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We'd be surprised if Steadfast Group Limited (ASX:SDF) shareholders haven't noticed that the Co-Founder, Robert Kelly, recently sold AU$637k worth of stock at AU$3.64 per share. On the bright side, that sale was only 4.9% of their holding, so we doubt it's very meaningful, on its own.
Check out our latest analysis for Steadfast Group
The Last 12 Months Of Insider Transactions At Steadfast Group
Notably, that recent sale by Co-Founder Robert Kelly was not the only time they sold Steadfast Group shares this year. They previously made an even bigger sale of -AU$3.5m worth of shares at a price of AU$3.07 per share. That means that an insider was selling shares at slightly below the current price (AU$3.57). We generally consider it a negative if insiders have been selling, especially if they did so below the current price, because it implies that they considered a lower price to be reasonable. However, while insider selling is sometimes discouraging, it's only a weak signal. This single sale was just 25% of Robert Kelly's stake.
Over the last year, we can see that insiders have bought 114250 shares worth AU$356k. But they sold 2605567 for AU$8.5m. Over the last year we saw more insider selling of Steadfast Group shares, than buying. You can see the insider transactions (by individuals) over the last year depicted in the chart below. If you want to know exactly who sold, for how much, and when, simply click on the graph below!
I will like Steadfast Group better if I see some big insider buys. While we wait, check out this free list of growing companies with considerable, recent, insider buying.
Insider Ownership
For a common shareholder, it is worth checking how many shares are held by company insiders. We usually like to see fairly high levels of insider ownership. Steadfast Group insiders own about AU$42m worth of shares. That equates to 1.4% of the company. This level of insider ownership is good but just short of being particularly stand-out. It certainly does suggest a reasonable degree of alignment.
What Might The Insider Transactions At Steadfast Group Tell Us?
Insiders haven't bought Steadfast Group stock in the last three months, but there was some selling. Despite some insider buying, the longer term picture doesn't make us feel much more positive. But it is good to see that Steadfast Group is growing earnings. While insiders do own shares, they don't own a heap, and they have been selling. So we'd only buy after careful consideration. Therefore, you should should definitely take a look at this FREE report showing analyst forecasts for Steadfast Group.
But note: Steadfast Group may not be the best stock to buy. So take a peek at this free list of interesting companies with high ROE and low debt.
For the purposes of this article, insiders are those individuals who report their transactions to the relevant regulatory body. We currently account for open market transactions and private dispositions, but not derivative transactions.
We aim to bring you long-term focused research analysis driven by fundamental data. Note that our analysis may not factor in the latest price-sensitive company announcements or qualitative material.
If you spot an error that warrants correction, please contact the editor at firstname.lastname@example.com. This article by Simply Wall St is general in nature. It does not constitute a recommendation to buy or sell any stock, and does not take account of your objectives, or your financial situation. Simply Wall St has no position in the stocks mentioned. Thank you for reading.


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Oneview Healthcare is delighted to announce the appointment of Mr James Fitter as CEO. James has over 25 years' experience in the global financial markets during which time he has lived and worked on four continents working as an investment banker, a portfolio manager and a professional investor.
James was formerly the global head of emerging market equities at Deutsche Bank and the CEO of Sovereign Asset Management, a multi-billion dollar family office. His appointment broadens the senior management team and allows founder of Oneview, Mark McCloskey to focus his efforts entirely on product development and the conversion of Oneview's considerable global pipeline.
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subset: commoncrawl


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EMMERDALE fans were left in tears tonight as Bernice Blackstock exited the soap in truly dramatic fashion.
She marched out of the Dales and away to Australia after calling off her wedding and breaking fiance Liam Cavanagh's heart in the process.
She told him the wedding was on hold because she had to take daughter Dee Dee to her critically injured dad Charlie - Bernice's third husband.
However, she failed to mention that she probably wasn't coming back.
Bernice told Liam: "I can't marry you tomorrow - because I'm going with her. I'm sorry my darling, I can't marry you."
She then said goodbye to him and her mum in a tear-jerking finale which is widely thought to be Samantha Giles' last scenes after revealing she'd quit in September.
One viewer wept: "Bernice what are you doing 😭😭😭😭😭Liam makes you happy! Now im sobbing! X."
Another wailed: "Why can't Bernice ever have happiness ? She is so precious and just deserves the world Damn it!"
A third said: "Bernice leaving for unselfish reasons. I'm quite emotional 🥺."
One other sobbed: "I'm actually gutted Bernice and Liam never made it down the aisle 🥺."
Viewers saw her explaining what had happened to Charlie, saying: "He's basically had a massive stroke and the effects can be catastrophic.
"He's going to need months of rehab. He could be brain-damaged or paralysed."
And, setting up the fact she won't be returning, Bernice was warned she would be "walking away indefinitely".
She replied: "What choice? Choose between putting my daughter first or my happiness with Liam?"
And fans agreed, with one writing: "First rule of Motherhood... There IS NO CHOICE. Your daughter comes first."
But she didn't tell Liam she wouldn't be back, with the pair sharing an emotional dance in their living room before she headed off.
One viewer wailed: "Wish they'd have given Bernice more serious stuff to act out. @sammeegiles is breaking my HEART.
"You had a good 'un here, #Emmerdale. I hope she comes back one day to some real gritty material x."


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Faculty and Staff
Kyle Walton
I am a graduate of Fordham University with a BA in Theater Performance and English. My theatrical experience encompasses both the artistic and educational. As an artist, I have worked in states as far north as Maine and in our home state of NJ. My repertoire of performances includes off Broadway with the Brooklyn Symphony Orchestra, at the prestigious Shakespeare Theater of NJ, and in several independent films. I have also worked on the technical end of theater in a props warehouse, as well as all facets of stage crew in many productions and as a production assistant on a Netflix television show.
My experience as an educator includes being a teaching artist with McCarter Theater and the Shakespeare Theater of NJ and teaching acting to students ages 5-17 in New York City and Maine. I am also an audiobook narrator and producer.
Upcoming Events
1st Marking Period Exams
December 10-13
End 1st Marking Period
December 13
Holiday Sing
December 19
Teacher In-Service (School Closed)
December 20School Calendar
School News
Cedar Hill Prep Continues to Win as they Win the First Debate of the School Year! Cedar Hill Prep School made a clean sweep at the first Debate Meet of the Garden State Debate League, held at the Hun School on October 26th, 2019. Arnav Upadhyay won the Golden Gavel for the Best Speaker at the tournament, out of approximately 100 speakers. Cedar Hill Prep School teams walked away with the most wins by percentage points and overall wins. Thirty-seven teams participated in all. Congratulations Cedar Hill Prep Debate Team!
Did You Know?
The Multiple Advantages of a Multilingual CurriculumThere have been a number of studies in recent years that suggest people who speak more than one language are in fact more intelligent. An article written by the New York Times highlights that learning multiple languages could improve cognitive skills beyond those related to language, which is a considerably different viewpoint than previously thought.


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See the article in its original context from
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NO MORE STEAM BOILER EXPLOSIONS.--We notice among the articles on exhibition at the Fair, a machine called the Hydrostat, for preventing steam boiler explosions. It may be seen in operation on one of the boilers in the engine room. It is really a beautiful and curious, yet very simple piece of mechanism. A circular View Full Article in Timesmachine »


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State Codes and StatutesStatutes > Nevada > Title-33 > Chapter-378a
378A.010 - Policy of this State; purpose of chapter.
378A.020 - "Board" defined.
378A.030 - Creation; membership; Secretary; recommendations to Legislature.
378A.040 - Qualifications, appointment and terms of members.
378A.050 - Meetings; salaries of members.
378A.060 - Duties.
378A.070 - Account for Historical Records: Creation; use; deposits; payment of claims.
378A.080 - Acceptance of gifts and grants of money.
378A.090 - Filing of documents with State Library and Archives.


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Tendinopathy is a common condition that can affect any tendon in the body. The most common which you are likely to have heard of are Achilles, Gluteal (check out our
Tag: Injury Management
Knee Ligament InjuriesKnee Ligament Injuries
October 2, 2019October 2, 2019| | 0 Comment
Winter is usually a time where we tend to see an increase in knee injuries, especially knee ligament injuries. This is largely thanks to the variety of 'change of direction'
Sever's DiseaseSever's Disease
September 27, 2019September 27, 2019| | 0 Comment
Aches and pains are relatively common in growing children, especially these days where it seems to be the norm for kids to play multiple different sports during the one season.
Sciatica. How do we treat it?Sciatica. How do we treat it?
September 26, 2019September 26, 2019| | 0 Comment
As discussed in part 1 there are numerous causes of sciatica, so if you missed part 1 you can read it HERE.… Part 2 of this blog will be discussing
Self management for long term pain reliefSelf management for long term pain relief
September 26, 2019September 26, 2019| | 0 Comment
At Active RX one of our main aims is to give our clients the basic knowledge and understanding of how the body is put together in the hope that one


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Heroes & Pioneers
On A Quest to End to Alzheimer's: Dr. Stephen Salloway in East Side Living Magazine
Dr. Stephen Salloway, director of the Memory and Aging Program at Butler Hospital, shares what drives his 25+ year quest to end Alzheimer's in East Side Living magazine.
Weathering the Journey After an Alzheimer's Diagnosis
Carole Rowe, a nurse by profession as well as a caregiver to her husband, shares her thoughts and advice on weathering the journey that comes after an Alzheimer's diagnosis.
World's First A4 Alzheimer's Prevention Study Participant Celebrates Completion of Study
The world's first participant in an Alzheimer's prevention study finished his 4 1/2-year clinical trial last month. Here's what he has to say about the experience.
CrossFitting for A Cure: How One Woman Is Turning Fear Into Action in The Fight Against Alzheimer's
Tracie Kurczy-Lussier of Auburn, MA isn't just dedicated to keeping fit through CrossFit – she's also using it to be a warrior in the fight to end Alzheimer's.
Caregiver Martha Langer on Positivity, Participation and Hope
Martha and her mother have been involved in the ENGAGE clinical study since 2015. The experience has given her special insights – as well as hope and perspective – in her role as caregiver to a loved one with Alzheimer's.
Volunteer Ernest Labbe: Why I'm A Member of the "Swab Squad"
"My volunteerism is based on my dad teaching me that we should all work to leave this place a little better. It does not matter your situation, there is always something that can be done to make a positive difference…"


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|In stock
There is No-Hiding it, finally, a healthier and more digestible alternative to Rawhide. No-Hide Chews gives new meaning to dog treats. They are more durable and digestible than any other chew on the market. When it comes to Earth Animal's true core values, we insist on quality and we never settle for second best. We were tired of unacceptable ingredients and chemicals. The inspiration for No-Hide was to make a nutritious treat that would keep our dogs busy, help maintain healthy teeth and gums and have a chew that is easily digestible. No-Hides are one of the first chews of its kind and Hide free! The people have spoken and the dogs are begging for more, they are simply the best treat on the planet! Durable and long lasting makes a joyously, satisfied happy pup!
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Holy Cow, We did it! At last, a healthier, better alternative to Rawhide. No-Hide Stix gives new meaning to dog treats. When it comes to our true core values, we insist on quality and we never settle for second best. We were tired of unacceptable ingredients and chemicals. This is why we created No-Hide they are one of the first chews of its kind and Hide free!
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Montlucon Hotel
Search 10 hotels in Montlucon
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Where to stay in Montlucon
What's Montlucon like?
Where are the best places to stay in Montlucon?
Here are our travelers' top 4 places to stay in Montlucon:
The Originals Boutique, Hôtel de l'Univers, Montluçon (Inter-Hotel)Art Deco hotel in city center
- • Free WiFi • Sauna • Fitness center • Bar
Hôtel des BourbonsCity-center hotel
- • Free parking • Free WiFi • 2 restaurants
B&B Hotel Montluçon
- • Free parking • Free WiFi • Attentive staff
Ibis Montluçon3-star hotel within walking distance of Wilson Garden
- • Free WiFi • Bar • 24-hour front desk • Central location
What types of hotels are available in Montlucon?
We have 15 Montlucon accommodations with prices starting at INR 2577. Choose one of our 196 deals and get discounts of up to 20%.
Below are the number of accommodations by star rating in Montlucon and the surrounding area:
- • 3 4-star accommodations from INR 11309 per night
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How to Get to Montlucon
What are the closest train stations to Montlucon?
- • Montluçon Station (0.3 mi/0.5 km from the city center)
- • Montlucon-Ville Station (0.3 mi/0.5 km from the city center)
- • Montluçon-Rimard Station (0.5 mi/0.8 km from the city center)
Things to See and Do in Montlucon
What is there to see in Montlucon?
- • Parc des expositions de Montlucon
- • Chateau des ducs de Bourbon
- • Church of Notre-Dame
- • Wilson Garden
What is there to do in Montlucon?
- • Theatre Municipal Gabrielle Robinne
- • Museum of Popular Music
When is the best time to visit Montlucon?
- • Hottest months: July, August, June, September (average 76°F)
- • Coldest months: January, December, February, March (average 33°F)
- • Rainiest months: May, July, June, and August (average 3.60 inches of rainfall)


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Acyl-CoA oxidase complexes control the chemical message produced by Caenorhabditis elegans
Edited by David W. Russell, University of Texas Southwestern Medical Center, Dallas, TX, and approved February 20, 2015 (received for review December 19, 2014)
Many free-living and parasitic nematode (roundworm) species use pheromones called ascarosides to control their development and behavior. The molecular mechanisms by which environmental factors influence pheromone composition are unknown. The side chains of the ascarosides are derived from long-chain fatty acids that are shortened through β-oxidation cycles. In this manuscript, we show that three acyl-CoA oxidases, which catalyze the first step in the β-oxidation cycles, form specific homo- and heterodimers with different side-chain length preferences. Stressful conditions induce the expression of specific acyl-CoA oxidases, resulting in corresponding changes in pheromone composition. Thus, our work demonstrates how environmental conditions can influence the chemical message that nematodes communicate to other roundworms in the population.
Caenorhabditis elegans uses ascaroside pheromones to induce development of the stress-resistant dauer larval stage and to coordinate various behaviors. Peroxisomal β-oxidation cycles are required for the biosynthesis of the fatty acid-derived side chains of the ascarosides. Here we show that three acyl-CoA oxidases, which catalyze the first step in these β-oxidation cycles, form different protein homo- and heterodimers with distinct substrate preferences. Mutations in the acyl-CoA oxidase genes acox-1, -2, and -3 led to specific defects in ascaroside production. When the acyl-CoA oxidases were expressed alone or in pairs and purified, the resulting acyl-CoA oxidase homo- and heterodimers displayed different side-chain length preferences in an in vitro activity assay. Specifically, an ACOX-1 homodimer controls the production of ascarosides with side chains with nine or fewer carbons, an ACOX-1/ACOX-3 heterodimer controls the production of those with side chains with seven or fewer carbons, and an ACOX-2 homodimer controls the production of those with ω-side chains with less than five carbons. Our results support a biosynthetic model in which β-oxidation enzymes act directly on the CoA-thioesters of ascaroside biosynthetic precursors. Furthermore, we identify environmental conditions, including high temperature and low food availability, that induce the expression of acox-2 and/or acox-3 and lead to corresponding changes in ascaroside production. Thus, our work uncovers an important mechanism by which C. elegans increases the production of the most potent dauer pheromones, those with the shortest side chains, under specific environmental conditions.
The nematode Caenorhabditis elegans secretes the ascarosides, structurally diverse derivatives of the 3,6-dideoxy-l-sugar ascarylose, as chemical signals to control its development and behavior. Under favorable growth conditions, C. elegans progresses from the egg through four larval stages (L1–L4) to the reproductive adult. At high nematode population densities, however, specific ascarosides, which are together known as the dauer pheromone, trigger entry into a specialized L3 larval stage called the dauer (Fig. 1A) (1⇓⇓⇓–5). Dauers have a thickened cuticle, do not feed, derive energy from fat stores, and up-regulate stress-resistance pathways (6). Certain ascarosides also influence behaviors, including male attraction to hermaphrodites, hermaphrodite attraction to males, avoidance, and aggregation (7⇓⇓⇓⇓–12). C. elegans senses the ascarosides using several classes of G protein-coupled receptors (GPCRs), which are expressed in specific chemosensory neurons (13, 14). The dauer pheromone ascarosides induce dauer formation by downregulating the insulin/insulin-like growth factor 1 (IGF-1) and TGF-β pathways, which control not only dauer development but also metabolism and lifespan in C. elegans (15).
All ascarosides have a simple, modular structure with a fatty acid-derived side chain (Fig. 1B). The fatty acid side chain is attached to the ascarylose sugar at either its penultimate (ω-1) or terminal (ω) carbon, and is sometimes unsaturated (Δ) at the α-β position. The ascarosides can be named according to the number of carbons in their side chains using the following rubric: head group-asc-(ω)(Δ)C#-terminus group (16). Head groups attached to the ascarylose sugar include the indole-3-carbonyl (IC) and glucosyl (Glu) modifications, and terminus groups on the fatty acid terminus include the methylketone (MK) and para-aminobenzoic acid (PABA) modifications (Fig. 1B). The dauer pheromone consists of at least five ascarosides, asc-C6-MK (C6; ascr#2), asc-ΔC9 (C9; ascr#3), asc-ωC3 (C3; ascr#5), IC-asc-C5 (C5; icas#9), and asc-ΔC7-PABA (ascr#8) (Fig. 1A) (2⇓⇓–5). Of the dauer pheromone ascarosides, only asc-ωC3 has a side chain that is attached to the ascarylose sugar at its ω-carbon [as opposed to its (ω-1)-carbon]. Intriguingly, this ascaroside works synergistically with the others to induce dauer formation (3), and targets a unique class of GPCRs (13, 14).
Biosynthesis of the side chains of the ascarosides requires peroxisomal β-oxidation cycles that shorten the side chains by two carbons per cycle (11, 17⇓–19). Four peroxisomal enzymes, an acyl-CoA oxidase (ACOX-1), enoyl-CoA hydratase (MAOC-1), (3R)-hydroxyacyl-CoA dehydrogenase (DHS-28), and 3-ketoacyl-CoA thiolase (DAF-22), have been implicated in the β-oxidation cycles for ascaroside biosynthesis (Fig. 1C) (11, 17⇓–19). These enzymes are homologous to mammalian peroxisomal enzymes that process very long chain fatty acids, branched-chain fatty acids, and bile acid intermediates (20). Worms with mutations in maoc-1, dhs-28, or daf-22 do not produce any of the short-chain, dauer-inducing ascarosides, but instead accumulate ascarosides with long-chain side chains, stalled at different steps in the β-oxidation process (11, 17). This phenotype suggests the following three models for the role of β-oxidation in ascaroside biosynthesis: (i) β-Oxidation shortens long-chain ω/(ω-1)-ascarosides to short-chain ω/(ω-1)-ascarosides; (ii) β-oxidation shortens long-chain ω/(ω-1)-hydroxylated fatty acids to short-chain ω/(ω-1)-hydroxylated fatty acids, which are then attached to the ascarylose sugar; or (iii) β-oxidation shortens long-chain fatty acids to short-chain fatty acids, which are then ω/(ω-1)-hydroxylated and attached to the ascarylose sugar (Fig. S1). Worms with mutations in acox-1 have a less severe phenotype; they produce very little asc-ωC3, asc-C5, asc-ΔC7, and asc-ΔC9 and, instead, accumulate asc-ωC5 and asc-C9, implicating ACOX-1 in the β-oxidation of both ω- and (ω-1)-ascarosides (11). This result suggests that fatty acid side-chain hydroxylation precedes β-oxidation during the biosynthetic process and that the third model is not correct.
Ascaroside production has been shown to be influenced by a variety of factors, including developmental stage (21), temperature (3, 19), and sex (8), although interpretation of these results is often complicated by the fact that they were obtained by changing multiple variables at the same time (21) or by using long-term, mixed-larval stage cultures (3, 19). Starvation has been suggested to increase the ratio of asc-ΔC9 to asc-ωC3 in long-term, mixed-stage cultures (11). On the other hand, starved L1 larvae, in comparison with those that are fed, have been shown to secrete a higher ratio of ascarosides with shorter, 5-carbon side chains to those with longer, 9-carbon side chains (12).
Here we show that specific acyl-CoA oxidases enable C. elegans to modulate the composition of the ascaroside pheromones that it produces under different environmental conditions. Specifically, an ACOX-1 homodimer acts on the CoA-thioester of an ascaroside with a 9-carbon (ω-1)-side chain, an ACOX-1/ACOX-3 heterodimer acts on one with a 7-carbon (ω-1)-side chain, and an ACOX-2 homodimer acts on one with a 5-carbon ω-side chain. Using quantitative (q)RT-PCR, we show that acox-2 and acox-3, which act as gatekeepers for the production of shorter-chain ascarosides, are regulated by changes in temperature and food availability, leading to corresponding changes in ascaroside production. Studies of the ascaroside biosynthetic pathway and how it is modulated by different factors to control pheromone production will shed light on chemical communication in the C. elegans model system, as well as in other free-living and parasitic nematode species, which also use ascarosides as pheromones (22, 23).
Role of the ACOX-1 Homodimer in Ascaroside Biosynthesis.
BLAST analysis of ACOX-1 indicated the existence of several homologs in C. elegans (Fig. S2). To characterize the role of acyl-CoA oxidases in ascaroside production, we cultivated mixed-stage cultures of wild-type (N2) worms, available deletion mutants of acox-1 and acox-3 (F08A8.3), as well as a recently isolated acox-2 (F08A8.2) nonsense mutant (24). The ascarosides secreted by these mutants were then analyzed by LC-MS/MS in precursor scanning mode, monitoring for the product ion m/z 73.0 (11, 16). The mutants displayed distinct defects in ascaroside biosynthesis (Fig. 2). Similar to as shown by von Reuss et al. (11), the acox-1 mutant produces an increased amount of asc-C9 and a reduced amount of asc-ΔC9 (Fig. 2A), suggesting that ACOX-1 is required in the first step of the β-oxidation cycle that processes a 9-carbon (ω-1)-side chain to a 7-carbon (ω-1)-side chain. The mutant produces a similar amount of asc-C7 as wild type but no asc-ΔC7 and very little asc-C5, suggesting that ACOX-1 is required for the β-oxidation cycle that processes a 7-carbon (ω-1)-side chain to a 5-carbon (ω-1)-side chain. The mutant produces an increased amount of asc-ωC5 and virtually no asc-ωC3, suggesting that ACOX-1 is also required in the cycle that processes a 5-carbon ω-side chain to a 3-carbon ω-side chain. In addition, the acox-1 mutant produces a reduced amount of asc-C6-MK, as determined by LC-MS (Fig. S3).
To investigate further the role of acox-1, we cloned the gene from a cDNA library, expressed it in Escherichia coli, and purified the protein for in vitro assays against synthetic substrates. Based on its retention time on a gel filtration column relative to protein standards, ACOX-1 was expressed as a homodimer. As the type(s) of substrates used by ACOX-1 has not been established, we synthesized CoA-thioesters of ascarosides (1–4), fatty acids (5–7), and hydroxylated fatty acids (8 and 9) of various carbon chain lengths (Fig. 3A). The CoA-thioesters of the ascarosides could be made by directly coupling the carboxylic acids of unprotected ascarosides with CoA with yields of 10–40%. The acyl-CoA oxidase activity of ACOX-1 against the different substrates was determined by monitoring H2O2 production using a coupled reaction catalyzed by peroxidase that generates a UV-active product (25). ACOX-1 demonstrated a broad substrate range against the CoA-thioesters of fatty acids and hydroxylated fatty acids (Fig. 3B), consistent with a general role for acox-1 in peroxisomal fatty acid β-oxidation. In terms of ascaroside substrates, ACOX-1 was specifically active toward asc-C9-CoA (4) (Fig. 3B). The kcat and Km of ACOX-1 against asc-C9-CoA (4) were of a similar order as those of mammalian acyl-CoA oxidases against fatty acyl-CoA substrates (26) (Table S1). Surprisingly, however, ACOX-1 was not active against any other ascaroside-based substrates, including asc-ωC5-CoA (1) or asc-C7-CoA (3), contrary to what might be expected from the LC-MS/MS data of the acox-1 mutant (Fig. 2A).
Role of the ACOX-1/ACOX-3 Heterodimer in Ascaroside Biosynthesis.
In comparison with wild-type worms, the acox-3 deletion mutant produces an increased amount of asc-C7 and a reduced amount of asc-ΔC7 (Fig. 2B). These data indicate that ACOX-3 is involved in the first step of the β-oxidation cycle that processes a 7-carbon (ω-1)-side chain to a 5-carbon (ω-1)-side chain. The phenotype of the acox-3 mutant, however, is more subtle than that of the acox-1 mutant (compare Fig. 2 A and B), suggesting that ACOX-3 is not solely responsible for this step.
To investigate further the role of acox-3, we cloned the gene from a cDNA library and attempted to express it in E. coli to purify the protein for in vitro activity assays. However, we were not able to overexpress ACOX-3 in either a soluble or insoluble form, even after synthesizing a codon-optimized version of the gene, cloning it into various expression plasmids with either an N- or a C-terminal His tag, and repeatedly attempting to optimize expression conditions. Because the LC-MS/MS data of the acox-1 mutant indicated that ACOX-1 is required for the β-oxidation cycle that processes a 7-carbon (ω-1)-side chain to a 5-carbon (ω-1)-side chain, we speculated that ACOX-1 and ACOX-3 may work together and that coexpression of ACOX-1 with ACOX-3 in E. coli may be required for the expression of ACOX-3. The acox-1 and acox-3 genes were cloned such that ACOX-1 would be expressed without a tag and ACOX-3 would be expressed with a His tag. This coexpression system enabled expression and purification of an ACOX-1/ACOX-3 heterodimer. The two proteins appeared on an SDS/polyacrylamide gel as two bands of slightly different apparent molecular weight in an approximate 1:1 ratio, and their identities were verified through tryptic digestion of the proteins and mass spectrometry of the fragments (Fig. S4 A and B). To confirm the identities of the two bands, the acox-1 and acox-3 genes were recloned such that ACOX-1 would be expressed with a FLAG tag and ACOX-3 would be expressed with a His tag. Western blot of the heterodimer supported the identification of the two bands as ACOX-1 and ACOX-3 (Fig. S4C). Thus, our results provide strong evidence that the expression and/or stability of ACOX-3 is dependent on the formation of a complex with ACOX-1.
The acyl-CoA oxidase activity of the ACOX-1/ACOX-3 heterodimer was monitored against different substrates (Fig. 3 A and C). Remarkably, the ACOX-1/ACOX-3 heterodimer was specifically active toward asc-C7-CoA (3) (Fig. 3C and Table S1). Unlike the ACOX-1 homodimer, the ACOX-1/ACOX-3 heterodimer was not active against any fatty acid or hydroxylated fatty acid-type substrates. This substrate specificity provides the first, to our knowledge, direct evidence that the acyl-CoA oxidases and other enzymes involved in the β-oxidation cycles in the ascaroside biosynthetic pathway act directly on the CoA-thioesters of ascarosides, rather than on the CoA-thioesters of either fatty acids or hydroxylated fatty acids (that is, model 1 in Fig. S1). The substrate specificity of the ACOX-1/ACOX-3 heterodimer is consistent with the LC-MS/MS ascaroside profiles of the acox-1 and acox-3 mutants, which both showed an increase in the ratio of asc-C7 to asc-ΔC7 relative to wild type. Thus, the ACOX-1/ACOX-3 heterodimer plays a key role in the β-oxidation cycle that shortens a 7-carbon (ω-1)-ascaroside to a 5-carbon (ω-1)-ascaroside. Furthermore, both the acox-1 and acox-3 mutants show a reduced amount of asc-C6-MK as analyzed by LC-MS (Fig. S3). Thus, the biosynthesis of asc-C6-MK occurs downstream of the conversion of asc-C7-CoA (3) to asc-ΔC7-CoA catalyzed by the ACOX-1/ACOX-3 heterodimer.
Role of the ACOX-2 Homodimer and ACOX-1/ACOX-2 Heterodimer in Ascaroside Biosynthesis.
In comparison with wild-type worms, the acox-2 nonsense mutant produces an increased amount of asc-ωC5 and a reduced amount of asc-ωC3 (Fig. 2C). These data indicate that ACOX-2 plays a role in the β-oxidation cycle that processes a 5-carbon ω-side chain to a 3-carbon ω-side chain. To determine the substrate specificity of ACOX-2, we expressed ACOX-2 in E. coli and purified the protein for in vitro activity assays. The ACOX-2 homodimer was not active against any fatty acid or hydroxylated fatty acid-type substrates, but was active specifically toward asc-ωC5-CoA (1) (Fig. 3 A and D). Thus, the substrate specificity of the ACOX-2 homodimer is consistent with the LC-MS/MS ascaroside profile of the acox-2 mutant.
Because the LC-MS/MS data of the acox-1 mutant suggested that ACOX-1 plays a role in the β-oxidation cycle that processes a 5-carbon ω-side chain to a 3-carbon ω-side chain, we speculated that ACOX-1 might also form a heterodimer with ACOX-2. The acox-1 and acox-2 genes were cloned such that ACOX-1 would be expressed without a tag and ACOX-2 would be expressed with a His tag. This coexpression system suggested that ACOX-1 and ACOX-2 interact, as verified through tryptic digestion of the proteins and mass spectrometry of the fragments (Fig. S5 A and B). In terms of ascaroside substrates, the purified proteins were most active toward asc-ωC5-CoA (1) and showed a greater catalytic efficiency toward asc-ωC5-CoA (1) than did the ACOX-2 homodimer (Fig. 3D, Fig. S5C, and Table S1). To confirm the interaction between ACOX-1 and ACOX-2, the acox-1 and acox-2 genes were recloned such that ACOX-1 would be expressed with a FLAG tag and ACOX-2 would be expressed with a His tag. Western blot confirmed that ACOX-2 could pull down ACOX-1, although the protein purified is likely a mixture of ACOX-1/ACOX-2 heterodimers and ACOX-2 homodimers (Fig. S5D). Thus, an ACOX-2 homodimer, as well as possibly an ACOX-1/ACOX-2 heterodimer, plays an important role in the β-oxidation cycle that processes a 5-carbon ω-side chain to a 3-carbon ω-side chain to produce asc-ωC3.
Another possible explanation for the effect of the acox-1 deletion on asc-ωC3 biosynthesis is that the deletion influences the expression of acox-2, because the deletion is only 1.299 kb upstream of the start codon of acox-2. However, analysis of acox-2 expression in wild-type worms and in the acox-1 mutant using qRT-PCR showed that acox-2 expression is not strongly down-regulated in the mutant (Fig. S6). Thus, it is unlikely that the primary reason that the acox-1 mutant does not make any asc-ωC3 is simply because the deletion in this mutant interferes with acox-2 expression.
Effect of Environmental Conditions on Ascaroside Production and Acyl-CoA Oxidase Gene Expression.
Our biochemical results suggested that ACOX-2 and ACOX-3 serve as specialized acyl-CoA oxidases that promote the biosynthesis of short-chain ω- and (ω-1)-ascarosides, respectively. Thus, we hypothesized that the expression of these proteins may be regulated by environmental conditions to enable the composition of the ascarosides produced by C. elegans to change in response to specific conditions, such as different temperatures and starvation. To investigate the effect of food availability, we grew synchronized cultures of wild-type worms to the L4 stage but gave the cultures minimal bacterial food (E. coli) such that once the worms reached the L4 stage, they were starving. At this point, the cultures were either fed or not fed additional bacterial food, and the culture medium and worms from the cultures were collected at 4 and 24 h. By performing this experiment over a relatively short period in synchronized adults, we attempted to minimize the indirect effect of food on ascaroside production due to the increased growth rate upon food addition. Acyl-CoA oxidase gene expression in the worms at 4 h was analyzed using qRT-PCR, and ascaroside secretion into the culture medium at 24 h was analyzed using LC-MS. Production of most ascarosides was increased in fed worms, as one might expect, because the fed worms presumably have a greater metabolic potential. However, production of shorter-chain (ω-1)-ascarosides, such as asc-ΔC7, asc-C5, and asc-C6-MK, was not increased in fed worms as much or not at all, and production of IC-asc-C5 was reduced (Fig. 4A). Correspondingly, the acox-3 gene was strongly down-regulated by food addition, suggesting a mechanism whereby the production of shorter-chain (ω-1)-ascarosides is suppressed by these conditions (Fig. 4B).
To determine the effect of dauer-inducing (DI) conditions on ascaroside production and acyl-CoA oxidase gene expression in early larval stage worms, we monitored pheromone production in cultures initiated with synchronized L1 larvae under either DI (high population density and low food) or non-DI (low population density and high food) conditions (Fig. S7). Although pheromone levels were monitored from the L2 to young adult stages, it should be noted that worms grown under non-DI conditions have an increased growth rate relative to worms grown under DI conditions, presumably giving them greater capacity to produce pheromones, especially at later time points. Analysis of an early time point (28 h) indicates that non-DI conditions strongly induced the production of the short-chain ω-ascaroside asc-ωC3 (Fig. 4C). Consistent with increased asc-ωC3 production under non-DI conditions, the acox-2 gene was very strongly up-regulated under these conditions relative to DI conditions (Fig. 4D). Given that in our hands asc-ωC3 is the most robust dauer pheromone component (3), it was surprising that asc-ωC3 was produced in higher amounts under non-DI conditions because, in general, C. elegans would presumably not need to enter dauer under these conditions. However, we speculate that perhaps asc-ωC3 enables dauer formation in the face of specific stressors despite the presence of food or otherwise favorable conditions. Indeed, unlike other dauer pheromone components, asc-ωC3 induces dauer, even when the dauer formation assay is performed using live (antibiotic-treated) food, rather than heat-killed food (Fig. S8).
To investigate the effect of temperature on ascaroside production and acyl-CoA oxidase gene expression, we grew cultures of wild-type worms from eggs to the young adult stage at either 20 °C or 25 °C. Once the worms reached the young adult stage, they were grown for an additional 24 h at the different temperatures. The production of most ascarosides increased at the higher temperature (Fig. S9A). The expression of acox-2 and -3 also increased at the higher temperature (Fig. S9B). Increased ascaroside production may contribute to the increased dauer formation seen in C. elegans at higher temperatures.
The ascarosides represent a nuanced chemical language that C. elegans and many other species of nematodes use in communication. Depending on its larval stage, larval history, sex, temperature, and nutritional environment, C. elegans produces a different mixture of ascarosides to communicate different information to other individuals in the population and to improve the survival of conspecifics. Here we have shown that C. elegans uses specific acyl-CoA oxidase complexes to control the production of the short-chain ascarosides. Importantly, we show that an ACOX-1/ACOX-3 heterodimer catalyzes the first step in the β-oxidation cycle that shortens an ascaroside with a 7-carbon (ω-1)-side chain to one with a 5-carbon (ω-1)-side chain (Fig. 5). Thus, ACOX-3 is important for the biosynthesis of the shorter-chain (ω-1)-ascarosides, including the dauer pheromone asc-C6-MK. An ACOX-2 homodimer, as well as possibly an ACOX-1/ACOX-2 heterodimer, catalyzes the first step in the β-oxidation cycle that shortens an ascaroside with a 5-carbon ω-side chain to one with a 3-carbon ω-side chain (Fig. 5). Thus, ACOX-2 is important for the biosynthesis of the short-chain ω-ascarosides, including the robust dauer pheromone asc-ωC3, which acts synergistically with other ascarosides to induce dauer (3).
Consistent with the role of ACOX-3 in the production of short-chain (ω-1)-ascarosides, the expression of acox-3 and the production of short-chain (ω-1)-ascarosides (e.g., asc-ΔC7, asc-C5, and asc-C6-MK) are both suppressed in synchronized L4 larvae upon the addition of bacterial food. Consistent with the role of ACOX-2 in the production of short-chain ω-ascarosides, the expression of acox-2 and the production of asc-ωC3 are both increased in early larval stage worms grown in non-DI conditions. Our results show that acox-2 and -3 are transcriptionally up-regulated in adult worms that have been grown at higher temperatures. These results agree with expression profiling experiments that showed that acox-2 and -3 are up-regulated by a shift to higher temperatures (27). Up-regulation of acyl-CoA oxidase genes may contribute to increased pheromone production at higher temperatures.
It is somewhat counterintuitive that production of asc-ωC3, a potent dauer pheromone, would be increased under non-DI conditions. Indeed, this ascaroside targets GPCRs in the ASI neuron, which plays a direct role in inducing dauer (14). Unlike several other dauer pheromone components, asc-ωC3 has a very sharp EC50 curve in the dauer formation assay and induces close to 90% dauers at higher concentrations (3). The EC50 of asc-ωC3 in the dauer formation assay is also similar at low and high temperatures (3, 4). Our results show that asc-ωC3 can induce dauer even in the presence of live (antibiotic-treated) bacteria. Thus, asc-ωC3 may enable dauer formation in response to certain stressors, even when conditions are otherwise favorable. Alternatively, it could be that asc-ωC3 has some biological activity or role that has yet to be identified. It is unknown whether asc-ωC3 production in non-DI conditions is a response to specific nutrients in the bacteria, the slight pathogenicity of E. coli (28), or other factors.
In mammals, the first step in peroxisomal β-oxidation is catalyzed by several acyl-CoA oxidases with different substrate preferences (20). Palmitoyl-CoA oxidase is specific to the CoA-thioesters of straight-chain fatty acids, pristanoyl-CoA oxidase is specific to the CoA-thioesters of 2-methyl-branched-chain fatty acids and long/very long straight-chain fatty acids, and cholestanoyl-CoA oxidase is specific to the CoA-thioesters of bile acids. Whereas palmitoyl-CoA oxidase and cholestanoyl-CoA oxidase exist as homodimers (29, 30), pristanoyl-CoA oxidase is thought to exist as a homooctamer (31). To our knowledge, our work is the first to provide evidence that acyl-CoA oxidases can function as heterodimers. The crystal structure of rat palmitoyl-CoA oxidase homodimer bound to a fatty acid suggests that substrate binding occurs at the dimer interface (32, 33). Thus, acyl-CoA oxidase heterodimers may form a composite binding site at their interface that is different from the corresponding homodimers and thus shows an altered substrate preference. In addition to ACOX-1, -2, and -3, there are several other acyl-CoA oxidases in C. elegans that may participate in ascaroside biosynthesis (Fig. S2). ACOX-1 is also predicted to have three different splice variants that could potentially play specific roles in ascaroside biosynthesis. Different splice variants of human palmitoyl-CoA oxidase have been reported to have different fatty acid chain length preferences (26).
Our work demonstrates that β-oxidation enzymes act directly on long-chain ascarosides to shorten them, as opposed to acting on long-chain fatty acids or hydroxylated fatty acids that once shortened are subsequently attached to the ascarylose sugar. Thus, we have established the order of the main steps in ascaroside biosynthesis, which will likely provide insight into pheromone biosynthesis not only in C. elegans but in other nematode species as well. Furthermore, our work provides a general framework for understanding the effects of environmental conditions on ascaroside side-chain length and pheromone composition. This knowledge will shed light on how nematodes respond to a changing environment and adapt accordingly.
Materials and Methods
C. elegans Liquid Cultures.
Strains are listed in SI Materials and Methods. Large-scale (150 mL) nonsynchronized worm cultures were fed E. coli (HB101) and grown for 9 d, and extracts were generated from the culture medium and analyzed by LC-MS/MS, as described (16). Small-scale (50 mL) synchronized worm cultures were used to test the effects of environmental conditions on ascaroside production and acyl-CoA oxidase gene expression, as described in SI Materials and Methods. For sample collection in these experiments, 5 mL culture was centrifuged (800 × g for 2 min), the worms at the bottom were removed, and the supernatant was centrifuged again (2,671 × g for 10 min). One milliliter of this supernatant was lyophilized and resuspended in 100 μL of 50% (vol/vol) methanol in water, and the ascarosides were analyzed by LC-MS as described (16) with modifications (SI Materials and Methods). The worms obtained were washed with cold M9 buffer several times, flash-frozen, and stored at −80 °C until they were used for qRT-PCR (SI Materials and Methods).
Acyl-CoA Oxidase Expression and Biochemical Assay.
acox-1a and acox-2 were each cloned by PCR from a C. elegans (N2) cDNA library, and a codon-optimized version of acox-3 was chemically synthesized (GenScript). acox-1a and acox-2 were inserted into a modified pET-16b vector at the NcoI/NotI restriction sites (C-terminal His tag). The plasmids were individually transformed into BL21(DE3) cells, and expression was induced overnight at 16 °C using 0.6 mM isopropyl β-d-1-thiogalactopyranoside (IPTG). For coexpression experiments, acox-1a was inserted into pACYCDuet-1 at the NcoI/NotI sites (with either no tag or a C-terminal FLAG tag), acox-2 was inserted into a modified pET-16b vector at the NcoI/NotI sites (C-terminal His tag), and acox-3 was inserted into a modified pET-16b vector at the NcoI/NotI sites (C-terminal His tag). The plasmids were cotransformed into BL21(DE3) cells, and expression was induced overnight at 20 °C using 0.5 mM IPTG. Cells were lysed using a microfluidizer. The protein was purified with Ni-NTA resin (Thermo Fisher), concentrated with a 10 kDa-cutoff Centricon (Millipore), further purified on a HiLoad 16/600 Superdex 200 column (GE Healthcare) attached to an FPLC, and concentrated again to 1 mg/mL for assay. The acyl-CoA oxidase activity assay was based on a previously reported method (25) with modifications (SI Materials and Methods).
Synthesis of CoA-Thioesters.
Ascarosides were synthesized as previously described (34), and (R)-6-hydroxyheptanoic acid and (R)-8-hydroxynonanoic acid were synthesized as described in SI Materials and Methods. CoA-thioesters of ascarosides, fatty acids, and hydroxylated fatty acids (1–9) were synthesized based on a previously described method for making fatty acyl-CoA thioesters (35) with modifications (SI Materials and Methods).
We are grateful to Kyle Hollister, Mark Spell, and Justin Ragains for providing initial stocks of several ascarosides that enabled us to obtain preliminary data. We thank Scott Neal and Sarah Hall for advice, and Shohei Mitani at the National BioResource Project (Japan) for strains. Some strains were also provided by the Caenorhabditis Genetics Center, which is funded by the NIH Office of Research Infrastructure Programs (P40 OD010440). This work was supported by the National Institutes of Health (GM87533) and the Ellison Medical Foundation.
- ↵1To whom correspondence should be addressed. Email: butcher{at}chem.ufl.edu.
Author contributions: X.Z., L.F., and R.A.B. designed research; X.Z., L.F., and Y.W. performed research; S.C., P.S., and J.K.N. contributed new reagents/analytic tools; X.Z., L.F., and R.A.B. analyzed data; and X.Z. and R.A.B. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1423951112/-/DCSupplemental.
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Peter Elliott tests the new Extreme 700 Game King, which is a trailer boat that he says has both fishing and family applications.+MORE
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2019 Fi-Glass Lightning
2019 Mercury 115hp 4 Stroke Pro XS
2019 Fi-Glass Multi Roller Trailer
Package includes:
Moulded Acrylic Top Screen
Carpet Flooring - Domed in for easy cleaning
Fi Glass Dominator in very good condition
NZs New Fi-glass dealer is pleased to announce the arrival of this stunning model now in stock at one of New Zealands busiest dealers located on the sunny North Shore.
This Sensational 6.4M Fi-Glass Warrior Is The Ultimate Boating Package In What Would Have To Be One Of The Biggest Selling Fiberglass Boat Brands In New Zealand
The Warrior Is Not Only An Incredible All-Rounder For The Lakes & Coastal Regions But It Handles Amazingly Well In Virtually All Conditions With The Bonus Of Fantastic Cabin Space & When Matched With The Smooth & Powerful 135 HP Mercury EFI 4 Stroke It Becomes The Perfect Combination For All Your Boating Adventures Definitely Call Today!
Fi Glass Firestar
Yamaha 90hp 2 stroke
Fi Glass single axle trailer
A deep-V, soft-riding boat that handles superbly on open or inland waters. It features a practical cabin area, and is an ideal open day-boat for fishing, skiing and cruising. The Senator offers excellent under-floor storage, and a number of seating options. This model has one of the most spacious cockpits of any 5.3 metre boat on the market. It is ideal for any fishing or watersport activities that you have in mind.
The Cavalier punches above its weight for value, flexibility and ease of use with a spacious cabin and ample storage capacity that will hold all of you gear. And at 6.5m trailer length Cavalier is easily stored in a standard sized garage and offers the features and specifcations of larger pleasure craft.
1990 Fi-Glass Warrior
2008 Evinrude 130hp 2 Stroke (310.7 hours)
1990 Single Axle Unbraked Trailer
Classic and power are always a good combination. This Fi-Glass has been repowered with a grea...
New Zealands New Figlass dealer is pleased to announce the arrival of this stunning model now in stock at one of New Zealands busiest dealers located on the sunny North Shore and the only factory authorised Dealer on the North Shore. Fi Glass boat are kiwi made.
With the finest pedigree born out of the ever-popular 'Lightning', the all new 'Cavalier' promises to be your dream come true. Industry leading construction embodies over fifty years of boat-building experience - with a generous cockpit that rivals the Lightning. Superb handling and versatility complete a standard specification package that you can adapt to call your own, with a generous array of options.
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Trade Marketing Specialist in St. Paul MN
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Dairy Farmers of America
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Marketing Representative, Trade Marketing/Manager
Responsible for the full cycle of trade promotions from Entry to Settlement in Anaplan. Help research, verify and resolve customer payment deductions across for the warehouse lines of business at Kemps. Works directly with our broker network and cross functionally with Sales and Trade Marketing to obtain and enter all Kemps promotions into Anaplan as well as verify performance and resolution.
Works with Field Sales and Broker network to obtain promotional plans and transfer those plans from excel & word into Anaplan
Secures and organizes backup from customers via website, email, or by phone.
Accurately verifies and resolves deduction information with promotions within Anaplan
Responsible for helping Deduction Specialists research and resolve promotional types of deductions (i.e. promotional chargebacks) in accordance with established policy and authorization limits.
Works with Accounting and Trade Marketing to ensure accurate General Ledger account coding is applied to resolved deductions. Coordinate with Trade Marketing, utilizing Promotional Calendars to match and apply deductions accurately.
Ability to research, summarize, and present facts in writing and verbally to internal teams
Enter market pricing data into our retail database and ability to use pivot tables to summarize the data for Trade Marketing and Sales consumption and reporting
Identify ways to improve consistency and streamline processes. Develop and maintain documentation for to use for training purposes.
Develop relationships to maintain and improve our internal business partnerships with Sales, Trade Marketing, and Finance. Actively participate in identifying ways to work more collaboratively.
Keeps supervisor informed of activities that may require their attention.
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After making our relationship "social media official" during Art Basel a few months after our first date, the real challenge came when we purchased our first house a few months after our wedding. We had a lot of wall space to fill but some pieces from our respective collections would inevitably not make the cut. Between her extensive collection of works by Cuban masters and my collection of nature-inspired textured series of original modern artworks, our curation process is to this day a work in progress! Recently, we attended the VIP Preview of the new Museum of Graffiti in Wynwood and picked up a limited-edition graffiti skateboard by Noah to add to our collection, appropriately titled "One Love." Next time you visit someone's home and admire their art, remember that each and every piece on display has a unique story behind it worth a million words.
Creative Space
Billionaire philanthropist, entrepreneur and contemporary art collector Jorge M. Pérez unveiled his highly anticipated experimental arts center, El Espacio 23, with an intimate preview event for local community leaders and South Florida cultural luminaries
Pass The Avocado
Turkey with stuffing. The Christmas ham. Blackeyed peas, cornbread and collard greens for good luck in the New Year. Enough holiday food. It's time to remember that we live in the south and nothing is more tropical than delicious, nutritious avocados.


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Dustee Miles Drawing, 2019-06-30 07:17:50. A finger chart is offered in tutor books in addition to separate fingerings on pages throughout in the detailed learning practice. The Logo drawing, on the flip side, was nearly a whole loss. It becomes interesting and easy to understand when introduced with alphabets. It is very important for young children. Drawing out what you would like to say can be fun.
Deonna Levy Drawing, 2019-07-04 10:06:39. You simply need to make sure to know why it's happening so that you can be more bold when you ink. Now you have ink on your pen, you're prepared to ink. To put it differently, you've got to ink somewhat faster than you feel you should. Put simply, you've got to ink a little faster than you are. Also there is absolutely no blending involved with drawing hair. Often it is a mixture of issues that result in a less than perfect'' portrait.
Neely Hendricks Drawing, 2019-07-04 10:06:39. Some ideas are extremely original and stick out between conventional strategies. These ideas are astoundingly simple, very simple drawing ideas for children. Finding a notion that seems worthwhile of your time can be a task alone.
Dawn Calhoun Drawing, 2019-07-04 10:06:39.Drawing or sketching are actions that begin with easy concepts which are easy to grasp and adhere to a few tries things will start to feel organic. Pictures could be performed anywhere without the problem of dragging a great deal of supplies round with you personally. Twist the paper so that the rest of the photograph is concealed. Simple drawings are not always straightforward to make. An excellent sketch is still possible to impress. You'd love to approach your sketchbook using a sense of existence and like what you are doing.
Marji Herrera Drawing, 2019-07-04 10:06:39. You won't require a computer to draw floor plans if you get an intelligent phone or tablet. The important thing here is to understand that the computer screen is not anything more than a fancier piece of graph paper. Different pictures will allow you to study your subject from various angles and in different lighting conditions. A cross-sectional view portrays a cut-away part of the object and is another means to show hidden components in a gadget.
Arlena Hammond Drawing, 2019-07-04 10:06:39. Unique textures can be achieved based on the method used to construct tone. The shape will be quite straightforward. The subsequent shape ought to be somewhat enjoy a carrot or an almond. Bear in mind your deviates can be accomplished with size, shape, or both. Start looking for two or three features that you may use to deviate from their usual dimensions or proportion. Run the tape along the surface of the baseboard so you find yourself with a whole wall-to-wall dimension. For instance, some objects need only two views, but others need four.
Mallory Guerrero Drawing, 2019-07-04 10:06:39. You may wish to begin the cartoon cow with a framework of some ovals and a couple lines. Other animals aren't that easy. Basically, you're combining body parts from assorted wild animals with that of a human for an enjoyable and imaginative spin on the cartooning approach. As an example, consider that you would like to draw a cartoon cat.
Deonna Levy Drawing, 2019-07-04 10:06:39. Bear in mind, not all the ideas will be winners. In reality, a few of the ideas may not even make sense. Quite often it can be hard brainstorming suggestions for our sketchbooks.
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TII India showcases
- Fresh from the factory in India: Tiiger SD modular platform trailer for loads up to 240 tonnes
- Designed in Germany, made in India, sold to regional markets: TII India is geared to the market requirements of lightweight cargo
For the transport of loads up to 240 tonnes on public roads or in industrial areas, TII India is launching a new vehicle on the market at bauma: the Tiiger Smart Duty (SD). The modular platform trailer has a 53 degree steering angle and hydraulically supported pendulum axles thus making it super light and resilient. Up to 18 tonnes can be transported per axle line. Developed in Germany and manufactured in India, the robust all-rounder is suitable for transporting construction machinery, cranes, wagons and concrete girders, among other things. The Tiiger SD is sold in India, Africa and Southeast Asia.
TII India is one of the four subsidiaries of the TII Group, and has a market share of around 70 per cent in India. In its home country, TII India is the market leader for heavy goods transport.
Managing Director of TII India Pvt. Ltd, Dr. Jochen Landes, is convinced: "With the Tiiger SD we have perfectly matched our customers' requirements regarding robust and cost-effective transport solutions for loads up to 240 tonnes. Our trailers have the strongest bending moment on the market, comply to international safety standards and regulations - all of this at competitive prices".
The TII Group with its daughter company TII India has been active in India since 2015. The TII India subsidiary manufactures transport vehicles under the Tiiger brand name for India, Africa and Southeast Asia. Designed in Germany and manufactured in India. With annual economic growth of around seven per cent and numerous infrastructure measures, India´s demand for high technology transport solutions is increasing. In particular, robust vehicle transportation of heavy loads (ODC/OWC) especially under challenging road conditions are in demand. This is precisely the TII Group's area of expertise: heavy-duty vehicles with a low deadweight for transporting excavators, transformers, ship components, wind power equipment and even entire plant facilities.
Product portfolio of TII India:
- TIIGER SD. modular platform trailer (18 tonnes per axle line)
- TIIGER MD: modular platform trailer (26 tonnes per axle line)
- TIIGER HD: modular platform trailer (36 tonnes per axle line)
- TIIGER Extender: telescopic semi-trailer up to 53 metres or 60 metres long
- Side girder bridge with loading capacity of up to 550 tonnes
- TIIGER semi-trailer
Energy (wind power, nuclear energy, heating); infrastructure
TII India is market leader in the heavy-duty range up to 240 t
Target markets:
India, South-East Asia, Africa


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Two pickup 4K versions.
34" P v 32" P v 30" P
skelf replied to obbm's topic in General DiscussionTalk to Eude he has a 30.5" TKO with a P Bass split coil in it.
They switch the coils on each pickup. Series/single/parallel. With regard to the tonal range from the filters. The lowpass filters in your version of the EQ01 go as high as 6.5kHz the newer version was brought down in line with the current DFM to 3.5kHz. This increases the resolution of the pot giving more control over the mids and it also reduces the overlap with the highpass filter. While both the EQ01 and DFM share the same control layout the newer restricted version of the lowpass filter does make it easier to dial in. Less being more in this case. This was done after the EQ03 where myself and John took things a bit to far. The preamp was hugely versatile but difficult to use. We only really worked that out once we went to far. Dialled it back to make the DFM which would be the EQ04. I don't see it being developed any further. As far as I am concerned it ticks all the boxes I set out to tick. It took 4 versions over 9 years a fair bit of cash a few mistakes and me being the blight of John existence. Full credit to John East for putting up with me and his brilliant design work to get me to where I am and wanted to be Regarding using it. I only use about 20% of what the preamp can do. That produces what I like so it is what I use. However the other 80% has to be there because it may well produce the 20% you like. Don't see anyway round that. If you want a versatile bass, and that has always been my goal, then there will be a degree of complexity and redundancy in the system. Bottom line use what you like ignore the rest. I look on it as a bass with two tone controls one for each pickup. I setup what I like on each pickup and use the blend. The blend is your friend. Because of the independent EQ it is way more powerful than in a 3 band EQ. All you have there is the difference between the pickups and their placement. With the filters the two pickups can have radically different sounds making the blend much more important. Live the blend makes changes easy. However once you have spent time with the preamp and I am the first to admit it has a learning curve changing on the fly is certainly possible I know many players who are totally comfortable with it live. Equally there are people that just don't get them or get on with them which is why the East Uni-Pre is always available in all my builds except for the Uber Series. The other thing to point out is that the filters give you an option you don't have. You have at least a 3 band cut and boost EQ in your amp Filters give you sounds you can't get with a 3 band. Conversely a 3 band is better at some things than the filters. You have the option of best of both. And finally for those that wish to know. Flat in a filter preamp is not entirely possible because the filters are engaged all the time.So the control setting for flat are not all in the middle as would be the norm. However you can get close with the following. Both lowpass filters fully open, upper ring fully of. Upper ring on the highpass fully of. With the active/passive that is available with the DFM A/B the above with the passive and tone fully up there is not a lot of difference. So should you want to try flat that is as close as you can get.
AC Guitars still make a filter per pickup system. While EQ01 is not currently available for sale I use the DFM (Dual Filter Modular) in my own instruments which is the fourth generation ACG dual filter preamp. There will also be a limited run of EQ02 preamps in the not to distant future.
*SOLD* Custom fretless ACG
skelf replied to Clarky's topic in Basses For SaleNot many can say they have number 1 of 1.
ACG Finn 4 (Premium Second Hand Bass)
skelf replied to skelf's topic in Commercial Items for saleDon't know at the moment.
ACG Finn 4 (Premium Second Hand Bass)
skelf posted a topic in Commercial Items for saleUnusual bass for sale with full explanation at the link below. Full details on this bass can be found at: http://www.acguitars.co.uk/project/0299finn4/ If interested in the bass please get in touch.
5 String Short Scales. Do they work?
skelf replied to Woodinblack's topic in Bass GuitarsUsed to chat with Tim on various forums. A really funny guy as well as great builder. I am a a big fan of short scales the more I build the more I like them. Improved string technology and the correct building techniques have made them more than a viable option for most players.
MINI SALE running until the end of my financial year 31.3.19.All stock basses and guitars are reduced some guitars up to 50%.Details in the Affiliates Forum.
MINI SALE starts on Friday morning ends when the show closes doors on Sunday night.Discounts available on all stock instruments during The Guitar Show in person at the show or via mail/messenger/phone.
3 Leaf Enabler and TC Polytune Pedal-Train Nano plus power supplies
skelf replied to skelf's topic in Effects For SaleNano etc on hold pending the usual.
3 Leaf Enabler and TC Polytune Pedal-Train Nano plus power supplies
skelf replied to skelf's topic in Effects For SaleJust about every thing has been sold the two last items are now bundled for £125.00 inc UK delivery.
3 Leaf Enabler and TC Polytune Pedal-Train Nano plus power supplies
skelf replied to skelf's topic in Effects For SalePolytune mini, Enabler and Phil Jones Headphones now sold.
3 Leaf Enabler and TC Polytune Pedal-Train Nano plus power supplies
skelf replied to skelf's topic in Effects For SalePolytone Standard now sold.
Hi A small clear out of gear because I have streamlined my show demo gear down to a Helix. Pretty much everything has now been sold. Just the Pedal-Train with Gig-Rig power supply is left so I will throw in the patch leads with it. £125 inc UK delivery. Pedal-Train Nano with gigbag and Gig Rig Generator plus 1 Distributor. All as new with velcro on the Nano. £125.00 inc UK SD. Would prefer not to split. 5 x Patch leads Nuetrik Jacks Klotz cable £30 inc UK Delivery. Any questions fire away. Cheers Alan


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In Belgium, if you look hard enough, you can still find dozens of restaurants where time seems to have stood still. Most of them aren't that well known, and that's a shame: these places really deserve to be cherished. Because they are witnesses of the past, with an interior which has been hardly touched for decades, and because they offer a taste of the cooking of the old days: hearty and heartily food, with straightforward and pure flavours. In Bistro Belge, Toni De Coninck tells the story of these establishments, and of the people who work there. He visited restaurants all over Belgium, from West-Flanders ('t Hommelhof in Watou) to Luxembourg (Le Moulin Hideux in Noirefontaine) and from Limburg (Tante ('Auntie') Julia in Stevoort) tot Brussels.
He learned all there is to know about the history of these places and the families who run them, and also gathered some recipes of speciality dishes that for generations have been prepared in the exact same way. The photos in this book complement the texts; they immerge the reader into the warm-heartedness that is so typical of the Belgian culinary heritage.


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Tomasz Kwiatkowski is a financial services sales executive, podcaster and a philanthropist. He was born and raised in Poland and moved to the U.S. as a first-generation immigrant in 2006 at the age of 19 by himself to pursue a better quality of life. He is an energetic, seven-feet tall leader and a self-starter whose passion lies at the intersection of people, sales and technology. Tomasz possesses a rare combination of a highly relationship-centric nature with the aggressive pursuit of impact and results to bring the vision to reality for the betterment of the community he serves in. He is an international citizen, a natural networker and a connector. "We all ought to develop meaningful symbiotic relationships with a purpose." – he says.
With 8 years of diversified business development and client-facing experiences, he has a proven track of record selling to many mid and large size organizations and helping them scale, improve operational efficiency and reduce cost. In the process, Tomasz has traveled extensively across the U.S. as well as covered large local territories to build relationships with C-level executives in the areas of financial services, technology, consulting and higher education.
In his current role as a Business Development Executive at PlainsCapital Bank (PCB) in Frisco, TX, Tomasz is responsible for developing new commercial lending relationships and increasing depository volume growth through extensive outside sales and community building efforts. In addition, he promotes and expands PCB's footprint in the Frisco Market, throughout Collin County and North Texas geographical area. He strives to establish business partnerships with local businesses, sports organizations, and charitable organizations. Lastly, he utilizes social media platforms such as LinkedIn for content creation and brand building to give back, build a better corporate brand and drive new business.
My Top 5 StrengthsFinder® Talents / Themes / Strengths
In addition to his day-to-day duties, Tomasz has a big heart for non-profit world. He serves extensively in several local non-profits such as the Boys & Girls Clubs of Collin County, American Heart Association and Project Unity. He genuinely believes that life becomes more fulfilling when we realize that our purpose is to pass the torch forward to those in need and those less fortunate.
As a hobby, he hosts "The Tomasz Show" podcast that features community leaders, entrepreneurs, and influential figures. The episodes are available anywhere you listen to the podcasts. They highlight respective guests' stories and their particular growth pillars that helped them get to where they are today. His objective is to help extract a lot of value around the areas of personal and professional development for the benefit of the next generation.
Tomasz is a graduate of Southern Methodist University (SMU) and SMU Men's Basketball Alum. He earned a BBA degree in Marketing as well as a Master of Arts degree in Dispute Resolution and Conflict Management. In addition, he is a certified mediator. He intends to graduate from Southwestern Graduate School of Banking at SMU in 2020 where he has a privilege to serve as President of Class 61. Tomasz currently resides in Aubrey, Texas with his wife Ashley and their dog Maverick. They love to travel and learn from different cultures and pathways of life.


text: "July Pyrra" wrote : | | 2008-06-04 15:16:41,348 DEBUG [org.jboss.ejb3.stateless.StatelessDelegateWrapper] Starting failed jboss.j2ee:jar=Project.jar,name=Update,service=EJB3 | java.lang.NullPointerException | at org.jboss.ejb.txtimer.DatabasePersistencePolicy.listTimerHandles(DatabasePersistencePolicy.java:165) | at sun.reflect.NativeMethodAccessorImpl.invoke0(Native Method) | at sun.reflect.NativeMethodAccessorImpl.invoke(NativeMethodAccessorImpl.java:39) | at sun.reflect.DelegatingMethodAccessorImpl.invoke(DelegatingMethodAccessorImpl.java:25) | | That looks like a DEBUG message. Probably you can ignore it. Or else, stop the server and delete the contents of the JBOSSHOME/server/myapp/data folder. Then start the server. "July Pyrra" wrote : | And please, could you precise if I had to add the sqljdbc.jar and the mysql-connector.jar y the deploy folder or put it in the lib folder would be enough? Since you are using MySQL database, you don't require the sqljdbc.jar file (which is the driver for MS-SQL server). You should not place the mysql-connector.jar in the deploy folder. It should be placed in the JBOSSHOME/server/myapp/lib folder. View the original post : http://www.jboss.com/index.html?module=bb&op=viewtopic&p=4155676#4155676 Reply to the post : http://www.jboss.com/index.html?module=bb&op=posting&mode=reply&p=4155676


text:
The Best Survival Movies Based on True Stories
List Rules No straight horror movies.
There are hundreds if not thousands of movies about survival, but only a choice few are based on true events. The survival movies based on true stories can offer harrowing examples of people performing extraordinary feats in the most desperate of circumstances, but those films are few and far between. Survival films often depict what really happened to people whose life is told in a story that could be inspirational or end in disaster.
You may think a survival movie like Tom Hanks' Castaway would wind up on a list like this one, but that film was based on multiple survival accounts of people who survived that sort of situation instead of a single, true event. The strict criteria for this list means you'll find the best survival movies - if and only if they are based on true events! Real stories about people surviving or struggling to overcome obstacles offer an interesting and sometimes difficult viewing of struggle, perseverance, and the will to overcome... but not everyone survives and often, these movies end in ways you may not desire...
The Mercy is a 2017 British biographical drama film directed by James Marsh. Amateur sailor Donald Crowhurst (Colin Firth) attempts to circumnavigate the globe alone as his family awaits his return. ...more on Wikipedia
Actors: Colin Firth, Rachel Weisz, David Thewlis, Ken Stott
Released: 2018
Directed by: James Marsh
The Great Alaskan Race is a 2019 American action-adventure film directed by Brian Presley. Set during the 1925 great race of mercy, a group of mushers travels 700 miles to save the children of Nome, Alaska from a deadly epidemic. ...more on Wikipedia
Actors: Brian Presley, Treat Williams, Brad Leland
Released: 2019
Directed by: Brian Presley
The Aeronauts is a 2019 independent biographical action-adventure film directed by Tom Harper. A Pilot (Felicity Jones) and scientist (Eddie Redmayne) find themselves in an epic fight for survival while attempting to make discoveries in a gas balloon. ...more on Wikipedia
Actors: Eddie Redmayne, Felicity Jones, Phoebe Fox, Himesh Patel, Vincent Perez, + more
Released: 2019
Directed by: Tom Harper
Jungle is a 2017 Australian biographical survival drama film directed by Greg McLean. An adventurer (Daniel Radcliffe) joins a group during an expedition into the Bolivian jungle, only to find themselves in a dangerous situation. ...more on Wikipedia
Actors: Daniel Radcliffe, Alex Russell, Thomas Kretschmann, Yasmin Kassim, Joel Jackson, + more
Released: 2017
Directed by: Greg McLean