 HC
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subset: commoncrawl


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Is Your Back Pain Caused by a Degenerated Disc or a Pinched Nerve?
Back pain can severely disrupt one's life, as sufferers already very well know. It can affect everything from your job performance and social interactions down to your general enjoyment of daily living. While temporary solutions such as pain medication and compresses can provide disc pain relief in Paramus, NJ, you might soon find the pain returning if the root causes of your pain are not addressed.
Sometimes, back pain can have a variety of causes such as a pinched nerve or a herniated disc, and treatment can vary depending on which one you are afflicted with.
Read more from this blog.


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Some thoughts and info about the photo (pensive) … The photo was taken at a School function, where the girl was somewhat bored or pensive. She was in the audience. Glad I had my cam with me, when I turned … Continue reading
Tag Archives: winner
2nd Winners Event-Wow Factor-Cheyl Penn Hidden & Evie Go
Chreyl Penn Hidden with Bourkes Luck Pot Holes
Sony DSC-P41
Focal length: 5mm
Exp. time: 1/400
F-stop: 5.6
Landscape mode
Gee, I was so amazed and honoured to have a 1st Placing for my photo. Being of such personal significance, it has made me quite teary.
The photo of Bourkes Luck Pot Holes came about after my parents passed away. I really wanted to go and visit the area, that they travelled to annually in Mpumalanga (North Eastern area of S. Africa). This has amazing natural geological rock formations, waterfalls and is also near our Kruger National Game Reserve. They would travel in their "Trekliner" – equivalent to an RV.
To be making this journey, which was an emotional tribute to my parents, for me, I needed to travel in the footsteps of my parents and visit all the places that they did – to discover the attraction that Mpumalanga (then called the Eastern Transvaal) held for them.
I was in awe of the vast mountainous area. Where to start ??
The Bourke's Luck Potholes are undoubtedly one of the area's finest attractions.
This natural (and most beautiful) water feature, formed over thousands of years by water erosion, marks the beginning of the stunning Blyde River Canyon – one of the largest canyons in the world and said to be the world's largest 'green canyon'.
The Bourke's Luck Potholes occur at the confluence of the Treur and the Blyde rivers. Over the eons, the swirling, eddying, and plunging waters of the Treur River, waterborne sand and pebbles in tow, formed huge cylindrical potholes in the bedrock of the Blyde River. Mother Nature's vast forces left us with a truly stunning natural phenomenon – absolutely fascinating geological sculptures delightfully 'painted' in fantastic white, yellow, and reddish brown hues.
I hadn't quite thought of potholes as being amazing, works of natural art…. potholes were to be filled in roads as quickly as possible, but I was enthralled. A 700 metre path alongside the potholes leads one to the bridge, which one can cross (safely) to the other side to sit on the rocks, dabble feet in the little stream making its way down to the water below and relax watching tiny fish.
Other places of interest in the area which we visited, which can be googled, are "God's Window" and "The Blyde River Canyon", "The Three Rondavels" apart from many waterfalls which my parents used to speak of, and other historical places.
Evie Go – Angkow Wat.
I dont know what to say abt the Angkor Wat pic…except that i have always wanted to see the ruins & never got the chance until some FB friends who loves to travel also started planning & we all just got together & set it! It was kinda scary, as I dont know these people. We all just met at the airport & it turned out to be one of our best vacations ever! We had so much fun! I love my FB friends all over the world! I used my Sony TX5


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Define LVN
An LVN is a licensed vocational nurse, alternately called an LPN, or licensed practical nurse. The training, licensing and responsibilities of LVNs and LPNs are similar. However, California and Texas use the designation LVN, while the other states use LPN. An LVN or LPN works under the supervision of physicians and registered nurses to provide basic patient care in accordance with the laws of each state.
Education and Licensing
Each LVN or LPN must complete an educational training program accepted by the state board of nursing. Hospitals, two-year colleges and techni,cal schools offer certificate programs, which take approximately one year. In addition, some high schools offer vocational nurse training. LVNs and LPNs must also pass the National Council Licensure Examination, or NCLEX, at the PN or practical nurse level, and complete other state requirements for licensing.
Job Environment
As of 2010, approximately 75 percent of LVNs and LPNs had full-time work schedules, according to the U.S. Bureau of Labor Statistics. Many vocational nurses work shifts longer than 8 hours, and many work holidays, weekends and nights. As of 2010, 29 percent worked in nursing care, 15 percent worked in hospitals and 12 percent worked in doctor's offices, according to the BLS. An additional 9 percent were employed in home heath, and 5 percent worked in community elder care facilities.
Job Duties
The duties of an LVN or LPN depend on state law and the needs of the particular employer. Under the supervision of other health professionals, they typically check blood pressure, help to bathe and feed patients, change dressings, keep records and collect lab samples. When allowed by law, some LVNs and LPNs also perform more advanced tasks, such as beginning intravenous drips or giving medicines.
As of May 2011, the average LVN or LPN nationwide earned $42,040 per year, according to the Bureau of Labor Statistics. Those working in nursing care facilities averaged $43,090 per year, while hospital nurses averaged $41,060 per year. Nurses working in doctor's offices averaged 38,440 per year. Home health nurses averaged $43,160 per year, while community care nurses received an average of $42,960. In those states using the LVN designation, Texas and California, vocational nurses averaged $42,260 and $51,130 per year respectively as of 2011.
The number of jobs for licensed vocational and practical nurses will increase by 22 percent nationwide between 2010 and 2020, which compares to a predicted average of 14 percent for all U.S., jobs, according to the Bureau of Labor Statistics. The aging of the U.S. population and the trend toward moving procedures out of hospitals will increase the need for LVNs in assisted living facilities and nursing homes. The expected retirement of many current nurses will contribute to excellent job prospects for licensed vocational nurses.
- U.S. Bureau of Labor Statistics Occupational Outlook Handbook: What Licensed Practical and Licensed Vocational Nurses Do
- American Nurses Association: What Nurses Do
- U.S. Bureau of Labor Statistics Occupational Outlook Handbook: Licensed Practical and Licensed Vocational Nurses -- How to Become One
- U.S. Bureau of Labor Statistics Occupational Outlook Handbook: Licensed Practical and Licensed Vocational Nurses -- Work Environment
- U.S. Bureau of Labor Statistics Occupational Employment and Wages, May 2011: Licensed Practical and Licensed Vocational Nurses
- U.S. Bureau of Labor Statistics Occupational Outlook Handbook: Licensed Practical and Licensed Vocational Nurses -- Job Outlook
Photo Credits
- Jupiterimages/Creatas/Getty Images


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Bamboo Cotton Buds1 × £1.95 ×
Subtotal: £1.95
Total: £1.95
Bright and uplifting orange, lemon and lime essential oil within our handmade shampoo bars. perfect for people with greasy hair as the citrus really helps to cut through any unwanted grease
Our Conditioner bars packed full of butters and oils to help dry hair. and help nourish the hair back to a natural glow. simply rub the bar over your hair or into your hands to transfer oils onto your hair. don't apply too much, as could become greasy
Birthday cake scented with sugar sprinkles on top… good enough too eat… but no… don't… as it doesn't taste nice…
only a few of these avalible. but thease are a complete mystery.. could be bath bombs / whipped soaps/ shower gels/. mini bombs/ dusts/ foot soaks.. but youll get a mystery bag worth at LEAST £20


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Download Smart Recovery Tools
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Using the software is child's play. PC INSPECTOR™ smart recovery runs on all Win 9x, ME, NT 4.0, XP and Windows™ 2000 systems.
PC INSPECTOR™ smart recovery the software with which you can recover your holiday pictures in seconds after unintentional deletion or a system error.
PC INSPECTOR™ smart recovery is also available bundled with many high-quality digital cameras – ask your salesperson.
PC INSPECTOR™ smart recovery supports the following picture formats: .jpg, .amr, .tif, .bmp, .gif, Canon .crw , Fuji .raf , RICOH .raw, Olympus .orf (E-XX), Olympus .orf (C5050), Nokia 3gp, Kodak .dcr, Minolta .mrw, Nikon .nef (D1H/D1X), Nikon .nef (D2H/D2X), Nikon .nef (E5000/E5700), Sigma – Foveon .x3f, mp4, QuickTime .mov (Konica Minolta). Video files in .avi, QuickTime .mov format and audio files in .wav, .dss format can also be recovered.
File Size: 5.36 MB
Filed under: Download Software, Hard Disk Utilities, System Utilities | Tagged: Data Recovery | Leave a comment »


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More events:
An absolute masterpiece, with its unique mix of fantasy elements, musical, horsemanship, sound and lighting effects and an integral part ofEQUITANA – Equestrian World Trade Fair
"The HOP TOP Show is considered to be the most exceptional and most impressive equestrian show on earth." - WDR
On five evenings you can experience this unique horse show.
The dates will be published here.
This unique equestrian event can only be experienced on a few evenings once every two years as part of EQUITANA. EQUITANA – the world's leading equestrian exhibition.
Immerse yourself in a vibrant world of innovative performances to the accompaniment of amazing sound and lighting effects.
The HOP TOP Show features leading lights from the equestrian show scene and top-class sports, performing their very latest thrilling landmark routines.
The 60m wide LED display and gantry spotlights under the roof transform the exhibition hall into an imposing amphitheatre.
Experience "LaOra" in two hours full of rousing effects, poetry and harmony between humans and animals.
There is no other equestrian gala on earth quite like the HOP TOP Show. In view of the huge demand for tickets in previous years, we recommend reserving your tickets early.
The Greek philosopher Plato once wrote that, "Time is the moving image of eternity". "LaOra", the HOP TOP SHOW for EQUITANA 2019 stopped time for two hours as leading stars of and newcomers to the equestrian show scene took to the arena with new spectacular, breathtaking performances. "LaOra" presented inspiring liberty dressage, quadrilles, dazzling acrobatics on horseback, colourful performances with ponies, and classical elements from the worlds of dressage, show-jumping and carriage-driving. The spotlight is always on the deep bond that exists between horse and human and on the subtle communication between the two that solely relies on virtually imperceptible signals.
Here are the stars for "LaOra" 2019.
Guillaume Mauvais' performances with his mighty, earth-shaking Comtois heavy horses impress thanks to their amazing partnership, the horses' will to work, their harmony and yet also their easy playfulness.
Guillaume's love of Comtois horses and his enthusiasm for horsemanship led to him founding his team, "Les Comtois en Folie", in 2001. They are based in Maîche in France, close to the Swiss border.
What connects the 30 or so members of Austria's Friesian Show Team is their love of their "black pearls".
Whether on a side saddle, in baroque dress, in front of a carriage or doing circus lessons, the outfit celebrates the beauty and versatility of the Friesian horse in performances throughout Austria and neighbouring countries at festivals, exhibitions and other events. At the HOP TOP SHOW "LaOra" we will see them deliver an imaginative quadrille together with the dance company "Dance Industry" from Austria.
Sisters Isaora and Jezabel Marques from Arles in the South of France whisked us away to the world of Andalusia.
Audiences are fascinated by the precision, perfect synchronization and top riding skills demonstrated in their perfectly choreographed performances that follow the doma vaquero tradition, including using the garrocha.
Will Rogers is originally from Australia, but now lives in Sassenberg near Warendorf. He has developed off his own method of accustomizing horses to fresh stimuli at an early age by working from the ground and using liberty work.
This way, his horses learn to master all the challenges that they will encounter in their lives as competition horses or in everyday life with people. Will's horses have such complete trust in him that they will follow him through the most unusual situations, whether under the saddle or completely at liberty at his side.
Anne Krüger-Degener runs a sheep farm and an animal school for working dogs. Over the years the seriousness for the daily work turned into fun on the challenge of really breathtaking tasks.
Involving almost the whole crew of the farm the shepherdess Anne arranges fascinating shows demonstrating the fantastic performance of sheepdogs and the incredible harmony between dog, horse and human being.
Mystical, labyrinthine, sometimes ensnared, but then managing to wriggle free after all.
Riders Gianluca Coppetta, Andrea Giovannini and the dance team SYANDA from Italy will be presenting a creative performance that centres around boundaries and freedom, proximity and flight, all against the background of a fascinating light show.
A regular and welcome visitor from France, Lorenzo has been a major star in the equestrian show scene for many years now.
It all started around 20 years ago with a few vaulting exercises on a Camargue horse in the South of France. Today, Lorenzo is one of the most famous and successful equestrian artists worldwide. Enthralling audiences with his spectacular breath-taking performances, Lorenzo stands totally relaxed on the backs of two horses without any form of bridle or halter, directing the rest of his team with the greatest of ease. It is as if they were all joined by invisible threads. So attuned are they to one another that they trust each other blindly and work as an integral unit. Such harmony and perfection reveal just how close the empathy between man and horse can be.
The NRW Pony Show Team won the KATINA Show Cup at EQUITANA Essen in 2017 with a derring-do jumping quadrille. They also featured in the EQUITANA OPEN AIR Evening Show in 2018.
In the meantime, the team comprises 16 young riders from all over North Rhine-Westphalia who regularly train at the weekend for appearances at all sorts of different events. In view of the demanding choreography of their jumping quadrilles, the riders have to be perfectly spaced and each routine finely tuned, demanding absolute concentration and lightning reactions of both riders and horses.
The current world champions demonstrated vaulting at the very highest level – a sport that demands perfect teamwork, acrobatics, creativity and elegance.
Under the direction of Patric Looser, our intrepid vaulters turn the wheel of time faster and faster in "LaOra", until they have become so fast that they manage to leap through time.
Living freedom combined with sport and excitement. The Instagram star Lisa puts a lot of emphasis on the varied training with their horses.
The Instagram star Lisa puts a lot of emphasis on the varied training with their horses.
Top-class carriage driving at the HOPTOP SHOW! 64 horses' legs burned 16 hot wheels in the arena!
Team Weinmayr & Dobrovitz brings together two of the leading families in carriage-driving and no less than four teams of four into the arena. Multiple German champion Ludwig Weinmayr, his wife Regina, József Dobrovitz, currently 7th in the FEI world-rankings, and his son József Junior, will be demonstrating just how agile and speedy good driving horses are - and how they have nerves of steel, even when making their "hot wheels" glow.
Our newsletter provides you with regular information about the HOP TOP SHOW, with special offers, competitions and the very latest news about EQUITANA - World Equestrian Sports Fair.
For over 30 years now, team EQUITANA has been putting together shows that are the most amazing multi-faceted works of art, with their mix of spectacular performances by a whole plethora of top international stars from the equestrian scene. The HOP TOP SHOW is an impressive world-class show that is only performed on four evenings once every two years as part of EQUITANA.
"EQUITANA also always means the HOP TOP SHOW. Even Olympic champions find it irresistible!"
"The HOP TOP SHOW is one of the best equestrian shows worldwide."
Equestre, Spanien
"The EQUITANA Hop Top Show is so beautiful it moves you to tears."
Reiter & Pferde in Westfalen
"HOP TOP– a name that has been synonymous with spectacular acts and unforgettable moments for many years now."
pferderevue.at, Österreich
"The Hop top Show, judged to be one of the world's best equestrian shows, has been a regular feature of EQUITANA for many years now."
"The HOP TOP SHOW has been one of the highlights of EQUITANA for many years now and is an absolute must-see for all horse lovers."
lovasok.hu, Ungarn
"The Hop Top Show is considered to be the most exceptional and most impressive horse show in the world."
Visitors' service hotline: +49 211 90191-201 Monday – Friday from 8 am to 6:30 email@example.com


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99 Locations of London
99 Locations of New York
Strange trip in NYC
Detective Minute
"Detective Minute", a live action comics. Set in a strange, fictive metropolis Magnetic City, the project was inspired by the concept of the future in the 1950s.
Death of Speech
A short story about how we communicate with each other nowadays.
Ready to Fly
Planet Madness
Inspired by detective comics, 1950s of America, Magical Realism style and Igor Stravinsky's Firebird Suite Introduction theme.


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Herd Medicine?
The nation has opted to create some form of universal health care. That is clear.
"…physicians, as an ethical duty owed to society, must practice efficient, parsimonious, and cost-effective health care." from The American College of Physicians Sixth Edition of its Physicians Ethics Manual
While this statement is reasonable and reasoned with regard to national health care, it is in clear contrast to the time-honored ethical position that 'Every life is beyond price".
Though an ambitious endeavor like the creation of national health care is a process, to be revised and restructured as required, strident debates continue. Consider these quotes:
"Traditional medical ethics, based on the doctor-patient dyad, must be reformulated to fit the new mold of the delivery of health care. . . The primary function of regulation in health care. . .is to constrain decentralized individualized decision making." From 'New Rules" by Troyen A. Brennan and Donald M. Berwick MD MPP
"…the GOD panels (Government Operatives Deliberating) – … which will determine the most cost-effective way to practice medicine, and… distribute rules down to American physicians for deciding who gets what, when and how – tell us that what's good for the herd is certainly what's good for the individual." From the DrRich blogsite January 3rd, 2012 "…
"M.D.s do not think of … patients as (a) herd (but) future health care decision-makers will." From 'Obama's Herd Health Program' by Heather McCauley DVM
herd (hûrd)
1. a) A group of cattle or other domestic animals of a single kind kept together for a specific purpose.
b) A number of wild animals of one species that remain together as a group:
2. a) A large number of people; a crowd:
b) The multitude of common people regarded as a mass:
I am not a veterinarian. Yet it is reasonable to observe that in general veterinary medical practice is found in two forms:
Herd Medicine -focused on the greatest benefit to the greatest number of animals, and the economics of health care; and 'Companion Animal Medicine'- fee for service care focused on individuals.
The term herd, when applied to human health care is disturbing to some, who see in it Huxley's Brave New World; on the other hand, some see in our current medical 'system' much of Dickens' 17th century industrial Britain. I contend that both views are flawed, and that the voting public in general, should realize that these two aspects of health care are not mutually exclusive; they are necessary in a national health program.
The table below provides side-by-side comparisons between herd and companion animal medicine. They are not altogether translatable to human health care, yet they are helpful in considering the implications of the national health care like that of the Affordable Care Act.
Herd-Vet Medicine Pet-Vet (Companion Animal) Medicine
Focus on large populations Focus on individuals
Collective outcome central Individual outcome central
Greatest benefit to greatest number Greatest benefit to the individual pet
Cost /benefit a major factor Cost limiting, but not central
Economic viability/profit critical to all Profit a consideration for 'providers'.
Herd owner pays Pet owner pays
Decisions based on outcomes , economics Decisions based on emotions and owner resources
Ethical Considerations: Ethical Considerations:
Euthanasia, cloning, etc. justified by economics. End of life and advanced tech decisions by law.
Animal Rights not a prime consideration Animal ('patient') Rights of prime importance
Special considerations & legislation Special considerations & legislation
Valuable animals treated as individuals People can be jailed longer for abusing a
Animal rights laws may affect outcomes pet than a person.
In a sense, 'Pet-vets' infringed the patent on physician fee for service medical care. (So did hospitals, phone companies, government agencies, and other entities that bill for 'services' through a lengthy and unintelligible list of charges; but that's different essay!)
Society needs now to infringe the patent on veterinary herd medicine, in a dual system to make national health care effective, practical and viable. In fact, dual- herd/individual care- systems have always existed in medicine, like the county hospital/private hospital dichotomy that survived until half a century ago,which was imperfect but functional. In fact, every nation that has a sustainable national health program allows or employs both types of health care: one focused on large numbers, outcomes, and economics, and the other focused on the rights of individuals. Both rest on ethical and practical considerations, which address different aspects of national health care. The ACA is such a duality, theoretically and functionally imperfect but subject to revision and improvement.
Every nation able to offer ongoing universal national health care to all its citizens has found it necessary to develop some sort of 'two tier system'. Can we do so now in our democracy? We'll see. I believe we can, and will. Any objective look at the life of people on this earth makes clear that over the centuries, people have lived progressively longer, healthier, more comfortable lives. I predict we will develop a functional, ethical, and affordable national health care consistent with our own culture and history. It will reflect one key word in the Affordable Care Act that implicates features of herd medicine; that word is: Affordable.


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- American Society of Plant Biologists
Shoot meristems harbor stem cells that provide key growing points in plants, maintaining themselves and generating all above-ground tissues. Cell-to-cell signaling networks maintain this population, but how are meristem and organ identities controlled? TERMINAL FLOWER1 (TFL1) controls shoot meristem identity throughout the plant life cycle, affecting the number and identity of all above-ground organs generated; tfl1 mutant shoot meristems make fewer leaves, shoots, and flowers and change identity to flowers. We find that TFL1 mRNA is broadly distributed in young axillary shoot meristems but later becomes limited to central regions, yet affects cell fates at a distance. How is this achieved? We reveal that the TFL1 protein is a mobile signal that becomes evenly distributed across the meristem. TFL1 does not enter cells arising from the flanks of the meristem, thus allowing primordia to establish their identity. Surprisingly, TFL1 movement does not appear to occur in mature shoots of leafy (lfy) mutants, which eventually stop proliferating and convert to carpel/floral-like structures. We propose that signals from LFY in floral meristems may feed back to promote TFL1 protein movement in the shoot meristem. This novel feedback signaling mechanism would ensure that shoot meristem identity is maintained and the appropriate inflorescence architecture develops.
Growth and development are coordinated through diverse signaling networks. Cell-to-cell signals communicate positional information, induce cellular and tissue identities, and ultimately lead to complex developmental patterns of differentiation. Animals and plants have evolved diverse modes of signaling to establish cell and organ identities. Diffusible protein morphogens, secreted peptides, and small hormones promote differential tissue development and growth (Crickmore and Mann, 2006; Xu et al., 2006). An important aim is to identify the range of such signals, how they are regulated, and how they coordinately act upon their targets.
Plant meristems integrate a diverse array of signals. Meristems are key growth points of plants, adding new organs appropriate to each developmental phase. They are composed of central cells, which maintain the population, and peripheral cells that are programmed to form the different types of organ primordia on the flanks. Various signaling networks maintain this pattern and regulate the size and form of the meristem (Clark et al., 1993; Long et al., 1996; Mayer et al., 1998; Fletcher et al., 1999; Wu et al., 2005). Overlaying this pattern are gene activities and signals that determine where and when primordia are made, what will be their identity, and, thus, what form of overall plant architecture will arise.
Shoot meristems give rise to above-ground organs of the plant. Organ identity is tightly controlled, reflecting the ability of the shoot meristem to precisely interpret a diverse array of signals that affect the growth and identity of primordia. In Arabidopsis thaliana, upon germination, the shoot meristem passes though a vegetative phase, making leaf primordia in a compact rosette, with the first leaves showing juvenile traits and later leaves adult traits (Poethig, 2003; Baurle and Dean, 2006). Later, in response to developmental and environmental signals, the shoot meristem changes its growth pattern: the stem elongates, makes modified leaves, and then switches to making flowers to give the inflorescence. Secondary shoots (branches) arise from the axils of leaves and have a similar growth pattern to the main shoot. However, which axillary meristems grow out, when this occurs, and to which degree are controlled by many axillary factors and branching signals (Beveridge, 2006). Shoot meristems therefore integrate many different signals, but what are these and how do they act?
Specific signals act upon the different growth phases. Various signaling pathways define the vegetative phase, its identity, and its length. Many of these signals are components of RNA interference pathways, affecting the expression of genes that influence the identity of leaves (Hunter et al., 2006). The number of leaves is also tightly controlled, with many pathways promoting a longer vegetative phase and others shortening it (Baurle and Dean, 2006; Corbesier and Coupland, 2006). The switch to flowering involves an integration of these signaling pathways, resulting in the upregulation of the flowering genes. Activation of key integrators, FLOWERING LOCUS T (FT) and SUPPRESSOR OF CONSTANS1, results in a switch in shoot meristem identity and the induction of flowering (Kardailsky et al., 1999; Kobayashi et al., 1999; Samach et al., 2000; Abe et al., 2005; Wigge et al., 2005). FT is expressed in leaves, but its mRNA appears to be a mobile signal that moves to activate APETALA1 (AP1) and LEAFY (LFY) on the flanks of the shoot meristem where they promote floral meristem development (Mandel et al., 1992; Weigel et al., 1992; Hempel et al., 1997; Takada and Goto, 2003; An et al., 2004; Huang et al., 2005).
TERMINAL FLOWER1 (TFL1) is a key signaling protein that controls shoot meristem identity during all of the plant's life cycle. TFL1 acts throughout development to influence each phase of growth; tfl1 mutants have a shorter vegetative phase, making less leaves, branches, and flowers than the wild type (Shannon and Meeks-Wagner, 1991; Alvarez et al., 1992; Schultz and Haughn, 1993). Also, the shoot meristem converts to a terminal flower in tfl1 mutants, unlike the wild type, where it grows indefinitely, never differentiating and only finally senescing. TFL1 is a temporal and spatial repressor of flowering genes and acts opposite to FT, which promotes flowering genes. Interestingly, these two proteins are also homologs (Kardailsky et al., 1999; Kobayashi et al., 1999; Bradley et al., 1997; Ohshima et al., 1997).
Expression of TFL1 is restricted to the inner cells of mature shoot meristems. TFL1 mRNA is very low during the vegetative phase, but its levels are strongly upregulated at the switch to flowering (Simon et al., 1996; Bradley et al., 1997; Ratcliffe et al., 1999). TFL1 expression does not extend into the outer cell layers, the epidermis of the shoot meristem, or into primordia, yet it controls the identity of all of these cells. First, TFL1 determines the identity of primordia made. Second, in tfl1 mutants, all cells, even the epidermis, ectopically express flowering genes (Weigel et al., 1992; Bowman et al., 1993; Gustafson-Brown et al., 1994; Bradley et al., 1997; Liljegren et al., 1999). Thus, TFL1 acts beyond its expression domain, but how does TFL1 achieve this? Where does the protein act to repress flowering genes?
We find that the TFL1 protein acts as a mobile signal to coordinate shoot meristem identity. TFL1 moves from inner cells to outer cells but is restricted from lateral primordia. Its pattern does not show a gradient, suggesting that it may be actively transported to coordinate shoot meristem identity and reach its targets. Strikingly, we found that LFY may control TFL1 movement, suggesting that floral meristems signal back to influence shoot development. This feedback may provide a general mechanism to keep floral meristem identity genes expressed on the flanks and thus generate an indeterminate growing inflorescence bearing many flowers. As lfy mutants terminate in floral structures, our observations suggest that TFL1 movement may also be necessary. Finally, we demonstrate that TFL1 expression is regulated differently in the mature shoot meristem compared with young axillary meristems, both in terms of RNA expression and protein patterns.
TFL1 Is a Cytoplasmic, Unmodified 20-kD Protein of Shoot Meristems
To analyze the amount, size, and distribution of the TFL1 protein, we raised antibodies against TFL1. This anti-TFL1 sera detected Escherichia coli–expressed TFL1, but not its Antirrhinum majus functional homolog CENTRORADIALIS (CEN), which is 69% identical (Figure 1A ) (Bradley et al., 1996; Cremer et al., 2001). A commercial anti-TFL1 serum gave only a weak signal.
We tested whether our anti-TFL1 sera detected TFL1 in plant extracts. Initial immunoblots with extracts of 35S-TFL1 control plants (Ratcliffe et al., 1998) showed that only two bands were detected: one of the predicted size of 20 kD and one of 30 kD. This 30-kD band was nonspecific, as it was detected by all antisera and controls and was found in tfl1 null mutant plants and only in leaf tissue. Therefore, to characterize the 20-kD band in more detail, we analyzed protein extracts of young wild-type Arabidopsis and two tfl1 mutant alleles (tfl1-1 and tfl1-18). The tfl1-18 mutant allele is a newly described insertion allele (available in the GABI-Kat collection; Y. Hanzawa and D. Bradley, unpublished data) that should delete most of the protein, and no significant mRNA was detected in long days (LD) by in situ analysis. Therefore, tfl1-18 was probably a very strong or null allele, and TFL1 protein was expected to be absent or much reduced in tfl1-18 compared with the wild type.
High-resolution gels showed that the initial 20 kD band detected could be resolved into two proteins of 20 and 17 kD. The 20-kD protein was present in E. coli TFL1 expression extracts, but wild-type plant extracts only had the 17-kD protein, and this was also present in tfl1-1 and tfl1-18 mutants, suggesting that it was a nonspecific cross-reaction of the anti-TFL1 sera (Figure1B). The 17-kD protein was enriched in extracts of young flowers and was not found in vegetative seedlings (e.g., Figure 1B, 35S-TFL1). The 20-kD TFL1 protein was detected in transgenic lines overexpressing TFL1.
To detect TFL1 in extracts, we used plant material enriched in meristems expressing TFL1, namely, the ap1 cal double mutant. This produces large cauliflower-like apices of proliferating meristems that have high levels of functional TFL1 mRNA (Bowman et al., 1993; Ratcliffe et al., 1999; Ferrandiz et al., 2000). The 20-kD TFL1 protein was clearly detected in these extracts, along with the 17-kD protein (Figure 1B). This reflected the dual nature of ap1 cal meristems being part inflorescence and part floral. Therefore, our anti-TFL1 sera could successfully detect the 20-kD TFL1 protein, which was stable in tissue extracts. TFL1 was apparently of the predicted size; repeated measurements gave a size of 19.5 ± 1.9 kD compared with the predicted 20.2 kD.
We checked whether there were any modifications in TFL1 not affecting apparent gel size. We affinity-purified TFL1 from transgenic plants expressing a constitutive TAPtagTFL1 fusion protein and subjected TFL1 to peptide fingerprinting/mass spectrometry (matrix-assisted laser-desorption ionization time of flight) (see Supplemental Figures 1 and 2 online). Repeat analyses provided extensive coverage (86%) of this version of TFL1 and spanned from V8 to R170. This revealed no novel peptide fragments, suggesting that no significant covalent modifications or processing of TFL1 occurs.
To determine where TFL1 was located inside cells, we fractionated native TFL1 from ap1 cal tissues. The final steps involved ultracentrifugation to separate the total mixture into soluble components and pelleted membranes/membrane vesicles (Figure 1C). This showed that TFL1 was a soluble 20-kD protein, while the nonspecific 17-kD protein was membrane-bound and thus separable from TFL1.
As the TFL1 homolog FT is found in the nucleus (from FT–green fluorescent protein [GFP] studies) and FT binds bZIP transcription factors, we asked whether TFL1 was also nuclear (Pnueli et al., 2001; Abe et al., 2005; Wigge et al., 2005). Fractionation of ap1 cal tissues yielded a nuclear-enriched fraction (as confirmed by histone detection) and a supernatant fraction, containing the cytosol and low-density membranes and organelles (Figure 1D). TFL1 was only found in the supernatant, suggesting that the majority of TFL1 is not in the nucleus (Figure 1D). To resolve exactly where TFL1 protein accumulated, in which specific tissues, and which domains of the inflorescence, we probed sections of plant tissue.
TFL1 Moves beyond Its mRNA Expression Domain in Inflorescence Meristems
Wild-type and control tfl1 mutant plants were grown in LDs and harvested at various time points for both protein (immunolocalization) and mRNA in situ analyses.
In wild-type plants, TFL1 mRNA was very low during the vegetative phase but was strongly upregulated in the shoot meristem when it switched to an inflorescence identity at about day 12 (Figure 2A ). TFL1 mRNA was strong in both the main and lateral shoot meristems at this stage and at later time points (Figure 2A, arrow). No tfl1 mRNA was detected in the tfl1-18 mutant plants at any time point (Figure 2B). Unlike tfl1-18 mutant plants, both tfl1-1 and tfl1-13 had tfl1 mRNA (Figures 2C and 2D). In these cases, their tfl1 mRNA was absent from the main shoot by day 12, as it had already converted into a floral meristem (as shown by AP1 expression). However, tfl1 mRNA was present in secondary shoots arising from the axils of rosette leaves, and this acted as a potential marker for inflorescence shoot meristem identity (Figure 2C). Therefore, as previously reported, TFL1 and tfl1-1 mRNA accumulated in the central, internal regions of inflorescence meristems. However, tfl1-18 mutant plants had no detectable tfl1 mRNA.
In wild-type Arabidopsis, Columbia (Col) or Landsberg erecta (Ler) backgrounds, TFL1 protein was not observed at early time points of 8 or 10 LD. Strong background, nonspecific signal was often observed but was confined to the pith region (the central body) of the rosette stem and more variably in older stems or leaves (Figure 3A ). This was confirmed as background by probing tfl1-18 mutants that had no TFL1 mRNA or protein. In the wild type, the TFL1 protein was detected only clearly at the switch to inflorescence identity at about day 12 onwards; TFL1 was then strong and found throughout upper axillary shoot meristems (Figures 3A to 3C). TFL1 protein was detected in the main shoot meristem only later, after it was seen in the secondary axillary meristems, where it again appeared throughout the meristem (Figures 3B and 3C). Therefore, independent of their position, wild-type inflorescences had TFL1 protein distributed throughout their entire meristems, unlike its mRNA, which was restricted to the central region.
Probing late wild-type time points showed that TFL1 protein signal was variable but present up to 28 LDs. TFL1 protein did not appear in floral meristems, despite being throughout the inflorescence meristem dome (Figures 3B and 3C). The floral meristems did have some weak background staining, but this was also found in tfl1-18 mutants and the wild type probed with preimmune sera (Figure 3D; see Supplemental Figure 3 online).
TFL1 protein did not appear to extend far below the mRNA expression domain (Figure 3C). This suggested that TFL1 moved toward the apex and to the outer layers (epidermis) of the meristem. If TFL1 did move inward, it did not accumulate, either because it was degraded or it moved rapidly away to distant tissues. We probed 35S-TFL1 plants and showed that TFL1 protein was present and stable in all tissues observed (see Supplemental Figure 3 online). This indicated that TFL1 was not rapidly degraded if present outside the inflorescence meristem.
We analyzed the effect of tfl1 mutations on protein levels and its distribution. The tfl1-1 and tfl1-13 alleles were chosen because they derive from single amino acid substitutions that map to two different sites of the TFL1 protein crystal structure, suggesting that they might affect two different domains of TFL1 function (Ahn et al., 2006). No tfl1-1 or tfl1-13 mutant protein was observed at any time point in mutant plants. This suggested that tfl1 mutant protein was perhaps not made or unstable. However, as tfl1 mutants are more advanced in their growth phase compared with the wild type and shoots converted to flowers, it was difficult to directly compare meristem stages. Therefore, to overcome these problems, we looked at short day (SD) grown plants, where tfl1 mutants have extensive vegetative and inflorescence phases.
TFL1 Protein Also Moves in Vegetative Meristems, but tfl1 Mutant Proteins Are Absent
Arabidopsis plants grown in SD have an extensive vegetative phase, during which time the main shoot meristem generates many leaves with axillary shoot meristems. These axillary meristems develop in a general gradient, being more developed in the older leaves (these axillary meristems make a few leaf primordia) and least developed nearer the main shoot meristem (the apex) (Grbic and Bleeker, 2000; Long and Barton, 2000). These meristems remain small and vegetative, until the main shoot has switched to inflorescence identity, at which point many grow out and switch to inflorescence identity. This switch can be simply controlled by shifting plants to LD.
We grew plants for 30 SDs before shifting them to LD to induce flowering and harvest at various time points for mRNA and protein in situ. This showed that TFL1 mRNA was strong in wild-type vegetative meristems before LD induction (Figure 4A ). However, this expression was only observed in axillary meristems, not the main shoot meristem. All axillary shoot meristems observed in sections appeared to have TFL1 mRNA, from young to old (Figure 4A). At the very youngest stages, TFL1 mRNA appeared to occupy all cell layers, but at later meristem stages, this resolved into TFL1 being restricted and only occupying the inner cells, at the base of the meristem, next to the leaf. After LD induction, TFL1 mRNA increased first in the upper developing secondary shoots of the elongating main inflorescence. TFL1 was still low in the main shoot meristem between +1 to +3 LD stages but generally increased. Eventually, by approximately +4 LD, TFL1 mRNA was strong in the main shoot in the central region, at the time when floral meristems developed as the uppermost lateral meristems, as shown by AP1 expression (Figure 4B; see Supplemental Figure 4 online) This suggested that the main shoot only expressed TFL1 strongly when it was in the inflorescence stage, but axillary meristems expressed TFL1 strongly in the vegetative phase.
In sections of tfl1-18 mutant plants, weak tfl1-18 mRNA was very occasionally observed in a few meristems (Figure 4C). After LD induction, no tfl1-18 mRNA was seen (Figure 4D). This was not due to conversion of all meristems to flowers directly, as both the main shoot meristem and many lateral meristems were vegetative or inflorescences, as shown by lack of AP1 expression (Figure 4D, arrows; see Supplemental Figure 4 online).
The tfl1-1 and tfl1-13 mutant plants had strong levels of tfl1 mRNA in SD, similar to the wild type (Figure 4E). After LD induction, both point mutation alleles again appeared like the wild type, with strong tfl1 mRNA accumulating in secondary meristems and later in the main shoot meristem (Figure 4F). Also, in some sections, the position of tfl1 mutant mRNA appeared altered compared with the wild type, occupying nearly the whole meristem. AP1 expression analyses also confirmed that many shoots remained as vegetative or inflorescence meristems throughout these time points for both tfl1 alleles, including the main shoot meristem (see Supplemental Figure 4C online).
Protein in situ showed high levels of TFL1 protein in axillary meristems in SD where TFL1 mRNA was also strong and not the main shoot meristem where mRNA was low (Figure 5A ). TFL1 protein was present throughout these SD vegetative meristems, in contrast with TFL1 mRNA, which was generally restricted to central/basal cells. After LD induction, the TFL1 protein appeared strong in the axillary shoot meristems of the elongating main shoot (Figures 5B and 5C). TFL1 protein was consistently not clearly detected in the main shoot meristem till about day 6 (Figure 5D). Again, the pattern of TFL1 protein did not match the mRNA pattern; the TFL1 protein was distributed throughout the meristems, vegetative or inflorescence, and so extended beyond the mRNA domain.
No tfl1-18 protein could be detected in mutant plants at any stage in SD or after LD induction, despite the presence of vegetative shoot meristems and inflorescences (Figures 5G and 5H). Also, no tfl1 mutant protein was seen in either tfl1-1 or tfl1-13 mutant plants at any time point, in any stage of meristem development (Figure 5I). Thus, despite significant levels of mRNA for these two different point mutation alleles, their proteins did not accumulate.
TFL1 protein had a different distribution compared with the wild-type mRNA. This difference might be explained by one possible artifact, namely, different fixation/embedding methods for protein versus mRNA. We tested this by probing consecutive sections of the same material for both protein and mRNA distribution. This showed that indeed the protein had a broader expression domain compared with the mRNA (Figures 5E and 5F). Thus, protein movement is most likely to account for TFL1 protein distribution inside shoot meristems. No obvious signal peptide could be found in the TFL1 protein sequence that suggested it might be secreted, moved, and taken up by target cells. This suggests that movement occurs cell-to-cell through plasmodesmata.
Floral Meristem Identity Genes Regulate the TFL1 Protein Pattern
TFL1 is antagonistic to the floral meristem identity genes. TFL1 mRNA is found in a separate domain from the floral meristem identity genes LFY, AP1, and CAL. This also appears true for the TFL1 protein in wild-type plants because although TFL1 protein moves beyond its mRNA domain, it does not move into floral meristems. To see how this pattern of protein movement might be regulated, we looked at the TFL1 protein in lfy and ap1 cal mutants.
In lfy mutants, shoots replace flowers and TFL1 mRNA is present in a similar pattern to wild-type shoot meristems, being strong in the central region (Huala and Sussex, 1992; Weigel et al., 1992; Liljegren et al., 1999; Ratcliffe et al., 1999). The shoot meristems eventually convert to terminal carpel/flower-like structures and TFL1 mRNA decreases. Samples from independent lfy alleles were harvested just as they began to elongate (bolt) at ∼23 LD. TFL1 protein was found to be strong in young, axillary meristems, similar to the wild type (Figure 6A , inset). Thus, the distribution and stability of TFL1 protein in young axillary shoot meristems was not affected in lfy mutants. However, in the elongated main, secondary, or tertiary shoots, the TFL1 protein was often weak or absent or in a novel pattern of a restricted domain in the center of the meristem (Figures 6A to 6C). Control wild-type plant segregants at the same time point showed TFL1 protein distribution throughout all shoot meristems, indicating that this restriction of TFL1 movement in lfy plants did not depend on the age of the meristem (Figure 6D).
This restriction of TFL1 protein to the center of the meristem was seen in more than four independent experiments, for two different alleles (lfy-6 [Ler] and lfy-14 [Col]), and for multiple meristems from any sample. However, it was only seen in 23 or 27 d samples, being absent from the four 36 d samples analyzed. This localized pattern was not seen in wild-type segregants (or other wild types analyzed in our experiments), and a possibly similar signal was only rarely found in tfl1 mutants grown in SD. Such a signal could arise if the antisera cross-reacted with pith/vascular tissue developing below the apex. However, such artifacts might be expected to be evident also at later time points or in the wild type, but this was not observed. Rather, given the high frequency of this novel TFL1 pattern in lfy mutants, it appears to show an inability for TFL1 protein to move in lfy shoots.
AP1 and CAL are also required restriction of TFL1 mRNA to the shoot. AP1 and CAL are required for floral meristem identity and are themselves activated by LFY. In ap1 cal mutants, TFL1 mRNA is present throughout the reiterated meristems of the cauliflower phenotype, in the same meristems as LFY, though the youngest meristems lack LFY (Weigel et al., 1992; Liljegren et al., 1999; Ratcliffe et al., 1999). Therefore, we looked at TFL1 protein distribution in ap1 cal. At day 12, ap1 cal mutant shoots looked similar to the wild type, and TFL1 protein was present at high levels in axillary shoot meristems similar to the wild type, while the main shoot meristem had moderate levels of TFL1 protein (Figure 6E). By day 14, the morphology of the main shoot had changed compared with the wild type, and the meristems arising from the flanks of the ap1 cal main shoot also had high levels of TFL1 protein (Figure 6F). By contrast, these meristems were floral in the wild type and had no TFL1 protein. In ap1 cal mutant plants, the lateral meristems proliferated to form more meristems on each of their flanks. Therefore, by days 16 or 20, a proliferating series of meristems was generated and these all had TFL1 protein (Figures 6G and 6H). In all meristems, at all time points, TFL1 protein appeared throughout the meristem, though levels were variable. Because LFY is not upregulated in the youngest stages of ap1 cal meristems, these data again show that early TFL1 protein movement is independent of all these floral genes. Thus, the ap1 cal mutations affected meristem identity but did not affect TFL1 movement.
We analyzed the biochemistry and tissue patterns of TFL1 to see how this signaling protein controls shoot meristem identity. Intriguingly, TFL1 mRNA is restricted to the inner cells of the meristem, but TFL1 protein is small and mobile and may thus coordinate meristem functions by moving to interact with its targets. Our data suggest a new model for how floral and shoot meristems may establish a feedback loop of expression and protein movement to control inflorescence architecture.
TFL1 Is a Small Cytoplasmic Protein of Shoot Meristems
The TFL1 protein accumulates in shoot meristems and is mostly cytoplasmic, not being readily detected in nuclear extracts. Consistent with this pattern, in situ studies showed that the TFL1 protein is found throughout the cell and not in a discrete nuclear-localized pattern similar to transcription factors, such as LFY (Levin and Meyerowitz, 1995; Parcy et al., 1998; Sessions et al., 2000). TFL1 affinity purification for peptide analysis indicated that extractable TFL1 is largely unmodified in plants. Therefore, to signal and regulate meristem identity genes, TFL1 may usually reside in the cytoplasm, where it may transiently associate with its interactors. If TFL1 moves to the membrane, nucleus, or any other organelle, then this may affect only a small amount of the TFL1 pool.
Many TFL1 plant homologs are known, and their functions, mRNA expression patterns, three-dimensional structures, and interactions are all very well described, yet only a few aspects of protein action have been described (Banfield and Brady, 2000; Pnueli et al., 2001; Abe et al., 2005; Hanzawa et al., 2005; Wigge et al., 2005; Ahn et al., 2006). Use of fusions to marker proteins showed that one protein homolog, FT, resides in both the cytoplasm and nucleus and interacts with a bZIP transcription factor to regulate its flowering target, AP1 (Abe et al., 2005; Wigge et al., 2005). Similarly, TFL1 can interact with bZIPs. Animal homologs have been shown to weakly associate with membranes but appear to be mostly cytoplasmic, interacting with a range of proteins, such as kinases and proteases (Hengst et al., 2001; Yeung et al., 2001). Therefore, one general model for this family of proteins may be that they reside in the cytoplasm and interact with potentially many partners, and this regulates their distribution and action throughout the cell. Since TFL1 moves from cell to cell, some of these interactions may guide TFL1 on its journey.
TFL1 Moves from Inner to Outer Meristem Cells
We have addressed a key question on how TFL1 regulates shoot meristems. Gene expression data show that TFL1 mRNA is restricted to the inner cells of the shoot meristem, yet TFL1 must prevent flowering genes from expressing in all layers and cells (Weigel et al., 1992; Bowman et al., 1993; Gustafson-Brown et al., 1994; Bradley et al., 1997; Liljegren et al., 1999). To achieve this, the TFL1 protein apparently moves beyond its gene expression domain. TFL1 moves from inner cells to outer cells. Therefore, TFL1 itself is likely a signal that relays and coordinates shoot identity. Subsequently, TFL1 may find its interactors to act upon its targets to regulate cell identity (Abe et al., 2005; Wigge et al., 2005). A future goal will be to find the distribution of these interactors and how they overlap with TFL1.
TFL1 movement may allow it to act, not only in central cells, but also in peripheral cells of the shoot meristem that proliferate to form floral meristems. LFY is expressed in this peripheral region, in anlagen of future primordia and floral meristems (Weigel et al., 1992; Hempel et al., 1997). Thus, TFL1 protein may overlap with LFY mRNA at the periphery, though it is not clear if LFY protein also accumulates to significant levels in these cells (Levin and Meyerowitz, 1995; Parcy et al., 1998). FD is also expressed in anlagen and is involved in AP1 activation (Abe et al., 2005; Wigge et al., 2005). By contrast, AP1 mRNA is not upregulated until cells emerge from the shoot meristem to give rise to a stage 1 floral meristem (Mandel et al., 1992; Hempel et al., 1997). In tfl1 mutants, AP1 becomes expressed in the shoot meristem. Thus, TFL1 movement may be critically involved in ensuring that LFY and AP1 are not expressed in the outer cells of shoot meristems, as even their epidermal expression is sufficient to change shoot identity (Sessions et al., 2000).
Other proteins can move in the shoot meristem. KN1 moves from inner to outer cells of the epidermis (Jackson et al., 1994). CLV3-GFP can move from outer cells to inner cells, to a limited extent (Rojo et al., 2002; Lenhard and Laux, 2003). Transgenic methods involving chimeras or cell-specific promoters show that LFY or GFP can move within shoot or floral meristems (Sessions et al., 2000; Wu et al., 2003; Kim et al., 2005). By contrast, some factors have more restricted movement, such as DEF in floral meristems, that indicate regulation of movement (Perbal et al., 1996). SHR moves within root tissues in functionally important and very precise patterns (Nakajima et al., 2001). Thus, TFL1 is not unique in moving but appears to have its own specific pattern, regulation, and function in generating a particular architecture.
Mechanism of TFL1 Movement
Two general modes of protein movement in meristems have been suggested. The first requires export of the protein from the expressing cell, extracellular diffusion to the target cells, and interaction with receptors. Examples, such as CLV3, reveal signal sequences that target such export (Fletcher et al., 1999; Rojo et al., 2002; Lenhard and Laux, 2003). TFL1, however, has no clear signal sequence.
The second mode involves proteins moving through plasmodesmata that directly connect cells (Kim and Zambryski, 2005; Lough and Lucas, 2006). This movement may be passive by simple diffusion or controlled and targeted. Studies suggest that LFY moves passively, a default situation of not being retained (Sessions et al., 2000; Wu et al., 2003). Similarly, the small protein GFP diffuses. Both show a gradient in protein levels across the tissue from their source. By contrast, KN1 has specific sequences that are sufficient and necessary for movement (Kim et al., 2005). Similarly, viruses make specific proteins to interact with plasmodesmata to change size of their pores and promote virus movement (Lough and Lucas, 2006). Such modes result in a more even distribution across tissues. The TFL1 pattern appears to be quite uniform across the shoot meristem, suggesting it is also actively transported, but this is not proof. If specificity is involved in TFL1 transport, protein structure motifs might be found through functional bioassays (Kim et al., 2005). However, it could still be that size and TFL1 properties (small, stable, and cytoplasmic) might allow it to passively diffuse and accumulate to equivalent levels across many cell distances in a tissue. It could possibly move in both directions to reach an even distribution at equilibrium. It may prove difficult to address whether size limits TFL1 movement, as our initial fusions of TFL1 to other proteins showed that activity was lost if domains as small as 20 kD were fused to either end (our unpublished data).
Our data suggest that TFL1 is readily moving at all stages of development and thoughout the shoot meristems. One key aspect for Arabidopsis and TFL1 may be limiting its movement so that it does not enter floral meristems. Such restriction of movement between floral and shoot meristems has been described for different proteins and markers (Gisel et al., 1999; Sessions et al., 2000; Wu et al., 2003).
TFL1 Is Regulated Differently in the Main versus Axillary Meristems
TFL1 mRNA is already strongly expressed in axillary meristems in SD during vegetative growth but is weak or absent from the main shoot. Why is TFL1 strong in axillary meristems when not induced to flower? Perhaps axillary meristems need stronger protection from florigen signals, as they are closely associated with leaves, the source of florigens, while the main shoot is perhaps separate and more isolated (Grbic and Bleeker, 2000; Long and Barton, 2000). LFY itself promotes flowering and is expressed weakly in leaves (Weigel and Nilsson, 1995; Blazquez et al., 1997). High levels of initial TFL1 protein may therefore be needed to inhibit LFY effects. Interestingly, tfl1 mutants make extra axillary meristems compared with wild-type plants (Grbic and Bleeker, 2000). Perhaps TFL1 plays additional roles compared with the main shoot and this high level of protein is needed for this function. Lower levels of TFL1 in the mature shoot may be sufficient to prevent florigen action, as the mature shoot may be less competent to respond to such signals.
Previously it was suggested that high levels of TFL1 mRNA could be a marker for a switch to bolting and inflorescence stages (Bradley et al., 1997; Ratcliffe et al., 1999). This now appears only true for the main shoot meristem, not axillary meristems. In SD, axillary meristems have high TFL1 levels, yet are not flowering. Could they be at the stage of induced to bolt but are simply not growing out? Or are they truly in a nonbolting, vegetative rosette-like state? Elegant and detailed analyses of traits have been extensively made on the main shoot to define its phases (Telfer et al., 1997; Poethig, 2003). However, less is known about the phase changes in axillary meristems in SD despite detailed analysis of their early development. It may be possible to find markers, perhaps STP, that are in vegetative, but not inflorescence meristems, and so follow the changes in axillary meristem identity (Wu et al., 2005).
TFL1 mRNA was present through all cells of young axillary meristems. Only as these meristems developed did TFL1 mRNA become restricted to inner cells like the main shoot meristem. How is early and late expression controlled? Many factors play specific roles in control of axillary meristem development. Genes expressed before TFL1 regulate axillary meristem initiation and growth and are specifically expressed there, while hormone targets influence axillary meristem growth (Schmitz and Theres, 2005; Beveridge, 2006; Keller et al., 2006; Muller et al., 2006). These are candidates for early regulators of TFL1 expression or protein levels in axillary meristems. Later, the TFL1 pattern is limited to central cells similar to the pattern of WUS or CLV1. How do these or regulators of this pattern affect TFL1 mRNA? Dissection of the TFL1 promoter will give insights as to how these and flowering signals are integrated to control shoot meristem identity.
Spatial Model for TFL1 Action
Our studies suggest a model for how spatial regulation of TFL1 is linked to the control of plant architecture (Figure 7 ). In young axillary meristems, TFL1 mRNA is first seen in all meristem cells and suggests early regulation of TFL1 expression by axillary factors. This pattern changes as the meristem develops, with the most apical cells no longer expressing TFL1. However, TFL1 protein moves into these cells, and this pattern further resolves into that seen in mature shoots.
In a mature inflorescence shoot, TFL1 mRNA is restricted to cells in the center of the meristem, in a domain similar to WUS or CLV1 (Castellano and Sablowski, 2005; Williams and Fletcher, 2005). TFL1 protein moves outward to the epidermal tissues but does not appear to enter primordia, suggesting barriers to movement. This barrier is necessary, as expression of TFL1 in primordia may change their identity. We propose that TFL1 movement is required to inhibit LFY and AP1 expression in the inflorescence meristem because expression of these genes, even when restricted to the shoot epidermis, is sufficient to change meristem identity (Sessions et al., 2000). TFL1 restricts AP1 expression to developing floral meristems outside of the shoot meristem. TFL1 cannot prevent LFY expression in some cells, in future primordia on the periphery of the meristem (anlagen), suggesting that activating factors are expressed in this region and these are independent of TFL1. It is not clear if LFY protein accumulates in anlagen, but only in floral meristems, where it also moves (Levin and Meyerowitz, 1995; Parcy et al., 1998). In young axillary meristems of lfy mutants, TFL1 was found throughout the meristem, showing that initial movement was independent of LFY. However, TFL1 protein was limited to central regions in mature lfy mutant shoots. This might reflect a change in protein stability in outer cells, but TFL1 appears stable in young lfy mutants and in extracts from diverse genotypes. Rather, it suggests that movement of TFL1 in the inflorescence meristem is regulated by LFY. In lfy mutants, TFL1 becomes restricted to central cells and the shoot meristem changes identity, differentiating into carpel/floral-like structures. We therefore propose that a LFY-TFL1 feedback loop may operate; LFY promotes TFL1 movement (to overcome its limited expression domain), and this ensures that LFY and AP1 are limited to lateral meristems, while shoot meristems proliferate. We expect these interactions to be indirect, as LFY and TFL1 proteins are apparently in separate domains and no reports of molecular association have been reported.
Plant Materials and Growth Conditions
We used wild-type Arabidopsis thaliana plants, ecotype Col or Ler, mutant lines tfl1-1, tfl1-13 (Shannon and Meeks-Wagner, 1991), ap1-1 cal-1 (Bowman et al., 1993), lfy-6, and lfy-14 (Huala and Sussex, 1992; Weigel et al., 1992), and transgenic line 35S-TFL1 (Ratcliffe et al., 1998). The tfl1-18 mutant is a T-DNA insertion allele (GABI-Kat ID 228A07; Rosso et al., 2003). Plants were grown as described (Ratcliffe et al., 1998). For LD experiments, plants were grown under a 16-h-light/8-h-dark photoperiod in a controlled environment at a temperature of 20 to 22°C. Light was cool light fluorescent tubes at an intensity of 90 to 120 μmol m−2 s−1.
For the SD to LD shift experiments, plants were grown in SDs (8 h light/16 h dark) in a controlled environment at 20 to 22°C, with a light fluency of 120 to 170 μmol m−2 s−1. Plants were transferred to a LD cabinet and grown under 90 μmol m−2 s−1 white cool white lamps at a temperature of 20 to 22°C. Five independent shift experiments were performed, with transfers to LD after a range of 20 to 30 SD.
Antisera and Recombinant Proteins
Anti-TFL1 sera were raised in rabbits by repeated injections of 100 μg of His-TFL1 protein. Recombinant proteins were produced in BL21 (DE3) pLysS Escherichia coli cells (Novagen). His-TFL1 was found in inclusion bodies but could be solubilized and purified with a nickel column kit according to the manufacturer's instructions (Novagen). Untagged recombinant TFL1 protein was produced from a CHITIN BINDING DOMAIN-INTEIN-TFL1 fusion (CBD-INTEIN-TFL1) via intein-mediated cleavage according to the manufacturer's instructions (New England Biolabs). CEN protein was purified as previously detailed (Banfield and Brady, 2000). Anti-CEN sera were raised in rabbits (S. Rothstein, R. Stahl, and D. Bradley, unpublished data).
For protein in situ analysis, anti-TFL1 sera (final bleed) were affinity-purified against a glutathione S-transferase-TFL1 (GST-TFL1) column. Glutathione affinity-purified GST-TFL1 (150 μg) was cross-linked to a Thiogel (Severn Biotech). Sera (300 μL) were applied to the column and treated according to instructions, except that column washes involved PBS containing 0.1% (v/v) Tween 20. High-affinity antibodies were eluted, identified by immunoblot, pooled, dialyzed against PBS, and concentrated to ∼400 μg/mL. Affinity-purified antibodies were mixed with an equal volume of 10% (w/v) BSA in PBS and frozen.
Plasmid Construction
To produce different recombinant TFL1-tag proteins, the TFL1 open reading frame (ORF) was amplified from the cDNA with different sets of oligonucleotides and cloned into pGEM-T vector (Stratagene). For poly-histidine-TFL1 (His-TFL1), the TFL1 ORF was amplified from the cDNA with oligos s169 (5′-CATATGGAGAATATGGGAACTAGAGT-3′) and s170 (5′-CTGCAGCTAGCGTTTGCGTGCAGCGGT-3′) and cloned into pGEM-T to give plasmid pLC3. pLC3 was digested with NdeI and the TFL1 insert isolated and cloned into the NdeI site of pET-19b (Novagen) to give plasmid pLC7.
For GST-TFL1, the TFL1 ORF was amplified with oligos s136 (5′-AACTAGCGTTTGCGTGCAGCGG-3′) and s145 (5′-CCTCCCGGGGAATTCCATGGAGAATATGGGAAC-3′) and cloned into pGEM-T vector to give plasmid pLC23. The EcoRI/SalI fragment from pLC23 was ligated into the EcoRI/SalI sites of pGEX5X-3 (Pharmacia) to give the GST-TFL1 fusion protein.
To produce CBD-INTEIN-TFL1, the TFL1 ORF was amplified with oligos s170 (5′-CTGCAGCTAGCGTTTGCGTGCAGCGGT-3′) and s171 (5′-GCTCTTCTAACATGGAGAATATGGGAACT-3′) and cloned into pGEM-T to give pLC1. pLC1 was digested with SapI and a vector-derived PstI site and ligated into pTWIN2 (New England Biolabs) previously cut with SapI/PstI to give plasmid pLC6, resulting in the CBD-INTEIN-TFL1 fusion.
To produce 35S-TAPtag-TFL1, the TFL1 ORF was amplified by PCR from the TFL1 cDNA with oligos s174 (5′-TAAGCTTATGGAGAAATATGGGAACT-3′) and s175 (5′-TAAGCTTGGATCCTCTAGACTAGCGTTTGCGTGCA-3′) and cloned into pGEM-T to give plasmid pLC10. The TAPtag ORF was amplified by PCR from pBS1761 (Puig et al., 2001) with oligos s176 (5′-AGGATCCATGGCAGGCCCCTTGCG-3′) and s177 (5′-AAAGCTTATCGTCATCATC-3′) and cloned in pGEM-T to give plasmid pLC11. pLC10 was digested with HindIII and ligated into pLC11 to give pLC14. This was digested with BamHI, and the TAPtag-TFL1 fragment was cloned into the BamHI site of pJIT62 to give plasmid pLC15 (CaMV35S-TAPtag-TFL1-CaMVTerm). pLC15 was cut with KpnI to release the overexpression cassette, and this was cloned into the KpnI site of the pGREEN 0229 Bar binary vector to give pLC37 (Hellens et al., 2000).
SDS-PAGE and Immunoblots
Proteins were size-fractionated on 15% SDS-PAGE gels and gels stained with Coomassie Brilliant Blue, or proteins were transferred to nitrocellulose filters for immunoblots (Sammy Dry; Schleicher and Schuell). High-resolution gels used modified conditions as described (Schagger and von Jagow, 1987).
Filters were incubated in TTBS (5% [w/v] dry nonfat milk, 25 mM Tris-HCl, pH 8, 150 mM NaCl, and 0.05% [v/v] Tween 20) before incubation with different primary antibodies: rabbit anti-TFL1 (diluted 1:1000), rabbit anti-CEN (diluted 1:1000), rabbit anti-histone H3 acetyl K9 (Abcam) (diluted 1:500), or goat anti-TFL1 antibody (Santa Cruz Biotechology) (diluted 1:200). Filters were washed and incubated with the secondary antibody (anti-rabbit or anti-goat horseradish peroxidase–conjugated [Pierce]), diluted 1:20,000 in TTBS. Filters were washed in TTBS and incubated with the peroxidase substrate (Super Signal West Pico; Pierce) before exposure to film (Hyperfilm ECL; Amersham Pharmacia Biotech).
Arabidopsis Protein Extractions
Arabidopsis proteins were extracted as described (Martinez-Garcia et al., 1999). Cell membranes were isolated from ap1 cal meristem tissues (2 g) as described (Kjellbom and Larsson, 1984). Total homogenates were clarified by centrifugation at 7800g, and the resulting solution referred to as the total (T) protein extract. This was spun at 140,000g to produce a supernatant soluble fraction (S) and a high-speed pellet. This pellet was washed and respun to give the final pelleted membrane fraction (P).
Nuclei-enriched fractions were prepared from ap1 cal meristem tissues (2 g) as described (Xia et al., 1997). Fresh tissues were ground in a chilled mortar with a pestle and clarified by filtration through Miracloth and nylon mesh (50-μM pore) to give the total protein extract (T). Nuclei were pelleted by low-speed centrifugation (1500g), and the resulting supernatant was referred to as the cytosolic fraction (C). The nuclear fraction was washed and repelleted to give the nuclei-enriched fraction (N).
Proteins were quantified with the Bradford assay kit (Sigma-Aldrich) using BSA as a standard. Forty micrograms of protein extracts were mixed with an equal volume of 2× Laemmli protein loading buffer, heated to 95°C for 5 min, and loaded onto gels.
Protein in Situ
Arabidopsis plants were harvested, and all visible leaves (>2 mm) were removed with a sharp razor blade. Samples were fixed in formaldehyde-acetic acid solution and embedded in Paramat extra (BDH Laboratory Supplies) as previously detailed (Sessions et al., 2000).
Sections (8 μm) were dewaxed and hydrated through a decreasing ethanol series. Slides were boiled in an antigen retrieval solution of 10 mM citrate (pH 6) for 10 min and then left to cool for ∼30 min at room temperature (Shi et al., 1997). Sections were rinsed in deionized water and incubated with BB (5% [w/v] nonfat dry milk, 0.9% [w/v] NaCl, and 10 mM Na phosphate, pH 7.4) overnight at 4°C in a humidified chamber. Sections were incubated with affinity-purified anti-TFL1 antibodies diluted 1:200 in BB for 6 h at 4°C. Slides were washed three times at room temperature with PBSX (PBS and 0.3% [v/v] Triton X-100) with gentle rocking. Secondary antibody (alkaline phosphatase-conjugated AffiPure goat anti-rabbit IgG, Fc fragment specific; Jackson Immunoresearch) was diluted in BB to 1:2000 and added to slides that were incubated again overnight at 4°C in a humidified chamber. The next day, slides were washed three times in PBSX, followed by 10 min in 100 mM Tris-HCl, pH 7.5, and 150 mM NaCl, followed by a 10-min wash in buffer 5 (100 mM Tris-HCl, pH 9.5, 100 mM NaCl, and 50 mM MgCl2). Slides were then incubated in the presence of the substrate solution with 5-bromo-4-chloro-3-indolyl phosphate (BCIP) and nitroblue tetrazolium (NBT) diluted in buffer 5 (0.33 mg/mL of BCIP and 0.165 mg/mL of NBT) and left in the dark to develop signal (∼2 to 6 h). The color reaction was stopped by several rinses in deionized water. Slides were then washed through an ethanol series, counterstained with 0.1% (w/v) calcofluor, and dried before mounting with Entellan (Merck) and a cover slip.
RNA in Situ Experiments
The RNA in situ experiments were performed as described (Ratcliffe et al., 1999).
Accession Numbers
Sequence data from this article can be found in the GenBank/EMBL data libraries under accession numbers U77674 (TFL1), M91208 (LFY), and Z16421 (AP1).
Supplemental Data
The following materials are available in the online version of this article.
Supplemental Figure 1. Purification of TAPtagTFL1.
Supplemental Figure 2. MALDI-TOF Analysis of Purified TAPtagTFL1.
Supplemental Figure 3. TFL1 Protein in Situ Controls.
Supplemental Figure 4. AP1 Is Found Only in Floral Meristems.
We thank Kim Baumann, Enrico Coen, and Yoshie Hanzawa for critical reading of the manuscript and stimulating discussions.
The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantcell.org) is: Desmond Bradley (desmond.bradley{at}bbsrc.ac.uk).
1 Current address: Biomedical and Life Sciences Department, University of Glasgow, Glasgow, G12 8QQ, UK.
[W] Online version contains Web-only data.
- Received December 18, 2006.
- Revised February 14, 2007.
- Accepted February 23, 2007.
- Published March 16, 2007.


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Six exo-planets are circling red dwarf star Gliese 581 20 light-years distant in the constellation Libra
Astronomy News – The human search for an exoplanet capable of being a cradle for a new human genesis found what many consider the first exoplanet with the physical makeup to make it possible. A team of planet hunters from the University of California (UC) Santa Cruz and the Carnegie Institute of Washington recently announced to the world the discovery of an exoplanet they believe has a few characteristics of an exoplanet with the right stuff to make life possible. Gliese 581g, as it's referred too, has about three times the mass of Earth and appears to be situated in the right spot in the solar system of the red dwarf star Gliese 581 for the ingredients of life to exist. This is about dead center in what planet scientists term the habitable zone of Gliese 581, a position planet scientists believe could make it possible for water and an atmosphere to exist on this exoplanet, necessary ingredients for the formation of life, planet scientists believe.
Astronomers search for a cradle for a new human genesis
These planet hunters have been using one of the largest time-machine-to-the-stars on the planet, the Keck I Telescope in Hawaii's W.M Keck Observatory, to journey 20 light years to the constellation of Libra to continue the search for more planets circling red dwarf star Gliese 581 that could be habitable. Planet hunters have been using the HIRES spectrometer to precisely measure the radial velocity of the host star – the motion of the star along the line of sight from Earth – and stars close to red dwarf star Gliese 581, in order to try to find other planets circling this red dwarf star. The gravitational pull of orbiting planets causes periodic changes in the radial velocity of the host star that astronomers can calculate using sophisticated mathematical techniques we'll cover on another day. These are the techniques planet hunters used in order to find all of the stars they have found circling red dwarf star Gliese 581, which after the two most recent planet discoveries, brings the total to six exoplanets circling this distant star.
Astronomers believe Gliese 581g is in the habitable zone of its home star
The discovery of six exoplanets circling red dwarf star Gliese 581 marks the high-planet mark for the human hunt for planets capable of being a cradle for a new human genesis. Gliese 581g is the only planet of the six exoplanets discovered that astronomers have indicated, so far, as being in the life zone of the red dwarf star Gliese 581. This exoplanet orbits its parent star in about 37 days and measurements planet scientists have made of its mass indicates it's probably a rocky planet with a definite surface and enough gravity to hang onto an atmosphere. Gliese 581g is also tidally locked to its parent star, which means that one side of the planet is always facing its host star and in perpetual daylight. This makes some planet scientists believe that the best place for life to exist would be in the terminator, the part of the planet between the day and night sides of the planet.
Check out my latest astronomy website at http://astronomytonight.yolasite.com/.
Learn why astronomy binoculars are a popular choice with amateur astronomers
Read about the Anasazi Indians
Read about astronomers viewing a supernova they think might have given birth to a black hole


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Utah's bill to opt out of the Real ID Act , HB 234, has passed both chambers of the State legislature and is now on the way to the Governor's desk today. The bill was signed by the Senate President late afternoon.
Rep. Stephen Sandstrom, author of the bill, introduced HB 234 at the beginning of the Utah legislative session. He then watched it rocket through the local legislature after mounting public outcry over decisions to move forward with implementation of Real ID regulations for drivers licenses.


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Massage Therapy and Chiropractic
If you are one of millions of Americans suffering from chronic ailments such as lower back or neck pain, it can be difficult to know whether going to the chiropractor or getting a massage will alleviate your pain. The answer, which may surprise you, is often both. Today, chiropractic work and massage therapy are often seen as two sides of the same coin. Rather than being incompatible like many once thought, these types of bodywork are actually complementary.
Here at PAMT we regularly work closely with our trusted network of local chiropractors to make sure our clients are receiving all of the care they need. Massage therapy works with soft tissues to improve mobility and pain levels while chiropractic work is used to resolve structural issues within the body. This means that massage is useful before and/or after a chiropractic adjustment so that muscles can easily adapt to the new structural foundation of your body. Massage used before adjustments relieves muscle tension that may be pulling joints out of alignment and aids the body's ability to receive a more complete adjustment. Massage used after adjustments helps the body to recover and to relearn a new 'normal' position.
Many clients have found that chiropractic care in tandem with massage therapy was the best answer for them. This approach ensures that any structural and soft tissue problems are addressed together. Both treatment options are holistic in nature and focus on the underlying conditions and sources of your pain rather than just the symptoms. Together these treatments can keep you Active for Life!
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CoreVR for Free Until December 20185394
74058 8 8
|mori private msg quote post Address this user
|Now on the MP Website:
ALL MATTERPORT SPACES CONVERTED TO CoreVR FOR FREE UNTIL DECEMBER 2018
All Matterport Spaces created through December 31st, 2017, will be converted to CoreVR for free. Beginning January 1st, 2018, Matterport Spaces can be converted to CoreVR for only $19 (£16).
|Post 1 IP flag post
North Palm Beach, FL
|hometakes private msg quote post Address this user
|If they start charging for it, they are mad. No ifs ands buts or whats, they are officially crazy. Not one real estate client of mine will want to purchase this. Wearing Villahermosa residence. After gate, first right, then left. 4th house on the left side of the street. Wearing glasses by the general public has not been accepted yet, let alone realtors who are ultra slow to embrace technology. I spend $20 per new Matterport client giving them a free pair of VR Glasses in order to try and promote VR.
It just won't work...... Dont they know anything about agents in the USA?
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|mori private msg quote post Address this user
|Here in munich we had a real estate agent who extra bought the latest Samsung Gear VR with an S8 and was not able to set it up correctly without any help.
I think VR is good for trade fairs like the Expo Real, but not yet with the current apps.
Navigation is to complex, no multi language and offline is also not working in the real world. Had this issue with Daydream and Gear VR. Without a good connection you are totally lost.
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|BrianAshley private msg quote post Address this user
|I do not understand what the term Core VR mean. As has been the case for over a year, I can drop my Galaxie phone into my Samsung Headgear VR and play any Matterport Tour I have selected.
What does Core VR do differently than that? Is it an enhanced experience? Is the imagery better, or is there better navigation?
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|justinv private msg quote post Address this user
I might be wrong but I think that is core VR.
I consider myself a tech geek and I have honestly never seen it that way. I can almost guarantee none of my clients have ever viewed it either.
When it first came out, it was a paid service. I assume no one bought it, so they stayed offering it for free. I believe two deadlines for free core vr has passed and it is still free.
I wouldn't worry too much about it until they start charging!
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|Queen_City_3D private msg quote post Address this user
|@BrianAshley That is CoreVR. There is a CustomVR option that you can order, but the cost is somewhere around $500US or so. With the custom option there's a chance that you'd get steroscopic views (truer sense of 3D), but I don't think that's guaranteed.
Right now spaces convert to CoreVR for free, but there's a threat that they will start charging for this in the future.
I think that's ridiculous. As @hometakes mentioned, the usage for clients just isn't there and there's no way I can upsell that.
IF... (and that's a big if) Matterport is adamant that they will charge for the CoreVR, then they should just do it for basic plans and have the Business and Professional plans continue to get the free conversion. I imagine that would push more MSPs into their higher priced plans and reward the ones that are already there.
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|BrianAshley private msg quote post Address this user
|@queencity3d That makes more sense and I agree with assessment of MP trying to charge for this. They have a few years to wait.
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|mori private msg quote post Address this user
|MP updated it (fixed typing error in the headline) and it now says "DECEMBER 2017".
So in 2018 we should be for a in my opinion still not really great working VR versions, especially not the WebVR base ones. Still it´s a mess to explore MP spaces with Daydream.
See: https://matterport.com/virtual-reality
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This topic is archived. Start new topic?


text: Eisman upbeat on U.S., down on Canadian housing The man who earned his street cred by betting against subprime mortgages in the U.S., said he is pessimistic about the Canadian housing market where rising prices haven't been accompanied by personal income growth. That phenomenon is a real concern across the breadth and scope of the Canadian market, suggested Steve Eisman, who also identified Home Capital Group, a mortgage provider that trades in Canada, as a "short sell" at the 18th Sohn Investment Conference in New York recently. "If housing rolls over, this company is going to have serious problems," he told Barron's Financial Investment News. Eisman, the founder of Emrys Partners LP and currently the money manager with FrontPoint Partners, a division of Morgan Stanley, estimated the domestic funding gap for the big six Canadian banks at around $427 billion. That seems far from the truth currently. Home Capital Group posted a strong Q1 with diluted earnings per share up 13.2 per cent year over year to $1.72, and adjusted diluted earnings per share up 19.1 per cent year over year to $1.81. Home Capital Group is a holding company that operates through its principal subsidiary, Home Trust. As federally regulated trust company, Home Trust offers deposit, mortgage lending, retail credit and credit card issuing services, with offices in Ontario, Alberta, British Columbia, Quebec and Nova Scotia. The company has also been touted for its sound lending practices and the strength of its book. While Eisman has reservations about the Canadian market, he's bullish on U.S. housing. He said investors should buy shares of homebuilders, land and mortgage lenders as the U.S. market improves. "Housing is attractively priced versus incomes," Eisman told reporters. "While homebuilders are cheap, they have not priced in how much fundamentals have improved." SHARE


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Isaiah 66 Modern English Version (MEV)
True Worship
66 Thus says the Lord:
Heaven is My throne,
But to this man I will look,
The Vindication of Zion
5 Hear the word of the Lord,
7 Before she was in labor,
12 For thus says the Lord:
I will extend peace to her like a river
God's Reign and Final Judgment
14 Then you shall see this, and your heart shall rejoice,
17 Those who sanctify themselves and purify themselves in the gardens behind one tree in the midst, eating swine's flesh, and abominable things, and mice shall be consumed together, says the Lord.
18 For I know their works and their thoughts. The time shall come to gather all nations and tongues. And they shall come and see My glory.
19 I will set a sign among them, and send from them survivors to the nations: to Tarshish, Pul, and Lud—who draw the bow—to Tubal, and Javan, to the coastlands afar off who have not heard My fame nor seen My glory. And they shall declare My glory among the nations. 20 They shall bring all your brothers out of all nations as an offering to the Lord on horses, and in chariots, and in litters, and on mules, and on swift beasts to My holy mountain Jerusalem, says the Lord, as the sons of Israel bring an offering in a clean vessel into the house of the Lord. 21 I will also take some of them for priests and for Levites, says the Lord.
22 For as the new heavens and the new earth which I will make shall remain before Me, says the Lord, so shall your descendants and your name remain. 23 From one New Moon to another, and from one Sabbath to another, all flesh shall come to worship before Me, says the Lord. 24 And they shall go forth and look on the corpses of the men who have transgressed against Me. For their worm shall not die, nor shall their fire be quenched. And they shall be an abhorrence to all flesh.
Modern English Version (MEV)
The Holy Bible, Modern English Version. Copyright © 2014 by Military Bible Association. Published and distributed by Charisma House.
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