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Hospital adopts 'Lean Thinking'
PAMPA - Nurses at Pampa Regional Medical Center now exchange reports about patients at shift changes. The hospital recently learned that a high number of errors occurs during shift change.
Consumers stocking up on stamps
WASHINGTON - With a postal rate increase two weeks away, Americans are buying 30 million Forever stamps a day.
Mars in sweet deal to buy gum maker Wrigley
The Oracle of Omaha is betting that the country's candy jar is recession-proof.
Fed poised to cut key rate
WASHINGTON - Battling risky economic crosscurrents, the Federal Reserve is ready to bump down a key interest rate again to brace the wobbly economy.
Hrycyk Financial: Life After Taxes
Tax season just ended, and you may be feeling the results of anxiety over the annual deadlines, paperwork, and soaring numbers that accompany tax preparation each April.
Oklahoma trumps Tech
LUBBOCK - J.T. Wise scored once and drove in the go-ahead run with a single in the seventh inning to give Oklahoma a 3-2 win over Texas Tech on Sunday.
Divorces Granted: Randall County
Marriage Licenses Issued: Randall County
Marriage Licenses Issued: Potter County
Randall County District Courts
Divorces Granted: Potter County
Kyla Elliott
Janet and Scott Elliott of Amarillo announce the engagement of their daughter, Kyla, to Greg Lott.
Shannan Dee Lacy
Susan and Danny Jackson of Amarillo announce the engagement of their daughter, Shannan, to John Gregory Godbey. Grandparents of the bride-elect are Mabel Reed and the late Warren Reed.
Ashley Gayle Jones
Cynthia Jones and Billy Fred Jones Jr., both of Amarillo, announce the engagement of their daughter, Ashley, to Steven Brodi Busch.
Taylor McCall Saikowski
Rita and Anthony Saikowski of Amarillo announce the engagement of their daughter, Taylor, to Braden Christian Barrick. Grandparents of the bride-elect are Mr. and Mrs. Dewey E. Isham Sr. and Mr. Alexander Saikowski, all of Amarillo.
Roxane Elena Melton
Pamela and Douglas Melton of Florissant, Colo., announce the engagement of their daughter, Roxane, to Staff Sgt. Bradley Scott Phillips.
Sarah Trosper
Peggy Trosper of Canyon and Jay Trosper of Amarillo announce the engagement of their daughter, Sarah, to Ryan McBride, both of Canyon. Grandparents of the bride-elect are Elsie Trosper, the late E.J. Trosper, Mary Creel and the late Gordon Creel.
Kendra Hicks
Marilyn and Gary Grimes of Amarillo announce the engagement of their daughter, Kendra of Dalhart, to Jason Cantrell of Dimmitt. Grandparents of the bride-elect are Bulah and Doyle Grimes, Hazel Noble and the late Gerald Noble.
Courtnay Coons
RuNell and David Coons of Amarillo announce the engagement of their daughter, Courtnay, to Jeff Van Why of Irving. Grandparents of the bride-elect are the late LaRue and Larry Stitt. Stepgrandmother is the late Kay Stitt.
Jessica Ann Jordan
Karen and Rusty Jordan of Houston announce the engagement of their daughter, Jessica, to Christopher Carl Hill. Grandparents of the bride-elect are Joann and Kenneth Askew, Janelle Jordan and the late Larry Jordan. Great-grandparents are Ima and Travis Smith.
Nicole J. Corrigan
Dr. Suzanne and John J. Corrigan III of Coppell announce the engagement of their daughter, Nicole, to Scott W. Garrett.
Jessica Ashlee Moore
Sharon and Kenneth Cooper and Suzanne and Sammy Moore, all of Amarillo, announce the engagement of their daughter, Jessica, to Jeffery Dale Snyder of Amarillo. Grandparents of the bride-elect are Mr. and Mrs. Glen Brooks of Waxahachie and Ruby and Johnnie Moore of Amarillo. Stepgrandparents are Marlene and Darrell Cooper of Amarillo.
Meghan Dennison
Paula and Terry Ford of Mertzon and Jon Dennison of San Antonio announce the engagement of their daughter, Meghan, to Nick Kauten.
Jacy Elder
Cynda and Jack Elder of Amarillo announce the engagement of their daughter, Jacy, to David Ferril.
Kendra Kay Jones
Lorrie and Gene Jones of Canyon announce the engagement of their daughter, Kendra, to Jason Kyle Boehning.
Dear Abby: Certain death follows release of dove
Dear Abby: The wedding season is fast approaching, and every year, starry-eyed brides plan to release white doves to "freedom" as part of their wedding celebration.
Dr. Gott: Don't rush to meds for treating diabetes
Dear Dr. Gott: My recent nonfasting glucose was 102, and my A1C was 6.21. My physician wants to repeat the A1C in three months, and, if it is still elevated, he will recommend oral diabetic medication. What is your opinion? I am 82.
Universal appeal
Greg Welch gets a lot of compliments on his skin.
Dear Abby: Gossip is topic of high interest
Dear Abby: I live in a medium-size community where a lot of people know one another. I often hear them discuss such private matters as the state of someone's marriage, their finances, etc.
Heloise: Another mammogram hint
Dear Heloise: I read your article on mammogram hints, and I'm hoping you will add another helpful hint.
Heloise: Double-check prescriptions
Dear Readers: When you called in or dropped off your last prescription, did you check it when you picked it up to be sure you got what the doctor ordered?
Dr. Gott: Reader sees red about grocery store meat
Dear Dr. Gott: My in-laws insist that beef sold in most supermarkets is dyed red, and they soak their meat to remove the dye. Is this true?
Tascosa senior inks soccer pact
It was a season of firsts for Tascosa's girls soccer team.
Athlete of the Week: Cameron Swarb
On the Week: The senior qualified for state in two sports for the second straight year after winning regional titles in golf and tennis in boys doubles, teaming with his bother, Cragar.
Bushland's Daniel to play for Buffs
Bushland's Patrick Daniel has agreed to play football at West Texas A&M as a preferred walk-on, Falcons coach David Flowers announced.
Randall stays positive
SAN ANGELO - On Page 2 of the results of the Region I-4A girls 1600 relay preliminaries was Randall.
Bidistrict pairings set for area baseball teams
After a week of battles for playoff berths, the high school baseball postseason begins this week as area teams begin journeys they hope end in the state tournament at Round Rock.
Athlete of the Week: Katie Grimes
On the Track: The junior helped the Lady Eagles nab the Region I-3A title in Odessa by winning five gold medals for the second straight year.
Canyon's record relay time changed
Canyon's girls 1600 relay was good but not that good.
Dillas manager ready to start season
Amarillo Dillas fans will see several familiar faces return this season in the place where new manager Brady Bogart thinks championships are built.
Rangers club the Twins, 10-0
ARLINGTON - Vicente Padilla did what right-handers seldom do - throw a shutout at hitter-friendly Rangers Ballpark in Arlington.
Another poor start raises concern
Ron Washington and his Texas Rangers got a much-needed day off Monday, the first time in three weeks they didn't have to be at a ballpark for a game.
Rodeo: Childress calf roper Smith adds to bank account
The Professional Bull Riders is bringing its top-tier tour to Texas twice in the next two months, and if recent trends hold true, fans can expect to see riders turn in some big scores.
Stars going home with 2-0 advantage
SAN JOSE, Calif. - Mike Modano and the Dallas Stars probably don't want to leave the Shark Tank.
NFL approves trade involving 'Pacman'
IRVING - The NFL formally approved Tennessee's trade of suspended cornerback Adam "Pacman" Jones to the Dallas Cowboys on Sunday.
Stung in their house
DALLAS - David West hardly said a word the past two days.
Arrest made in attacks
A Honduras man was arrested Sunday on two counts of burglary of a habitation with the intent to commit sexual assault and one count of aggravated sexual assault stemming from three reported incidents in Wellington on women age 60 or older.
Foxcroft Drive to be closed for repairs
Foxcroft Drive will be closed for repairs from Coulter Street to Spencer Street beginning Thursday.
Docket: Potter County District Courts
Window on the Past: Racing to the finish line
Riders gasped for breath, some wobbling, some walking, as they came to the end of a 15-mile American Cancer Society Bike-A-Thon in this Amarillo Globe-News file photo from Saturday, May 2, 1981.
Court nixes holy icon
Potter County won't be tempting legal fate by allowing the placement of a monument with the Ten Commandments and various quotes inscribed on its surface at the Potter County Courthouse.
Blind, deaf man an entrepreneur
Preston Fansler's parents don't want him to be defined by what he can't do.
1 hurt in hit-and-run
One person was taken to the hospital and another was still on the run after a two-vehicle accident Monday night at South Western Street and Paramount Boulevard.
Families to get kindergarten preview
Moms and dads don't have to wait until the first day of school to introduce their kindergartners to the campus.
Tech to screen for depression
Seniors who suffer from depression are twice as likely to die of a stroke or heart attack, said Steve Burgess, a fourth-year medical student at Texas Tech University.
Cactus city leader resigns
CACTUS - Jeff Jenkins is resigning as city manager of Cactus, a Panhandle city that continues to struggle to recover from a devastating tornado that leveled nearly half the town.
Eye on Austin: Dem donors spread wealth
Apparently all that talk of a divided Democratic Party is not hurting Texas Democrats.
Men can be moms, too
Glenn Tignor, 30, and wife Kathy Burdge made a rather unpopular choice almost four years ago. The choice is one that Tignor often takes a great deal of flak over - especially from his own family members.
Food Establishment Inspection Report
The Food Establishment Inspection Report is a weekly report that appears Mondays. Information is taken from reports provided by the Bi-City-County Health District and individual reports can be viewed at their Web site at www.ci.amarillo.tx.us/departments/environhealth/foodestablishments.htm.
AISD goes green
School cafeteria operators for Amarillo Independent School District began participating in a new recycling program this month.
'Grand Theft' ready
Gamers are already making plans for this week's release of "Grand Theft Auto IV."
Another for the shelf
Shelly Patterson, left, sits with a first-place trophy on her 1929 Ford Ratrod at the Route 66 Roller Derby Car Show on Sunday at Hillside Road and Bell Street. Aspen Eager, 6, and Maeci Day, 6, climb over the seats.
Docket: Randall County District Courts
WT to study African tale
West Texas A&M University on Monday announced the next book in its common-readership program: Dave Eggers' "What is the What."
Uninsured set to rise in city
Amarillo's uninsured population will more than double by 2040 as more people choose to live without health insurance, according to data released Monday by a Texas health policy think tank.
It's All Trew: Lots of laughter in Old West
No matter how serious the history of the Old West gets, there is always a little humor included if you keep reading.
Living the dream
Standing ovations inside Radio City Music Hall are generally bestowed upon renowned recording artists and movie stars.
Jones on second day: Let's make a deal
IRVING - Jerry Jones has lived up to his reputation as a dealer.
Nancy Carol Simmons-Alexander
Nancy Carol Simmons-Alexander, 61, of Amarillo died Sunday, March 23, 2008.
Jewell Lambert Culp
HEREFORD - Jewell Lambert Culp, 92, died Sunday, April 27, 2008.
Mary D. Harman
TULIA - Mary D. Harman, 84, died Saturday, April 26, 2008.
Jerry W. Slaton
HANSTON, Kan. - Jerry W. Slaton, 62, died Thursday, April 24, 2008.
Rose Elizabeth "Betty" Blankenship Renfro
Rose Elizabeth "Betty" Blankenship Renfro, of Amarillo, died Sunday, April 27, 2008.
Billy McClelland Gillman
Billy McClelland Gillman, 73, of Amarillo died Thursday, April 24, 2008.
Raymond G. Foster
Raymond G. Foster, 84, of Amarillo, died Saturday, April 26, 2008.
Jamie Alejandro Martinez
BEAVER, Okla.- Jamie Alejandro Martinez, 25, died Thursday, April 24, 2008.
Earl B. Gullett
DUMAS - Earl B. Gullett, 91, died Monday, April 28, 2008.
Virgil Wade Wallin Jr.
PAMPA - Virgil Wade Wallin Jr., 55, died Sunday, April 27, 2008.
Joaquin De La Cruz
DIMMITT - Joaquin De La Cruz, 87, died Sunday, April 27, 2008, in Amarillo.
Janice Margaret Stevenson
HEREFORD - Janice Margaret Stevenson, 67, died Sunday, April 27, 2008.
J.B. Bennett
J.B. Bennett, 91, of Amarillo died Saturday, April 26, 2008.
Joe Moore
TULIA - Joe Moore, 65, died Sunday, April 27, 2008, in Amarillo.
Peggy Lee Mink
CHILDRESS - Peggy Lee Mink, 81, died Saturday, April 26, 2008.
Michael E. Stanley
BORGER - Michael E. Stanley, 45, died Saturday, April 26, 2008.
Ruth Geneva Wallace Mills
CLARENDON - Ruth Geneva Wallace Mills, 94, died Sunday, April 27, 2008.
Mary D. Harman
TULIA - Mary D. Harman, 84, died Saturday, April 26, 2008.
Leona Esther Schwope Kivlehen
CONROE - Leona Esther Schwope Kivlehen, 88, died Saturday, April 26, 2008.
Elsie Louetta Slaughter Hawkins
SHAMROCK - Elsie Louetta Slaughter Hawkins, 61, died Saturday, April 26, 2008.
Bryce R. Cook
Bryce R. Cook, 75, of Amarillo, died Friday, April 25, 2008.
Elsie Louetta Slaughter Hawkins
SHAMROCK - Elsie Louetta Slaughter Hawkins, 61, died Saturday, April 26, 2008.
Letter: Public safety needs protection
"Business owner rights!" What about workplace and public safety?
Letter: 'Rights' take back seat to public health
Those who decry the proposed smoking ordinance because of a perceived loss of "freedom" are blowing smoke in their own eyes. It's not about a business owner's "freedom" to run his business as he chooses. It's not about a smoker's "freedom" to light up anywhere he chooses, nor is it about one group's rights vs. those of another group.
Greg Sagan: What else should we put on courthouse lawn?
Shall we erect the Ten Commandments on the courthouse lawn? If you believe we shouldn't then the next question is, why not?
Dominick Miller: Founders excluded religion
I am writing in response to two articles:Keith Pelton's April 17 guest column, "What church-state separation?" and an April 16 letter to the editor by Doug Solsbery, which argues that the United States is a religious country, per the Declaration of Independence, and should be treated as such.
Letter: Everyone wins with water settlement
I would like to respond to a recent editorial praising the City of Amarillo's settlement with Premium Standard Farms related to its claims of alleged overproduction of groundwater. I think it is important to point out that during this entire process, Premium Standard Farms has been trying to be a good neighbor, but your readers need to know the "rest of the story."
Editorial: Let's start voting trend
The last time Amarillo had a public smoking ban on the ballot in 2005, nearly 30 percent of registered voters went to the polls.
Editorial: Long arm of law crosses state line
States have the power to prosecute the accused as the see fit. The U.S. Constitution gives that power specifically to each state.
Editorial: Written in stone and concrete
Potter County Commissioner Lacy Borger commented Monday that there are two things that are difficult to get done before dark - a discussion on religion and pouring concrete.
Letter: Let people decide for themselves on smoking
I am not a smoker. In fact I am moderately allergic to cigarette smoke. I am still against the sweeping ban that is to be voted on in May.
Stallings to speak at WT luncheon
West Texas A&M athletics will host a luncheon featuring former Alabama and Texas A&M football coach Gene Stallings on Thursday at the Amarillo Civic Center Grand Plaza Ballroom.
Lady Buffs drop finale
Abilene Christian pushed across a run in the top of the eighth inning, sending the Lady Wildcats past West Texas A&M, 3-2, Sunday afternoon in its softball season finale at Lady Buff Yard.
Browns ink WT's Kapanui
When NFL training camps start this summer, seven former West Texas A&M players are expected to be participating in workouts.
WT's Lee scores 14 in All-Star contest
West Texas A&M senior Robert Lee made 5-of-12 shots from the floor, finishing with 14 points on the West All-Star team during the Collegiate Basketball Invitational All-Star game Saturday night in Wadsworth, Ohio.
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Gold Virgin Media TiVo box pictures and hands-onPocket-lint
To celebrate reaching one million customers for its TiVo service, Virgin Media commissioned an extremely limited edition gold version of its 1TB set-top-box. So limited, in fact, that there's only one in existance.
And, like with the vajazzled version of the box made in honour of The Only Way is Essex, Pocket-lint has managed to get its hands-on it long enough to take a batch of exclusive photos.
The box itself has also been in the hands of a lucky couple from Bramcote in Nottinghamshire, Allan and Christine Ward, who, thanks to Virgin Media, got to meet their favourite celebrity, actor and ex-Spandau Ballet bassist Martin Kemp (Gold, get it?). He performed a celebrity install of their Virgin Media TiVo service at their home, and the Wards also got a brand new 3D TV to watch it on.
Looking at the gold box itself, while it's not exactly covered in the precious metal (rather, spray painted by experts) it's definitely a desirable colour scheme. We also put it up against the chrome VM TiVo remote that was created near the beginning of the service's introduction for VIPs and celebrities, and it fits right in.
It also brings back a few memories of yesteryear, as we'd swear blind that the sparkling finish is identical to the one adopted for the Golden Gamesmaster Joysticks in later series of the Channel 4 show from the Nineties. Originally, the joysticks were gold plated, but further down the line they were sprayed in gold car paint. Some walls in the Docklands probably still show the gold-rimmed sillouettes of Quickshot Pros - like games-oriented Banksy graffiti.
We digress though. Unfortunately, we couldn't actually plug the gold TiVo box into our cable service as it hadn't been set up for our account, so only got to play with the exterior, but from what we have seen, we reckon owners would lap up a limited run of these boxes should Virgin Media make them more widely available.
What do you think of the gold box? Would you want one? Let is know in the comments below...
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Best TV catch-up on Freeview Play: Robot Wars, Kinky Britain and more
Pokemon Go the Movie planned, live-action film rights snapped up
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subset: commoncrawl


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We've found a little gem tucked away in a tiny corner of southwest France—the Lot Valley, a secluded world of medieval cliff-top hamlets, Romanesque abbeys, open-air markets, ancient chateaux, robust red wines, and a savory cuisine flavored with truffles and wild mushrooms. On our enchanting exploration of this deeply French realm, our hikes bring you along the tranquil Lot River and across pilgrim paths between spectacular hilltop villages. You'll also discover the Pech Merle prehistoric cave, filled with extraordinary rock art, and sample the delicious wares of local wineries and small farms, including tangy Cabécou goat cheese. Overnights are in hand-selected historic hotels, including the majestic 13th century Château de Mercuès overlooking the Lot River.
We expected this would be a great trip and we were not disappointed. We saw beautiful villages, lovely valleys, stayed at great hotels, had interesting tours, picturesque hikes, and interesting excursions. It was a section of France we would never have explored on our own.
— Cynthia S., San Diego, CA
Days 1-3
Rodez / Conques
Our hikes bring you through picturesque hamlets north of Rodez, to the hilltop village of Conques, whose 9th century abbey is a spectacular example of Romanesque architecture, and along a pilgrim path to medieval Estaing, officially listed as one of les plus beaux villages de France.
Days 4-5
Célé Valley / Rocamadour
After exploring Figeac, a charming 15th century town with a perfectly preserved medieval center, we hike on an ancient path along the Célé River. Our next hike leads across the dramatic gorge of the Alzou River for spectacular views of cliff-perched Rocamadour, a pilgrim destination since 1166.
Day 6
Pech Merle Cave / St. Cirq Lapopie
At Pech Merle, one of the few prehistoric caves where the original 25,000-year-old paintings are on view, take in a treasury of vivid rock art as mammoths, bison, and dappled horses gallop across the cave walls. Later, hike a riverside trail to St. Cirq Lapopie, with its steep cobbled streets and terraced gardens. Our hotel for the next three nights is the majestic 13th century Château de Mercuès near Cahors.
Days 7-9
Château de Cénevières / Cahors
From the rock-perched village of Calvignac, we hike a forested path to the 13th century Château de Cénevières, where the lord of the chateau welcomes us for a visit. In sunny Cahors, explore the lively open-air market and enjoy scenes of daily life including beret-topped old men and housewives seeking out the freshest produce. A sumptuous wine-tasting in our hotel's vaulted cellar is followed by our farewell dinner in its Michelin-starred restaurant. Depart on Day 9 via Toulouse.
A mix of easy to moderate walks and hikes, 2-4 hours a day, van support (for luggage or if you wish to skip a day's hike)


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Dried Blood Plums
Total Time
24hrs 30mins
Prep 30 mins
Cook 24 hrs
Blood plums are delicious Japanese plums with bright red flesh. I usually can them, but I decided to try drying them, even though I couldn't find any instructions for doing so. Here is what I did, which was simple and produced dried plums with incredible flavor. I have included a photo sequence of the process.
Ingredients Nutrition
- 10 lbs of blood plums (or any amount you want to dry. I buy a twenty pound box at the farmer's market)
- water (for washing plums, recipezaar said I needed to have more than one ingredient, lol)
- Wash plums.
- Cut into wedges, removing pit (quarters or sixths depending on the size of the plum).
- Place on dehydrator rack (or some ovens have low temperature setting for drying fruit).
- Dry at 135 degrees for about 24 hours. Drying time may vary considerably depending on humidity levels.
- They are done when they are leathery and sticky, but no longer juicy.
- Store them in ziplock freezer bags in the freezer until ready to use.
- FYI - Some varieties of blood plum are Ruby Blood, Satsuma and Mariposa.


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A unique social movement has taken root in Atlanta. Small urban farms continue to pop up within the city limits, bringing farm closer to table than ever before. And with the growing popularity of farmers markets, and general food awareness, it's likely that this new trend is here to stay.
Love is Love Farm at Gaia Gardens
Driving south on Glenwood Avenue, East Atlanta quickly fades into East Lake. A right turn down a residential street dead-ends at a fence with a sign that reads: ORGANIC FARM, DO NOT SPRAY. The gate is overrun with honeysuckle. It smells like flowers and the forest floor.
Once inside Gaia Gardens, the only sign of city life is an occasional plane flying overhead.
Love is Love farmer Joe Reynolds is the fifth farmer to grow at Gaia Gardens. "Judith, my sweetie, and I don't own this, we are landless farmers that grow and sell as Love is Love Farm."
The land is owned by East Lake Commons, a cooperative housing community in East Lake founded 15 years ago by a group of forward-thinking individuals. "When they bought the land," says Reynolds, "they challenged the developer with creating a really amazing, tight-knit community, while reserving a little less than half of the land for a working organic farm."
As the current leaseholder, Reynolds' operation must honor guidelines set by the housing community. For example, the farm is required to be certified organic and grow a wide diversity of crops at all times.
Another stipulation of the lease is that the farm must provide a Community Supported Agriculture program, a CSA for short. With a CSA, members or subscribers pay for a share of the anticipated harvest before the season; once harvesting begins, they receive weekly shares of vegetables and fruit.
"It's helpful because I can't really predict every detail that the farm has to deal with," says Reynolds. "But with the CSA I know how much food we need to have, and I can really do a lot of budgeting around that information," he continues. "Another reason I love the CSA is I get a really deep feeling of community with those folks. Some of them have been with me ever since the first day that we sold food."
Often, the criticism of buying food from local farmers is that it costs more, especially when mainstream food prices are so low in comparison. "I get that," says Reynolds, "but nobody's making a fortune. I make a really modest salary. We run a pretty tight ship cost-wise."
For Reynolds, the bigger questions is: How do you make it so that farmers don't have to drop their prices while people who don't have the economic means to meet those prices can still have access to that food?
Even with the obstacles Reynolds feels lucky to be a part of the significant social movement toward localism in Atlanta. "When I entered adulthood, I just wanted to do something meaningful and farming is just kind of a win on so many levels. It's not only meaningful, but it's also good work, and there's a lot of it."
Just a few blocks south of Ralph David Abernathy Boulevard, near the heart of Atlanta's historic West End neighborhood, Cecilia Gatungo and Jamila Norman of Patchwork City Farms prepare for market. Huge greenish-purple leaves are carefully snipped, bundled, and placed tenderly in a basket. "It's actually a mustard green," says Gatungo. "An Asian variety. It's red and tastes just like wasabi, but it mellows out when you cook it."
Gatungo and Norman met and bonded while cleaning up parks and volunteering at the Good Shepherd Community Garden in West End. "We were talking about how lousy our Kroger was when we realized that we had a shared goal," says Norman. "We both wanted to grow food for ourselves and our families." Jumping in headfirst, the two began growing a variety of fruits and vegetables in Gatungo's front yard. "It was insane. Our neighbors thought we were crazy."
Soon, what began as a successful home garden grew into a commercial operation on half an acre of land leased from the Atlanta Public Schools System. By farming in West End, Gatungo has found a way to reconnect with her roots. "As a kid, everything we ate came from our garden. I wanted the kids in the neighborhood to have that, too."
During a time when diet-related illnesses such as diabetes and heart disease plague Americans everywhere, Norman is grateful for the health benefits of urban farming. "Nature's already got an eating plan that we should be working with that's helping us stay healthy and in the right rhythms."
As small urban farms like Patchwork City begin to take root in Atlanta, progress depends on whether or not consumers are willing to move away from buying cheap, processed food, and buy into ideas like seasonal eating and farmers markets despite relatively higher prices.
"When people support local agriculture, they're making an economic and political choice," says Gatungo. "A choice to support the local economy and to pay a fair price for their food. A lot of times we get asked why our food is so expensive. My quickest answer is, 'You can pay me more, or you can pay your doctor more.' There are hidden costs to everything."
"Supporting local farms creates jobs and keeps people employed in an industry that is helping your health and helping the environment," says Norman. "It's just the right thing to do."


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Indiana Basketball: Ride or Die with Yogi Ferrell
Last season, the man who loomed largest on the Indiana Hoosiers was often the most physically overshadowed on the court. Six-foot-10 NBA lottery pick Noah Vonleh drew the hungry eyes of pro scouts while departing 6-foot-7 senior Will Sheehey reveled in the cheers of his send-off season in Bloomington. Yet, on most nights, all the towering trees and tantalizing prospects galloping...
Via isportsweb.com
August 25, 2014 | Discuss
Report: Will Sheehey to play professionally in Montenegro
It appears that former Indiana forward Will Sheehey has found a home for his first season of professional basketball. Sheehey will join KK Buducnost of the Erste Liga in Montenegro, according to the team's website (translated text below): Two new players of today are officially members of the Future I-LOVE. Croatian center Robert Rikic (219cm, 24g), […] The post Report: Will...
Via Inside the Hall
August 01, 2014 | Discuss
Sheehey to play for Knicks in NBA summer league
Former Indiana forward Will Sheehey wasn't selected in Thursday's NBA draft, but he'll still get a shot to play in the NBA. After working out for 10 teams in the pre-draft process, Sheehey had some second round interest, but didn't hear his name called. He'll now have to play his way onto a roster and […] The post Sheehey to play for Knicks in NBA summer league appeared...
Via Inside the Hall
June 27, 2014 | Discuss
Sheehey continuing to adjust to new role
Indiana coach Tom Crean knows he is asking for a lot from Will Sheehey. In the Hoosiers' first 10 games of the 2013-2014 season, Crean has bestowed the role of starter, leader, defender, scorer and rebounder on a player who has typically come off the bench for much of his career as a sixth man. […]The post Sheehey continuing to adjust to new role appeared first on Inside the...
Via Inside the Hall
December 13, 2013 | Discuss
Video: Crean, Ferrell, Sheehey react to win over Washington
NEW YORK — Tom Crean, Will Sheehey (16 points, five assists) and Yogi Ferrell (20 points, five assists) met with the media following Indiana's 102-84 win over Washington on Thursday in the 2K Sports Classic at Madison Square Garden. Watch their postgame comments in the media player below:The post Video: Crean, Ferrell, Sheehey react to win over Washington appeared first on Inside...
Via Inside the Hall
November 22, 2013 | Discuss
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Phylogeny and evolution of Rab7 and Rab9 proteins
Mackiewicz, Paweł
Wyroba, Elżbieta
BMC Evolutionary Biology
An important role in the evolution of intracellular trafficking machinery in eukaryotes played small GTPases belonging to the Rab family known as pivotal regulators of vesicle docking, fusion and transport. The Rab family is very diversified and divided into several specialized subfamilies. We focused on the VII functional group comprising Rab7 and Rab9, two related subfamilies, and analysed 210 sequences of these proteins. Rab7 regulates traffic from early to late endosomes and from late endosome to vacuole/lysosome, whereas Rab9 participates in transport from late endosomes to the trans-Golgi network.
Although Rab7 and Rab9 proteins are quite small and show heterogeneous rates of substitution in different lineages, we found a phylogenetic signal and inferred evolutionary relationships between them. Rab7 proteins evolved before radiation of main eukaryotic supergroups while Rab9 GTPases diverged from Rab7 before split of choanoflagellates and metazoans. Additional duplication of Rab9 and Rab7 proteins resulting in several isoforms occurred in the early evolution of vertebrates and next in teleost fishes and tetrapods. Three Rab7 lineages emerged before divergence of monocots and eudicots and subsequent duplications of Rab7 genes occurred in particular angiosperm clades. Interestingly, several Rab7 copies were identified in some representatives of excavates, ciliates and amoebozoans. The presence of many Rab copies is correlated with significant differences in their expression level. The diversification of analysed Rab subfamilies is also manifested by non-conserved sequences and structural features, many of which are involved in the interaction with regulators and effectors. Individual sites discriminating different subgroups of Rab7 and Rab9 GTPases have been identified.
Phylogenetic reconstructions of Rab7 and Rab9 proteins were performed by a variety of methods. These Rab GTPases show diversification both at the phylogenetic, expression and structural levels. The presence of many Rab7 and Rab9 isoforms suggests their functional specialization and complexity of subcellular trafficking even in unicellular eukaryotes. The identified less conserved regions in analysed Rab sequences may directly contribute to such a differentiation.
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Thousands of Rab GTPases for the Cell Biologist
Diekmann, Yoan
Seixas, Elsa
Gouw, Marc
Tavares-Cadete, Filipe
Seabra, Miguel C.
Pereira-Leal, José B.
PLoS Computational Biology
Rab proteins are small GTPases that act as essential regulators of vesicular trafficking. 44 subfamilies are known in humans, performing specific sets of functions at distinct subcellular localisations and tissues. Rab function is conserved even amongst distant orthologs. Hence, the annotation of Rabs yields functional predictions about the cell biology of trafficking. So far, annotating Rabs has been a laborious manual task not feasible for current and future genomic output of deep sequencing technologies. We developed, validated and benchmarked the Rabifier, an automated bioinformatic pipeline for the identification and classification of Rabs, which achieves up to 90% classification accuracy. We cataloged roughly 8.000 Rabs from 247 genomes covering the entire eukaryotic tree. The full Rab database and a web tool implementing the pipeline are publicly available at www.RabDB.org. For the first time, we describe and analyse the evolution of Rabs in a dataset covering the whole eukaryotic phylogeny. We found a highly dynamic family undergoing frequent taxon-specific expansions and losses. We dated the origin of human subfamilies using phylogenetic profiling, which enlarged the Rab repertoire of the Last Eukaryotic Common Ancestor with Rab14, 32 and RabL4. Furthermore, a detailed analysis of the Choanoflagellate Monosiga brevicollis Rab family pinpointed the changes that accompanied the emergence of Metazoan multicellularity, mainly an important expansion and specialisation of the secretory pathway. Lastly, we experimentally establish tissue specificity in expression of mouse Rabs and show that neo-functionalisation best explains the emergence of new human Rab subfamilies. With the Rabifier and RabDB, we provide tools that easily allows non-bioinformaticians to integrate thousands of Rabs in their analyses. RabDB is designed to enable the cell biology community to keep pace with the increasing number of fully-sequenced genomes and change the scale at which we perform comparative analysis in cell biology.
Author Summary
Intracellular compartmentalisation via membrane-delimited organelles is a fundamental feature of the eukaryotic cell. Understanding its origins and specialisation into functionally distinct compartments is a major challenge in evolutionary cell biology. We focus on the Rab enzymes, critical organisers of the trafficking pathways that link the endomembrane system. Rabs form a large family of evolutionarily related proteins, regulating distinct steps in vesicle transport. They mark pathways and organelles due to their specific subcellular and tissue localisation. We propose a solution to the problem of identifying and annotating Rabs in hundreds of sequenced genomes. We developed an accurate bioinformatics pipeline that is able to take into account pre-existing and often inconsistent, manual annotations. We made it available to the community in form of a web tool, as well as a database containing thousands of Rabs assigned to sub-families, which yields clear functional predictions. Thousands of Rabs allow for a new level of analysis. We illustrate this by characterising for the first time the global evolutionary dynamics of the Rab family. We dated the emergence of subfamilies and suggest that the Rab family expands by duplicates acquiring new functions.
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Comprehensive Analysis Reveals Dynamic and Evolutionary Plasticity of Rab GTPases and Membrane Traffic in Tetrahymena thermophila
Bright, Lydia J.
Kambesis, Nichole
Nelson, Scott Brent
Jeong, Byeongmoon
Turkewitz, Aaron P.
PLoS Genetics
Cellular sophistication is not exclusive to multicellular organisms, and unicellular eukaryotes can resemble differentiated animal cells in their complex network of membrane-bound structures. These comparisons can be illuminated by genome-wide surveys of key gene families. We report a systematic analysis of Rabs in a complex unicellular Ciliate, including gene prediction and phylogenetic clustering, expression profiling based on public data, and Green Fluorescent Protein (GFP) tagging. Rabs are monomeric GTPases that regulate membrane traffic. Because Rabs act as compartment-specific determinants, the number of Rabs in an organism reflects intracellular complexity. The Tetrahymena Rab family is similar in size to that in humans and includes both expansions in conserved Rab clades as well as many divergent Rabs. Importantly, more than 90% of Rabs are expressed concurrently in growing cells, while only a small subset appears specialized for other conditions. By localizing most Rabs in living cells, we could assign the majority to specific compartments. These results validated most phylogenetic assignments, but also indicated that some sequence-conserved Rabs were co-opted for novel functions. Our survey uncovered a rare example of a nuclear Rab and substantiated the existence of a previously unrecognized core Rab clade in eukaryotes. Strikingly, several functionally conserved pathways or structures were found to be associated entirely with divergent Rabs. These pathways may have permitted rapid evolution of the associated Rabs or may have arisen independently in diverse lineages and then converged. Thus, characterizing entire gene families can provide insight into the evolutionary flexibility of fundamental cellular pathways.
Author Summary
Single-celled organisms appear simple compared to multicellular organisms, but this may not be true at the level of the individual cell. In fact, microscopic observations suggest that protists can possess networks of organelles just as elaborate as those in animal cells. Consistent with this idea, recent analysis has identified large families of genes in protists that are predicted to act as determinants for complex membrane networks. To test these predictions and to probe relationships between cellular structures across a wide swath of evolution, we focused on one gene family in the single-celled organism Tetrahymena. These genes control the traffic between organelles, with each gene controlling a single step in this traffic. We asked three questions about each of 56 genes in the family. First, what is the gene related to in humans? Second, under what conditions is the gene being used in Tetrahymena? Third, what is the role of each gene? The results provide insights into both the dynamics and evolution of membrane traffic, including the finding that some pathways appearing both structurally and functionally similar in protists and animals are likely to have arisen independently in the two lineages.
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Toward a Comprehensive Map of the Effectors of Rab GTPases
Gillingham, Alison K.
Sinka, Rita
Torres, Isabel L.
Lilley, Kathryn S.
Munro, Sean
Developmental Cell
The Rab GTPases recruit peripheral membrane proteins to intracellular organelles. These Rab effectors typically mediate the motility of organelles and vesicles and contribute to the specificity of membrane traffic. However, for many Rabs, few, if any, effectors have been identified; hence, their role remains unclear. To identify Rab effectors, we used a comprehensive set of Drosophila Rabs for affinity chromatography followed by mass spectrometry to identify the proteins bound to each Rab. For many Rabs, this revealed specific interactions with Drosophila orthologs of known effectors. In addition, we found numerous Rab-specific interactions with known components of membrane traffic as well as with diverse proteins not previously linked to organelles or having no known function. We confirm over 25 interactions for Rab2, Rab4, Rab5, Rab6, Rab7, Rab9, Rab18, Rab19, Rab30, and Rab39. These include tethering complexes, coiled-coiled proteins, motor linkers, Rab regulators, and several proteins linked to human disease.
Graphical Abstract
•Proteomic screen identifies effectors of Drosophila Rabs with a human ortholog•Specific hits include orthologs of numerous known effectors of mammalian Rabs•Validated effectors include traffic proteins and those of unknown function•Orthologs of disease genes CLEC16A, LRRK2, and SPG20 are validated as effectors
Rab GTPases organize cellular compartments by recruiting specific effectors to organelle membranes. This paper describes affinity chromatography using all Drosophila Rabs with a mammalian ortholog. The Rab interactors found include known effectors, tethering complexes, coiled-coil proteins, motor proteins, proteins of unknown function, and several proteins linked to human disease.
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Structural Insights into a Unique Legionella pneumophila Effector LidA Recognizing Both GDP and GTP Bound Rab1 in Their Active State
Cheng, Wei
Yin, Kun
Lu, Defen
Li, Bingqing
Zhu, Deyu
Chen, Yuzhen
Zhang, Hao
Xu, Sujuan
Chai, Jijie
Gu, Lichuan
PLoS Pathogens
The intracellular pathogen Legionella pneumophila hijacks the endoplasmic reticulum (ER)-derived vesicles to create an organelle designated Legionella-containing vacuole (LCV) required for bacterial replication. Maturation of the LCV involved acquisition of Rab1, which is mediated by the bacterial effector protein SidM/DrrA. SidM/DrrA is a bifunctional enzyme having the activity of both Rab1-specific GDP dissociation inhibitor (GDI) displacement factor (GDF) and guanine nucleotide exchange factor (GEF). LidA, another Rab1-interacting bacterial effector protein, was reported to promote SidM/DrrA-mediated recruitment of Rab1 to the LCV as well. Here we report the crystal structures of LidA complexes with GDP- and GTP-bound Rab1 respectively. Structural comparison revealed that GDP-Rab1 bound by LidA exhibits an active and nearly identical conformation with that of GTP-Rab1, suggesting that LidA can disrupt the switch function of Rab1 and render it persistently active. As with GTP, LidA maintains GDP-Rab1 in the active conformation through interaction with its two conserved switch regions. Consistent with the structural observations, biochemical assays showed that LidA binds to GDP- and GTP-Rab1 equally well with an affinity approximately 7.5 nM. We propose that the tight interaction with Rab1 allows LidA to facilitate SidM/DrrA-catalyzed release of Rab1 from GDIs. Taken together, our results support a unique mechanism by which a bacterial effector protein regulates Rab1 recycling.
Author Summary
Legionella pneumophila delivers 275 validated substrates into the host cytosol by its Dot/Icm type IV secretion system. Several substrates including SidM/DrrA and LidA directly interact with the host Rab GTPases and interfere with the vesicle secretion pathway. SidM/DrrA is necessary for Rab1 recruitment, function as a Rab1 specific GDI displacement factor and guanine nucleotide exchange factor. LidA has the auxiliary activity for Rab1 recruitment, whereas it is more important for the formation of the replication vacuole compared with SidM/DrrA. LidA is predicted to be the first substrate secreted by the Dot/Icm system and is critical for maintaining the integrity of the bacterial cell. Moreover, it expresses throughout the intracellular growth phase, localizes to early secretory compartments, and interacts with several members of Rab family. Here we present the crystal structures of LidA coiled-coil domain in complex with two different states of Rab1, GDP- and GTP-bound. The GDP-bound Rab1 in the complex surprisingly has the same conformation with the GTP-bound Rab1, revealing that LidA can retain Rab1 persistently in its active state. Our structures add a new insight into the regulation of the host Rab1 membrane cycle by pathogen-secreted coiled-coil effector.
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Vimentin phosphorylation and assembly are regulated by the small GTPase Rab7a
Cogli, Laura
Progida, Cinzia
Bramato, Roberta
Bucci, Cecilia
Biochimica et Biophysica Acta
Intermediate filaments are cytoskeletal elements important for cell architecture. Recently it has been discovered that intermediate filaments are highly dynamic and that they are fundamental for organelle positioning, transport and function thus being an important regulatory component of membrane traffic. We have identified, using the yeast two-hybrid system, vimentin, a class III intermediate filament protein, as a Rab7a interacting protein. Rab7a is a member of the Rab family of small GTPases and it controls vesicular membrane traffic to late endosomes and lysosomes. In addition, Rab7a is important for maturation of phagosomes and autophagic vacuoles. We confirmed the interaction in HeLa cells by co-immunoprecipitation and pull-down experiments, and established that the interaction is direct using bacterially expressed recombinant proteins. Immunofluorescence analysis on HeLa cells indicate that Rab7a-positive vesicles sometimes overlap with vimentin filaments. Overexpression of Rab7a causes an increase in vimentin phosphorylation at different sites and causes redistribution of vimentin in the soluble fraction. Consistently, Rab7a silencing causes an increase of vimentin present in the insoluble fraction (assembled). Also, expression of Charcot–Marie–Tooth 2B-causing Rab7a mutant proteins induces vimentin phosphorylation and increases the amount of vimentin in the soluble fraction. Thus, modulation of expression levels of Rab7a wt or expression of Rab7a mutant proteins changes the assembly of vimentin and its phosphorylation state indicating that Rab7a is important for the regulation of vimentin function.
► We searched for new Rab7a interacting proteins and we found vimentin. ► We demonstrated that Rab7a interacts directly with vimentin. ► Rab7a influences vimentin's phosphorylation and soluble/insoluble ratio. ► Rab7a regulates vimentin organization and function.
GST, Glutathione-S-Transferase; HA, Hemagglutinin; EGFP, Enhanced Green Fluorescent Protein; Rab7a; Vimentin; Rab protein; Intermediate filaments; Phosphorylation; Two-hybrid
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The Trypanosome Rab-Related Proteins RabX1 and RabX2 Play No Role in IntraCellular Trafficking but May Be Involved in Fly Infectivity
Natesan, Senthil Kumar A.
Peacock, Lori
Leung, Ka Fai
Matthews, Keith R.
Gibson, Wendy
Field, Mark C.
PLoS ONE
Rab GTPases constitute the largest subgroup of the Ras superfamily and are primarily involved in vesicle targeting. The full extent of Rab family function is unexplored. Several divergent Rab-like proteins are known but few have been characterized. In Trypanosoma brucei there are sixteen Rab genes, but RabX1, RabX2 and RabX3 are divergent within canonical sequence regions. Where known, trypanosome Rab functions are broadly conserved when orthologous relationships may be robustly established, but specific functions for RabX1, X2 and X3 have yet to be determined. RabX1 and RabX2 originated via tandem duplication and subcellular localization places RabX1 at the endoplasmic reticulum, while RabX2 is at the Golgi complex, suggesting distinct functions. We wished to determine whether RabX1 and RabX2 are involved in vesicle transport or other cellular processes.
Methodology/Principal Findings
Using comparative genomics we find that RabX1 and RabX2 are restricted to trypanosomatids. Gene knockout indicates that RabX1 and RabX2 are non-essential. Simultaneous RNAi knockdown of both RabX1 and RabX2, while partial, was also non-lethal and may suggest non-redundant function, consistent with the distinct locations of the proteins. Analysis of the knockout cell lines unexpectedly failed to uncover a defect in exocytosis, endocytosis or in the morphology or location of multiple markers for the endomembrane system, suggesting that neither RabX1 nor RabX2 has a major role in intracellular transport. However, it was apparent that RabX1 and RabX2 knockout cells displayed somewhat enhanced survival within flies.
RabX1 and RabX2, two members of the trypanosome Rab subfamily, were shown to have no major detectable role in intracellular transport, despite the localization of each gene product to highly specific endomembrane compartments. These data extend the functional scope of Rab proteins in trypanosomes to include non-canonical roles in differentiation-associated processes in protozoa.
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Rab11 Regulates the Compartmentalization of Early Endosomes Required for Efficient Transport from Early Endosomes to the Trans-Golgi Network
Wilcke, Mona
Johannes, Ludger
Galli, Thierry
Mayau, Véronique
Goud, Bruno
Salamero, Jean
The Journal of Cell Biology
Several GTPases of the Rab family, known to be regulators of membrane traffic between organelles, have been described and localized to various intracellular compartments. Rab11 has previously been reported to be associated with the pericentriolar recycling compartment, post-Golgi vesicles, and the trans-Golgi network (TGN). We compared the effect of overexpression of wild-type and mutant forms of Rab11 on the different intracellular transport steps in the endocytic/degradative and the biosynthetic/exocytic pathways in HeLa cells. We also studied transport from endosomes to the Golgi apparatus using the Shiga toxin B subunit (STxB) and TGN38 as reporter molecules. Overexpression of both Rab11 wild-type (Rab11wt) and mutants altered the localization of the transferrrin receptor (TfR), internalized Tf, the STxB, and TGN38. In cells overexpressing Rab11wt and in a GTPase-deficient Rab11 mutant (Rab11Q70L), these proteins were found in vesicles showing characteristics of sorting endosomes lacking cellubrevin (Cb). In contrast, they were redistributed into an extended tubular network, together with Cb, in cells overexpressing a dominant negative mutant of Rab11 (Rab11S25N). This tubularized compartment was not accessible to Tf internalized at temperatures <20°C, suggesting that it is of recycling endosomal origin. Overexpression of Rab11wt, Rab11Q70L, and Rab11S25N also inhibited STxB and TGN38 transport from endosomes to the TGN. These results suggest that Rab11 influences endosome to TGN trafficking primarily by regulating membrane distribution inside the early endosomal pathway.
Rab11; intracellular trafficking; endosomes; Shiga toxin; TGN38
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Novel interaction of Rab13 and Rab8 with endospanins☆
Hirvonen, Mirkka J.
Büki, Kálmán G.
Sun, Yi
Mulari, Mika T.K.
Härkönen, Pirkko L.
Väänänen, Kalervo H.
FEBS Open Bio
Rab GTPases regulate vesicular traffic in eukaryotic cells by cycling between the active GTP-bound and inactive GDP-bound states. Their functions are modulated by the diverse selection of effector proteins that bind to specific Rabs in their activated state. We previously described the expression of Rab13 in bone cells. To search for novel Rab13 interaction partners, we screened a newborn rat bone marrow cDNA library for Rab13 effectors with a bacterial two-hybrid system. We found that Rab13 binds to the C-terminus of Endospanin-2, a small transmembrane protein. In addition to Rab13 also Rab8 bound to Endospanin-2, while no binding of Rab7, Rab10, Rab11 or Rab32 was observed. Rab13 and Rab8 also interacted with Endospanin-1, a close homolog of Endospanin-2. Rab13 and Endospanin-2 colocalised in perinuclear vesicular structures in Cos1 cells suggesting direct binding also in vivo. Endospanin-2 is implicated in the regulation of the cell surface growth hormone receptor (GHR), but the inhibition of Rab13 expression did not affect GHR cell surface expression. This suggests that the Rab13–Endospanin-2 interaction may have functions other than GHR regulation. In conclusion, we have identified a novel interaction for Rab13 and Rab8 with Endospanin-2 and Endospanin-1. The role of this interaction in cell physiology, however, remains to be elucidated.
▸ Rab13 and Rab8 both interact with Endospanin-2 and Endospanin-1. ▸ Rab13 and Rab8 binding to endospanins is specific; Rabs 7, 10, 11 and 32 do not bind. ▸ Rab13 binding to Endospanin-2 is nucleotide-dependent. ▸ Rab13 and Endospanin-2 colocalise in perinuclear vesicles and at the cell periphery.
Vesicle trafficking; Rab13; Rab effector; Protein interaction; Endospanin; Osteoclast; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; GFP, green fluorescent protein; GHR, growth hormone receptor; GST, glutathione-S-transferase; HA, human influenza hemagglutinin; MBP, maltose binding protein; OB-R, leptin receptor; VPS55, vacuolar protein sorting 55.
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An Overexpression Screen of Toxoplasma gondii Rab-GTPases Reveals Distinct Transport Routes to the Micronemes
Kremer, Katrin
Kamin, Dirk
Rittweger, Eva
Wilkes, Jonathan
Flammer, Halley
Mahler, Sabine
Heng, Joanne
Tonkin, Christopher J.
Langsley, Gordon
Hell, Stefan W.
Carruthers, Vernon B.
Ferguson, David J. P.
Meissner, Markus
PLoS Pathogens
The basic organisation of the endomembrane system is conserved in all eukaryotes and comparative genome analyses provides compelling evidence that the endomembrane system of the last common eukaryotic ancestor (LCEA) is complex with many genes required for regulated traffic being present. Although apicomplexan parasites, causative agents of severe human and animal diseases, appear to have only a basic set of trafficking factors such as Rab-GTPases, they evolved unique secretory organelles (micronemes, rhoptries and dense granules) that are sequentially secreted during invasion of the host cell. In order to define the secretory pathway of apicomplexans, we performed an overexpression screen of Rabs in Toxoplasma gondii and identified Rab5A and Rab5C as important regulators of traffic to micronemes and rhoptries. Intriguingly, we found that not all microneme proteins traffic depends on functional Rab5A and Rab5C, indicating the existence of redundant microneme targeting pathways. Using two-colour super-resolution stimulated emission depletion (STED) we verified distinct localisations of independent microneme proteins and demonstrate that micronemal organelles are organised in distinct subsets or subcompartments. Our results suggest that apicomplexan parasites modify classical regulators of the endocytic system to carryout essential parasite-specific roles in the biogenesis of their unique secretory organelles.
Author Summary
Eukaryotic cells evolved a highly complex endomembrane system, consisting of secretory and endocytic organelles. In the case of apicomplexan parasites unique secretory organelles have evolved that are essential for the invasion of the host cell. Surprisingly these protozoans show a paucity of trafficking factors, such as Rabs and it appears that they lost several factors involved in endocytosis. Here, we demonstrate that Rab5A and Rab5C, normally involved in endocytic uptake, actually regulate secretion in Toxoplasma gondii, since functional ablation of Rab5A or Rab5C results in aberrant transport of proteins to specialised secretory organelles called micronemes and rhoptries. Furthermore, we demonstrate that independent transport routes to micronemes exist indicating that apicomplexans have remodelled Rab5-mediated vesicular traffic into a secretory system that is essential for host cell invasion.
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Construction of a Plasmodium falciparum Rab-interactome identifies CK1 and PKA as Rab-effector kinases in malaria parasites
Rached, Fathia Ben
Ndjembo-Ezougou, Carinne
Chandran, Syama
Talabani, Hana
Yera, Hélène
Dandavate, Vrushali
Bourdoncle, Pierre
Meissner, Markus
Tatu, Utpal
Langsley, Gordon
Biology of the Cell
Background information
The pathology causing stages of the human malaria parasite Plasmodium falciparum reside within red blood cells that are devoid of any regulated transport system. The parasite, therefore, is entirely responsible for mediating vesicular transport within itself and in the infected erythrocyte cytoplasm, and it does so in part via its family of 11 Rab GTPases. Putative functions have been ascribed to Plasmodium Rabs due to their homology with Rabs of yeast, particularly with Saccharomyces that has an equivalent number of rab/ypt genes and where analyses of Ypt function is well characterized.
Rabs are important regulators of vesicular traffic due to their capacity to recruit specific effectors. In order to identify P. falciparum Rab (PfRab) effectors, we first built a Ypt-interactome by exploiting genetic and physical binding data available at the Saccharomyces genome database (SGD). We then constructed a PfRab-interactome using putative parasite Rab-effectors identified by homology to Ypt-effectors. We demonstrate its potential by wet-bench testing three predictions; that casein kinase-1 (PfCK1) is a specific Rab5B interacting protein and that the catalytic subunit of cAMP-dependent protein kinase A (PfPKA-C) is a PfRab5A and PfRab7 effector.
The establishment of a shared set of physical Ypt/PfRab-effector proteins sheds light on a core set Plasmodium Rab-interactants shared with yeast. The PfRab-interactome should benefit vesicular trafficking studies in malaria parasites. The recruitment of PfCK1 to PfRab5B+ and PfPKA-C to PfRab5A+ and PfRab7+ vesicles, respectively, suggests that PfRab-recruited kinases potentially play a role in early and late endosome function in malaria parasites.
Interactom; Kinase; Plasmodium; Rab; Yeast; Ypt
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Rab11-FIP2 influences multiple components of the endosomal system in polarized MDCK cells
Ducharme, Nicole A
Ham, Amy-Joan L
Lapierre, Lynne A
Goldenring, James R
Cellular Logistics
The Rab11 Family Interacting Proteins (Rab11-FIPs) are hypothesized to regulate sequential steps in the apical recycling and transcytotic pathways of polarized epithelial cells. Previous studies have suggested that Rab11-FIP proteins assemble into multi-protein complexes regulating plasma membrane recycling. Rab11-FIP2 interacts with both myosin Vb and Rab11. Recent investigations have noted that that Rab11-FIP2 mutants [Rab11-FIP2(129–512), also designated Rab11-FIP2(ΔC2) and Rab11-FIP2(S229A, R413G), also designated Rab11-FIP2(SARG)], are potent inhibitors of transcytosis in polarized MDCK cells. Interestingly, Rab11-FIP2(ΔC2), but not Rab11-FIP2(SARG), also altered the morphology of the EEA-1 positive early endosomal compartment. These findings suggested that Rab11-FIP2 mutants could differentiate different points along the recycling pathway. We therefore sought to investigate whether Rab11-FIP2 is a general regulator of the early endosomal system. Both Rab11-FIP2 mutants altered the localization and co-localized with dynein heavy chain. In contrast, both clathrin heavy chain and AP-1 accumulated with membranes containing Rab11-FIP2(SARG), but not with Rab11-FIP2(ΔC2). Expression of Rab11-FIP2(ΔC2), but not Rab11-FIP2(SARG), caused clustering of early endosomal markers Rab5b, Epsin 4 and IQGAP1, around a collapsed Rab11-FIP2 containing membranous cisternum. Interestingly, neither Rab11-FIP2 mutant had any effect on the distribution of Rab5a, a classical early endosome marker. The results support the view that Rab11-FIP2 may influence microtubule-dependent centripetal movement of subsets of early endosomes as well as processing through the common and recycling endosomal systems.
Rab11-FIP2; Rab11; trafficking; apical recycling; endosome; MDCK cells; clathrin; dynein; Rab5; epsin
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The Glyceraldehyde-3-Phosphate Dehydrogenase and the Small GTPase Rab 2 Are Crucial for Brucella Replication
Fugier, Emilie
Salcedo, Suzana P.
de Chastellier, Chantal
Pophillat, Matthieu
Muller, Alexandre
Arce-Gorvel, Vilma
Fourquet, Patrick
Gorvel, Jean-Pierre
PLoS Pathogens
The intracellular pathogen Brucella abortus survives and replicates inside host cells within an endoplasmic reticulum (ER)-derived replicative organelle named the "Brucella-containing vacuole" (BCV). Here, we developed a subcellular fractionation method to isolate BCVs and characterize for the first time the protein composition of its replicative niche. After identification of BCV membrane proteins by 2 dimensional (2D) gel electrophoresis and mass spectrometry, we focused on two eukaryotic proteins: the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the small GTPase Rab 2 recruited to the vacuolar membrane of Brucella. These proteins were previously described to localize on vesicular and tubular clusters (VTC) and to regulate the VTC membrane traffic between the endoplasmic reticulum (ER) and the Golgi. Inhibition of either GAPDH or Rab 2 expression by small interfering RNA strongly inhibited B. abortus replication. Consistent with this result, inhibition of other partners of GAPDH and Rab 2, such as COPI and PKC ι, reduced B. abortus replication. Furthermore, blockage of Rab 2 GTPase in a GDP-locked form also inhibited B. abortus replication. Bacteria did not fuse with the ER and instead remained in lysosomal-associated membrane vacuoles. These results reveal an essential role for GAPDH and the small GTPase Rab 2 in B. abortus virulence within host cells.
Author Summary
A key determinant for intracellular pathogenic bacteria to ensure their virulence within host cells is their ability to bypass the endocytic pathway and to reach a safe replication niche. Brucella bacteria reach the endoplasmic reticulum (ER) to create their replicating niche called the Brucella-containing vacuole (BCV). The ER is a suitable strategic place for pathogenic Brucella. Bacteria can be hidden from host cell defences to persist within the host, and can take advantage of the membrane reservoir delivered by the ER to replicate. Interactions between BCV and the ER lead to the presence of ER proteins on the BCV membrane. Currently, no other proteins (eukaryotic or prokaryotic) have yet been associated with the BCV membrane. Here we show that non-ER related proteins are also present on the BCV membrane, in particular, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the small GTPase Rab 2 known to be located on secretory vesicles that traffic between the ER and the Golgi apparatus. GAPDH and the small GTPase Rab 2 are involved in Brucella replication at late post-infection. Similarly, integrity of secretory vesicle trafficking is also necessary for Brucella replication. Here, we show that recruitment of the two eukaryotic proteins GAPDH and Rab 2 on BCV membranes is necessary for the establishment of the replicative niche by sustaining interactions between the ER and secretory membrane vesicles.
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Rab41 Is a Novel Regulator of Golgi Apparatus Organization That Is Needed for ER-To-Golgi Trafficking and Cell Growth
Liu, Shijie
Hunt, Lauren
Storrie, Brian
PLoS ONE
The 60+ members of the mammalian Rab protein family group into subfamilies postulated to share common functionality. The Rab VI subfamily contains 5 Rab proteins, Rab6a/a', Rab6b, Rab6c and Rab41. High-level knockdown of Rab6a/a' has little effect on the tightly organized Golgi ribbon in HeLa cells as seen by fluorescence microscopy. In striking contrast, we found Rab41 was strongly required for normal Golgi ribbon organization.
Treatment of HeLa cells with Rab41 siRNAs scattered the Golgi ribbon into clustered, punctate Golgi elements. Overexpression of GDP-locked Rab41, but not wild type or GTP-locked Rab41, produced a similar Golgi phenotype. By electron microscopy, Rab41 depletion produced short, isolated Golgi stacks. Golgi-associated vesicles accumulated. At low expression levels, wild type and GTP-locked Rab41 showed little concentration in the Golgi region, but puncta were observed and most were in ruffled regions at the cell periphery. There was 25% co-localization of GTP-locked Rab41 with the ER marker, Sec61p. GDP-locked Rab41, as expected, displayed an entirely diffuse cytoplasmic distribution. Depletion of Rab41 or overexpression of GDP-locked Rab41 partially inhibited ER-to-Golgi transport of VSV-G protein. However, Rab41 knockdown had little, if any, effect on endosome-to-Golgi transport of SLTB. Additionally, after a 2-day delay, treatment with Rab41 siRNA inhibited cell growth, while overexpression of GDP-locked Rab41, but not wild type or GTP-locked Rab41, produced a rapid, progressive cell loss. In double knockdown experiments with Rab6, the Golgi ribbon was fragmented, a result consistent with Rab41 and Rab6 acting in parallel.
We provide the first evidence for distinctive Rab41 effects on Golgi organization, ER-to-Golgi trafficking and cell growth. When combined with the evidence that Rab6a/a' and Rab6b have diverse roles in Golgi function, while Rab6c regulates mitotic function, our data indicate that Rab VI subfamily members, although related by homology and structure, share limited functional conservation.
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Regulation of Synaptic Transmission by RAB-3 and RAB-27 in Caenorhabditis elegans
Mahoney, Timothy R.
Liu, Qiang
Itoh, Takashi
Luo, Shuo
Hadwiger, Gayla
Vincent, Rose
Wang, Zhao-Wen
Fukuda, Mitsunori
Nonet, Michael L.
Molecular Biology of the Cell
Rab small GTPases are involved in the transport of vesicles between different membranous organelles. RAB-3 is an exocytic Rab that plays a modulatory role in synaptic transmission. Unexpectedly, mutations in the Caenorhabditis elegans RAB-3 exchange factor homologue, aex-3, cause a more severe synaptic transmission defect as well as a defecation defect not seen in rab-3 mutants. We hypothesized that AEX-3 may regulate a second Rab that regulates these processes with RAB-3. We found that AEX-3 regulates another exocytic Rab, RAB-27. Here, we show that C. elegans RAB-27 is localized to synapse-rich regions pan-neuronally and is also expressed in intestinal cells. We identify aex-6 alleles as containing mutations in rab-27. Interestingly, aex-6 mutants exhibit the same defecation defect as aex-3 mutants. aex-6; rab-3 double mutants have behavioral and pharmacological defects similar to aex-3 mutants. In addition, we demonstrate that RBF-1 (rabphilin) is an effector of RAB-27. Therefore, our work demonstrates that AEX-3 regulates both RAB-3 and RAB-27, that both RAB-3 and RAB-27 regulate synaptic transmission, and that RAB-27 potentially acts through its effector RBF-1 to promote soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) function.
Related Articles
A Rab1 mutant affecting guanine nucleotide exchange promotes disassembly of the Golgi apparatus
The Journal of Cell Biology
The Golgi apparatus is a dynamic organelle whose structure is sensitive to vesicular traffic and to cell cycle control. We have examined the potential role for rab1a, a GTPase previously associated with ER to Golgi and intra-Golgi transport, in the formation and maintenance of Golgi structure. Bacterially expressed, recombinant rab1a protein was microinjected into rat embryonic fibroblasts, followed by analysis of Golgi morphology by fluorescence and electron microscopy. Three recombinant proteins were tested: wild-type rab, mutant rab1a(S25N), a constitutively GDP-bound form (Nuoffer, C., H. W. Davidson, J. Matteson, J. Meinkoth, and W. E. Balch, 1994. J. Cell Biol. 125: 225- 237), and mutant rab1a(N124I) defective in guanine nucleotide binding. Microinjection of wild-type rab1a protein or a variety of negative controls (injection buffer alone or activated ras protein) did not affect the appearance of the Golgi, as visualized by immunofluorescence of alpha-mannosidase II (Man II), used as a Golgi marker. In contrast, microinjection of the mutant forms promoted the disassembly of the Golgi stacks into dispersed vesicular structures visualized by immunofluorescence. When S25N-injected cells were analyzed by EM after immunoperoxidase labeling, Man II was found in isolated ministacks and large vesicular elements that were often surrounded by numerous smaller unlabeled vesicles resembling carrier vesicles. Golgi disassembly caused by rab1a mutants differs from BFA-induced disruption, since beta- COP remains membrane associated, and Man II does not redistribute to the ER. BFA can still cause these residual Golgi elements to fuse and disperse, albeit at a slower rate. Moreover, BFA recovery is incomplete in the presence of rab1 mutants or GTP gamma S. We conclude that GTP exchange and hydrolysis by GTPases, specifically rab1a, are required to form and maintain normal Golgi stacks. The similarity of Golgi disassembly seen with rab1a mutants to that occurring during mitosis, may point to a molecular basis involving rab1a for fragmentation of the Golgi apparatus during cell division.
Related Articles
AMPylation Is Critical for Rab1 Localization to Vacuoles Containing Legionella pneumophila
Hardiman, Camille A.
Roy, Craig R.
Legionella pneumophila is an intracellular pathogen that resides within a membrane-bound compartment that is derived from vesicles exiting the endoplasmic reticulum (ER). To create this compartment, these bacteria use a type IV secretion system to deliver effector proteins that subvert host cell functions. Several Legionella effector proteins modulate the function of the host protein Rab1, which is a GTPase that is recruited to the Legionella-containing vacuole (LCV). Here, we examined which of the Rab1-directed enzymatic activities displayed by Legionella effectors are important for localizing the Rab1 protein to the LCV membrane. The guanine nucleotide exchange factor (GEF) domain in the effector protein DrrA (SidM) was essential for Rab1 recruitment to the LCV and Rab1 AMPylation by the nucleotidyltransferase domain in DrrA was important for Rab1 retention. Legionella organisms producing mutant DrrA proteins that were severely attenuated for GEF activity in vitro retained the ability to localize Rab1 to the LCV. Rab1 localization to the LCV mediated by these GEF-defective mutants required AMPylation. Importantly, we found that efficient localization of Rab1 to the LCV occurred when Rab1 GEF activity and Rab1 AMPylation activity were provided by separate proteins. Rab1 phosphocholination (PCylation) by the effector protein AnkX, however, was unable to substitute for Rab1 AMPylation. Lastly, the defect in Rab1 localization to the LCV in AMPylation-deficient strains of Legionella was partially suppressed if the GTPase-activating protein (GAP) LepB was eliminated. Thus, our data indicate that AMPylation of Rab1 is an effective strategy to maintain this GTPase on the LCV membrane.
Activities that enable the intracellular pathogen Legionella pneumophila to subvert the function of the host protein Rab1 were investigated. Our data show that a posttranslational modification called AMPylation is critical for maintaining a pool of Rab1 on the LCV membrane. AMPylation of Rab1 led to the accumulation of GTP-bound Rab1 on the LCV membrane by protecting the protein from inactivation by GAPs. Importantly, PCylation of Rab1 by the Legionella effector protein AnkX was neither necessary nor sufficient to maintain Rab1 on the LCV, indicating that AMPylation and PCylation represent functionally distinct activities. We conclude that modification of Rab1 by AMPylation is an effective strategy to spatially and temporally regulate the function of this GTPase on a membrane-bound organelle.
Related Articles
A Rab8-specific GDP/GTP Exchange Factor Is Involved in Actin Remodeling and Polarized Membrane Transport
Hattula, Katarina
Furuhjelm, Johanna
Arffman, Airi
Peränen, Johan
Molecular Biology of the Cell
The mechanisms mediating polarized delivery of vesicles to cell surface domains are poorly understood in animal cells. We have previously shown that expression of Rab8 promotes the formation of new cell surface domains through reorganization of actin and microtubules. To unravel the function of Rab8, we used the yeast two-hybrid system to search for potential Rab8-specific activators. We identified a coil-coiled protein (Rabin8), homologous to the rat Rabin3 that stimulated nucleotide exchange on Rab8 but not on Rab3A and Rab5. Furthermore, we show that rat Rabin3 has exchange activity on Rab8 but not on Rab3A, supporting the view that rat Rabin3 is the rat equivalent of human Rabin8. Rabin8 localized to the cortical actin and expression of Rabin8 resulted in remodeling of actin and the formation of polarized cell surface domains. Activation of PKC by phorbol esters enhanced translocation of both Rabin8 and Rab8-specific vesicles to the outer edge of lamellipodial structures. Moreover, coexpression of Rabin8 with dominant negative Rab8 (T22N) redistributes Rabin8 from cortical actin to Rab8-specific vesicles and promotes their polarized transport to cell protrusions. The C-terminal region of Rabin8 plays an essential role in this transport. We propose that Rabin8 is a Rab8-specific activator that is connected to processes that mediate polarized membrane traffic to dynamic cell surface structures.
Related Articles
Overexpression of a Mutant Form of EhRabA, a Unique Rab GTPase of Entamoeba histolytica, Alters Endoplasmic Reticulum Morphology and Localization of the Gal/GalNAc Adherence Lectin▿
Welter, B. H.
Temesvari, L. A.
Eukaryotic Cell
Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. Vesicle trafficking events, such as phagocytosis and delivery of plasma membrane proteins, have been implicated in pathogenicity. Rab GTPases are proteins whose primary function is to regulate vesicle trafficking; therefore, understanding the function of Rabs in this organism may provide insight into virulence. E. histolytica possesses a number of unique Rabs that exhibit limited homology to host Rabs. In this study we examined the function of one such Rab, EhRabA, by characterizing a mutant overexpressing a constitutively GTP-bound version of the protein. Overexpression of mutant EhRabA resulted in decreased adhesion to and phagocytosis of human red blood cells and in the appearance of large tubular organelles that could be stained with endoplasmic reticulum (ER)-specific but not Golgi complex-specific antibodies. Consistent with the adhesion defect, two subunits of a cell surface adhesin, the galactose/N-acetylgalactosamine lectin, were mislocalized to the novel organelle. A cysteine protease, EhCP2, was also localized to the ER-like compartment in the mutant; however, the localization of two additional cell surface proteins, Igl and SREHP, remained unchanged in the mutant. The phenotype of the mutant could be recapitulated by treatment with brefeldin A, a cellular toxin that disrupts ER-to-Golgi apparatus vesicle traffic. This suggests that EhRabA influences vesicle trafficking pathways that are also sensitive to brefeldin A. Together, the data indicate that EhRabA directly or indirectly influences the morphology of secretory organelles and regulates trafficking of a subset of secretory proteins in E. histolytica.
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Rab6 and Rab11 Regulate Chlamydia trachomatis Development and Golgin-84-Dependent Golgi Fragmentation
Lipinski, Anette Rejman
Heymann, Julia
Meissner, Charlotte
Karlas, Alexander
Brinkmann, Volker
Meyer, Thomas F.
Heuer, Dagmar
PLoS Pathogens
Many intracellular pathogens that replicate in special membrane bound compartments exploit cellular trafficking pathways by targeting small GTPases, including Rab proteins. Members of the Chlamydiaceae recruit a subset of Rab proteins to their inclusions, but the significance of these interactions is uncertain. Using RNA interference, we identified Rab6 and Rab11 as important regulators of Chlamydia infections. Depletion of either Rab6 or Rab11, but not the other Rab proteins tested, decreased the formation of infectious particles. We further examined the interplay between these Rab proteins and the Golgi matrix components golgin-84 and p115 with regard to Chlamydia-induced Golgi fragmentation. Silencing of the Rab proteins blocked Chlamydia-induced and golgin-84 knockdown-stimulated Golgi disruption, whereas Golgi fragmentation was unaffected in p115 depleted cells. Interestingly, p115-induced Golgi fragmentation could rescue Chlamydia propagation in Rab6 and Rab11 knockdown cells. Furthermore, transport of nutrients to Chlamydia, as monitored by BODIPY-Ceramide, was inhibited by Rab6 and Rab11 knockdown. Taken together, our results demonstrate that Rab6 and Rab11 are key regulators of Golgi stability and further support the notion that Chlamydia subverts Golgi structure to enhance its intracellular development.
Author Summary
Chlamydia trachomatis is the leading cause of sexually transmitted disease and preventable blindness in humans. These bacteria survive and replicate within a membrane bound niche inside host cells, termed the inclusion. The inclusion expands as Chlamydia replicate, likely fuelled by the interception of cellular trafficking pathways to acquire essential nutrients. Certain host proteins, the Rab GTPases, have previously been observed at inclusion membranes, but the function of this host–pathogen interaction remains unknown. Here, we show distinct Rab proteins are important for the outcome of an infection. Depletion of Rab6 and Rab11 by RNA interference inhibited Chlamydia-induced fragmentation of the Golgi apparatus (GA), a key organelle functioning in the processing and packaging of cell synthesized macromolecules, accompanied by a decrease in bacterial progeny and a reduction in lipid transport to the inclusion. Our investigation of the cross-talk between Chlamydia and a host cell trafficking pathway not only further strengthens our group's previous finding that Chlamydia subverts GA structure to enhance replication, but has identified Rab6 and Rab11 as key regulators of GA stability. In the wider context, our results also raise the provocative notion of a novel anti-chlamydial treatment based on interference with GA function.
Related Articles
Rab8, a small GTPase involved in vesicular traffic between the TGN and the basolateral plasma membrane
The Journal of Cell Biology
Small GTP-binding proteins of the rab family have been implicated as regulators of membrane traffic along the biosynthetic and endocytic pathways in eukaryotic cells. We have investigated the localization and function of rab8, closely related to the yeast YPT1/SEC4 gene products. Confocal immunofluorescence microscopy and immunoelectron microscopy on filter-grown MDCK cells demonstrated that, rab8 was localized to the Golgi region, vesicular structures, and to the basolateral plasma membrane. Two-dimensional gel electrophoresis showed that rab8p was highly enriched in immuno-isolated basolateral vesicles carrying vesicular stomatitis virus-glycoprotein (VSV-G) but was absent from vesicles transporting the hemagglutinin protein (HA) of influenza virus to the apical cell surface. Using a cytosol dependent in vitro transport assay in permeabilized MDCK cells we studied the functional role of rab8 in biosynthetic membrane traffic. Transport of VSV-G from the TGN to the basolateral plasma membrane was found to be significantly inhibited by a peptide derived from the hypervariable COOH-terminal region of rab8, while transport of the influenza HA from the TGN to the apical surface and ER to Golgi transport were unaffected. We conclude that rab8 plays a role in membrane traffic from the TGN to the basolateral plasma membrane in MDCK cells.
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Rab18 Facilitates Dengue Virus Infection by Targeting Fatty Acid Synthase to Sites of Viral Replication
Tang, Wei-Chun
Lin, Ren-Jye
Liao, Ching-Len
Lin, Yi-Ling
Journal of Virology
Positive-sense RNA viruses, such as dengue virus (DENV), hijack the intracellular membrane machinery for their own replication. The Rab18 protein, a member of the Rab GTPase family, key regulators of membrane trafficking, is located on the organelles involved in DENV infection, such as the endoplasmic reticulum (ER) and lipid droplets (LDs). In this study, we addressed the potential involvement of Rab18 in DENV infection by using cells overexpressing the wild-type, GTP-bound active form, or GDP-bound inactive form of Rab18 and cells with Rab18 knockdown. DENV replication, measured by viral protein, viral RNA, and viral progeny production, as well as LD induction, was reduced in cells with inactive Rab18 and in cells deprived of Rab18 expression, suggesting a positive role of Rab18 in the DENV life cycle. Interestingly, the interaction of fatty acid synthase (FASN), a key lipogenic enzyme in lipid biosynthesis, with DENV NS3 protein relied on the conversion of the GDP-bound to the GTP-bound form of Rab18. Furthermore, the targeting of FASN to sites participating in DENV infection, such as the ER and LDs, depends on functional Rab18. Thus, Rab18-mediated membrane trafficking of FASN and NS3 facilitates DENV replication, probably by ensuring a sufficient and coordinated lipid supply for membrane proliferation and arrangement.
IMPORTANCE Infection by dengue virus (DENV), an important mosquito-borne virus threatening ∼40% of the world's population, can cause mild dengue fever or severe dengue hemorrhagic fever and dengue shock syndrome. The pathogenesis mechanisms of DENV-related diseases are not clear, but high viral replication is believed to be a risk factor for the severe form of DENV infection. Thus, understanding the detailed mechanism of DENV replication might help address this devastating virus. Here, we found that Rab18, a small GTPase involved in vesicle trafficking and located in the endoplasmic reticulum network and on the surfaces of lipid droplets, positively regulates DENV replication. The functional machinery of Rab18 is required to recruit the enzyme fatty acid synthase to sites of DENV replication and to interact with DENV NS3 protein to promote fatty acid biosynthesis. Thus, DENV usurps Rab18 to facilitate its own replication.
Related Articles
Evidence for a recycling role for Rab7 in regulating a late step in endocytosis and in retention of lysosomal enzymes in Dictyostelium discoideum.
Buczynski, G
Bush, J
Zhang, L
Rodriguez-Paris, J
Cardelli, J
Molecular Biology of the Cell
The mammalian small molecular weight GTPase Rab7 (Ypt7 in yeast) has been implicated in regulating membrane traffic at postinternalization steps along the endosomal pathway. A cDNA encoding a protein 85% identical at the amino acid level to mammalian Rab7 has been cloned from Dictyostelium discoideum. Subcellular fractionation and immunofluorescence microscopy indicated that Rab7 was enriched in lysosomes, postlysosomes, and maturing phagosomes. Cell lines were generated that overexposed Rab7 wild-type (WT), Rab7 Q67L (constitutively active form), and Rab7 T22N (dominant negative form) proteins. The Rab7 T22N cell line internalized fluid phase markers and latex beads (phagocytosis) at one-third the rate of control cells, whereas Rab7 WT and Rab7 Q67L cell lines were normal in uptake rates but exocytosed fluid phase faster than control cells. In contrast, fluid phase markers resided in acidic compartments for longer periods of time and were more slowly exocytosed from Rab7 T22N cells as compared with control cells. Light microscopy indicated that Rab7-expressing cell lines contained morphologically altered endosomal compartments. Compared with control cells, Rab7 WT- and Rab7 Q67L-expressing cells contained a reduced number of vesicles, the size of postlysosomes (> 2.5 microns) and an increased number of smaller vesicles, many of which were nonacidic; in control cells, > 90% of the smaller vesicles were acidic. In contrast, Rab7 T22N cells contained an increased proportion of large acidic vesicles relative to nonacidic vesicles. Radiolabel pulse-chase experiments indicated that all of the cell lines processed and targeted lysosomal alpha-mannosidase normally, indicating the lack of a significant role for Rab7 in the targeting pathway; however, retention of mature lysosomal hydrolases was affected in Rab7 WT and Rab7 T22N cell lines. Contrary to the results observed for the fluid phase efflux experiments, Rab7 T22N cells oversecreted alpha-mannosidase, whereas Rab7 WT cells retained this hydrolase as compared with control cells. These data support a model that Rab7 may regulate retrograde transport of lysosomal enzymes and the V-type H(+)-ATPase from postlysosomes to lysosomes coupled with the efficient release of fluid phase from cells.
Related Articles
From Sorting Endosomes to Exocytosis: Association of Rab4 and Rab11 GTPases with the Fc Receptor, FcRn, during RecyclingV⃞
Ward, E. Sally
Martinez, Cruz
Vaccaro, Carlos
Zhou, Jinchun
Tang, Qing
Ober, Raimund J.
Molecular Biology of the Cell
A longstanding question in cell biology is how is the routing of intracellular organelles within cells regulated? Although data support the involvement of Rab4 and Rab11 GTPases in the recycling pathway, the function of Rab11 in particular is uncertain. Here we have analyzed the association of these two Rab GTPases with the Fc receptor, FcRn, during intracellular trafficking. This Fc receptor is both functionally and structurally distinct from the classical Fcγ receptors and transports immunoglobulin G (IgG) within cells. FcRn is therefore a recycling receptor that sorts bound IgG from unbound IgG in sorting endosomes. In the current study we have used dual color total internal reflection fluorescence microscopy (TIRFM) and wide-field imaging of live cells to analyze the events in human endothelial cells that are involved in the trafficking of FcRn positive (FcRn+) recycling compartments from sorting endosomes to exocytic sites at the plasma membrane. Our data are consistent with the following model for this pathway: FcRn leaves sorting endosomes in Rab4+Rab11+ or Rab11+ compartments. For Rab4+Rab11+ compartments, Rab4 depletion occurs by segregation of the two Rab proteins into discrete domains that can separate. The Rab11+FcRn+ vesicle or tubule subsequently fuses with the plasma membrane in an exocytic event. In contrast to Rab11, Rab4 is not involved in exocytosis.
Related Articles
Rab11A-Controlled Assembly of the Inner Membrane Complex Is Required for Completion of Apicomplexan Cytokinesis
Agop-Nersesian, Carolina
Naissant, Bernina
Rached, Fathia Ben
Rauch, Manuel
Kretzschmar, Angelika
Thiberge, Sabine
Menard, Robert
Ferguson, David J. P.
Meissner, Markus
Langsley, Gordon
PLoS Pathogens
The final step during cell division is the separation of daughter cells, a process that requires the coordinated delivery and assembly of new membrane to the cleavage furrow. While most eukaryotic cells replicate by binary fission, replication of apicomplexan parasites involves the assembly of daughters (merozoites/tachyzoites) within the mother cell, using the so-called Inner Membrane Complex (IMC) as a scaffold. After de novo synthesis of the IMC and biogenesis or segregation of new organelles, daughters bud out of the mother cell to invade new host cells. Here, we demonstrate that the final step in parasite cell division involves delivery of new plasma membrane to the daughter cells, in a process requiring functional Rab11A. Importantly, Rab11A can be found in association with Myosin-Tail-Interacting-Protein (MTIP), also known as Myosin Light Chain 1 (MLC1), a member of a 4-protein motor complex called the glideosome that is known to be crucial for parasite invasion of host cells. Ablation of Rab11A function results in daughter parasites having an incompletely formed IMC that leads to a block at a late stage of cell division. A similar defect is observed upon inducible expression of a myosin A tail-only mutant. We propose a model where Rab11A-mediated vesicular traffic driven by an MTIP-Myosin motor is necessary for IMC maturation and to deliver new plasma membrane to daughter cells in order to complete cell division.
Author Summary
Apicomplexan parasites are unusual in that they replicate by assembling daughter parasites within the mother cell. This involves the ordered assembly of an Inner Membrane Complex (IMC), a scaffold consisting of flattened membrane cisternae and a subpellicular network made up of microtubules and scaffold proteins. The IMC begins to form at the onset of replication, but its maturation occurs at the final stage of cytokinesis (the last step during cell division) upon the addition of motor (glideosome) components such as GAP45 (Glideosome Associated Protein), Myosin A (MyoA), and Myosin-Tail-Interacting-Protein (MTIP, also known as Myosin Light Chain 1) that are necessary to drive the gliding motility required for parasite invasion. We demonstrate that Rab11A regulates not only delivery of new plasmamembrane to daughter cells, but, importantly, also correct IMC formation. We show that Rab11A physically interacts with MTIP/MLC1, implicating unconventional myosin(s) in both cytokinesis and IMC maturation, and, consistently, overexpression of a MyoA tail-only mutant generates a default similar to that which we observe upon Rab11A ablation. We propose a model where Rab11A-mediated vesicular traffic is required for the delivery of new plasma membrane to daughter cells and for the maturation of the IMC in order to complete cell division.
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It's just a little confusing, is all.
The good news is the republican convention is over. The bad news is if I can escape this police car how do I anull a marriage??
Title of this one pretty much says it all. If it's what you're looking for, then uh, look no further
There is literally nothing sexier than eye contact.
A dandy fop dog gets a rhino piss party and ant enema in the Jungle. Weldcome to Hell.
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The UAB National Alumni Society hosted the 7th annual Uncork Education on Sunday, November 8.
This popular wine and beer tasting event had a silent auction for guests and benefited UAB student scholarships. This year, the generosity of those who attended Uncork Education allowed the NAS to raise more than $48,500 to go towards UAB student scholarships, gifts, and grants.
In 2014-2015, the UAB National Alumni Society had the opportunity to provide approximately $775,000 in scholarships, gifts, and grants back to the University. The success of Uncork Education and the support of our sponsors and guests allow us to continue to give money to deserving students and sincerely express our gratitude to everyone for their continued support as we continue to closer to reaching our goal of providing more than $1 million in scholarships by 2020.
The evening gave our guests a chance to taste wines from around the globe from International Wines and local craft beer from Cahaba Brewing. They also had an opportunity to purchase a key that may open a fully stocked bar that included an assortment of premium liquor. The 2015 Blazer Wine label design was also unveiled and bottles were available to be purchased. Eric Essix provided live music.


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Experienced Pediatricians in McHenry County
Centegra Physician Care pediatricians and family practice doctors provide the leading pediatrics care in the McHenry County region.The doctors and nurse practitioners of Centegra Physician Care provide complete care for children from birth to 18 years of age.
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this is a wordpress blog that has been set up simply for money. :) -- ok, not totally for money, but you get the idea.
Given my wierd nature, I had to completely hide the admin and includes area, and I have the script built that will actually go out and grab data from CL for the 'big love' category, but I'm hesitant to use it, since I actually get a laugh out of reading some of the ads.


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Breast Biopsy Options
Dear OncoLink "Ask The Experts,"
What are the different ways to get a sample of my breast lump?
Lawrence J. Solin, MD, FACR, Professor of Radiation Oncology at the University of Pennsylvania, responds:
When an abnormality is found on screening mammography, the next step is to determine whether or not the abnormality is malignant (cancer). There are a number of different ways to obtain a sample of tissue for the pathologist to make this determination. Often, the physician cannot feel such abnormalities, and so, the biopsy must be guided by some form of radiologic imaging. Obtaining a small sample of tissue (core biopsy or aspiration cytology) is sometimes sufficient to determine if the abnormality is benign or malignant, and allows the treating doctors to better plan the most appropriate treatment. Doing a lumpectomy may be necessary when more tissue needs to be obtained or when there is a likely or known cancer in the breast.
Often, there are different ways that can be used to evaluate a breast abnormality, and no single way is best for all situations. Physician judgment on how best to do such an evaluation is necessary to design the approach for the individual patient.
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