New & Noteworthy

Can’t Get There Like That

May 04, 2016

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As HBO’s Silicon Valley scathingly relates, the mapping app from Apple was truly terrible when it was first launched. There are all kinds of funny (scary?) stories in which people following the directions ended up in the wrong place. (Click here for a few more of the epic fails.)

And sometimes it would show impossible ways to get from one location to the other. For example, to get to a certain place, my iPhone would recommend two different routes. After choosing the seemingly easiest route I quickly realized that it would take me through a building. Sometimes, even though it seems there are a couple of different ways to get somewhere, there is actually only one.

It turns out that this can be true in gene expression too. While you might increase expression by either mutating the promoter or duplicating the whole gene, sometimes only duplication is enough. There is just one route from here to there.

This point is driven home in a new study out in GENETICS by Rich and coworkers. Here they show that, under sulfate-limiting conditions, the only way that yeast can boost the expression of the SUL1 high affinity sulfate transporter enough to thrive is by duplicating it.

In many previous experiments, whenever yeast is starved for sulfate, after 100 generations or so the population almost always ends up selecting for a SUL1 duplication rather than increasing expression with a promoter mutation. This duplication results in a fitness advantage of 35% or more, which makes sense – when there isn’t much sulfate available, those cells that can get more of what’s there will outcompete their neighbors.

There are a couple of possible reasons things go down like this. It could be that the duplication is simply the most likely way to increase expression enough to survive, meaning that promoter mutations are possible but rare. Alternatively there may be no way to mutate the promoter enough to adequately increase the activity in such a short window of time. You simply can’t get to enough increased fitness by this route.

The first step in figuring out how to get somewhere is to map the roads in the area. This is also what Rich and coworkers needed to do – they needed to figure out how SUL1 is regulated. They did this in the standard way by nibbling away bits of the sequences upstream of the gene until there was a significant impact of gene expression. This is how they identified the 493 base pairs upstream of SUL1 as its promoter.

In the meantime, they also managed to develop a broadly applicable methodology for investigating any promoter – saturation mutagenesis, chemostat selection, and DNA sequencing to track variants.

The next step was to generate a library of mostly single point mutations in this promoter using error-prone PCR. As might be expected, most of the mutants had no effect or decreased gene expression but a few did increase activity. However, none of these last set increased expression as much as duplicating the gene.

They found 8 mutants that gave a 5% or better increase in fitness with the best being a 9.4% increase. Even when they combined these point mutations they could not increase the fitness much beyond 11%, not even half of the increase in fitness that an extra SUL1 gene gives. It seems that to get the most bang for its buck, yeast needs to duplicate SUL1. At least in the time frame of the experiment, that is.

Using what is essentially the scanning mutagenesis of this promoter, they were able to identify three sites that were important for SUL1 regulation. One site at -465 to -448 corresponded to a Cbf1-Met28-Met4 regulatory site, the second site at around -407 was most similar to either a Met31 or Met32 site and the third site at around -350 matched a Met32 site.

Mutations that resulted in increased activity tended to bring one of these three sites closer to the consensus transcription factor binding site. For example, the strongest point mutation, -353T>G, did this with the Met32 site at -350.

Even with these stronger consensus sequences for sulfate-regulatory transcription factors, none of these point mutations could get the yeast to where it needed to be in the fitness landscape in order to be able to thrive under sulfur limitation. Point mutations just can’t hold a candle to simply duplicating the SUL1 gene. Sometimes there really is just one way to get from point A to point B.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: adaptation, chemostat selection, fitness, gene duplication, saturation mutagenesis

Sign Up Now for the Next SGD Webinar: May 4th, 2016

April 26, 2016

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If you’re not already using YeastMine to answer all your questions about S. cerevisiae genes and gene products…you should be! SGD’s YeastMine is a powerful search tool that can retrieve, compare, and analyze data on thousands of genes at a time, greatly reducing the time needed to answer real, practical research questions. Through YeastMine, questions such as “What proportion of plasma membrane proteins are essential?” or “How many different gene products physically interact with the mitochondrial ribosome?” can be answered within minutes.

Next week on May 4th, 2016 at 9:30 AM PDT, we will provide a brief webinar tutorial on how to run queries and create gene lists in SGD’s YeastMine. As a practical demonstration of YeastMine, we will also showcase a research scenario in which yeast-human homology data is used to predict potential chemotherapy targets for human cancers.

Space for this webinar is limited. To reserve your spot, please register for the event using this online form: http://bit.ly/registerFor2ndSGDwebinar

This is the second episode of the SGD Webinar Series. If you missed the first one, or if you’d like to find out more about upcoming webinars, please visit our wiki page: SGD Webinar Series.

Categories: Announcements, Yeast and Human Disease

Chocolate and Coffee Too?

April 20, 2016

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Most of us know about yeast’s big part in making bread and booze. But those aren’t yeast’s only wonderful gifts. It also plays a big role in chocolate and coffee too. Is there anything this marvelous microorganism can’t do?

A new study by Ludlow and coworkers in Current Biology set out to look at the strains involved in cacao and coffee fermentation. Unlike the extensively studied wine strains, these have mostly been ignored up until now.

These researchers found that cacao, coffee and wine strains were very much different from one another. And they also found that unlike wine yeasts, which are pretty much the same most everywhere in the world, coffee and cacao strains are different depending on where they come from.

But the cacao and coffee strains did have one thing in common. Each had a lot in common with the other strains in its country and even on its continent.

So, for example, coffee strains from all over South America are very similar to each other but different from the coffee strains from Africa. And the opposite is true as well. African coffee strains are all pretty similar to each other but different from the South American strains. The same sort of thing is true for cacao strains as well.

You can think of coffee and cacao strains as the large, flightless birds of the yeast universe. Like a rhea, emu, or ostrich, they stay on their own continent. Wine yeasts are more like those chickens that are basically the same worldwide because humans have taken them along with them in their migrations.

The first step in their analysis was for Ludlow and coworkers was to get a hold of a bunch of different samples of cacao and coffee yeast strains from all over the world. They managed to get 78 cacao strains from 13 different countries and 67 coffee strains from 14 countries. The countries were from Central and South America, Africa, Indonesia and the Middle East.

The next step was to compare the genomes of these strains with each other and with the wine strains. They decided to use a technique called restriction site-associated DNA sequencing, or RAD-seq, that would give them an in depth look at around 3% of the yeast genome. As there was already a database with 35 wine strains that used the same method, Ludlow and coworkers only needed to generate data for their newly isolated strains.

These data revealed that coffee and cacao yeast strains were very different from one another. It also showed that the closer two coffee or cacao strains were to each other geographically, the closer they were together genetically. Using just these strains they could accurately predict the country of origin for a coffee yeast strain 79% of the time and 86% of the time for those associated with cacao.

The researchers next expanded the number of species they compared by using two additional databases. One included RAD-seq data from 262 strains from a wide variety of different places while the other contained another 57 strains.

This analysis generated 12 distinct groups of yeast, many of which had been identified before. Their new strains formed four new groups which they called South America Cacao, Africa Cacao, South America Coffee and Africa Coffee.

By comparing the four groups to the eight older ones, they were able to see that the new groups were not novel but instead were made up of mixtures of some of the other known groups. And who they shared alleles with depended on where the yeast strain was located. So, for example, the two South America groups shared alleles with the North American Oak group while the two African groups shared alleles with the Asian and European groups.

It looks like that, unlike with wine where there are a limited set of best yeast strains that most people use, the yeast strains involved with making chocolate and coffee are continent and even country specific. This suggests that people either took their wine yeast with them when they moved or that cross contamination throughout the world resulted in homogenization of wine yeasts. Sort of like the chickens Pacific Islanders brought with them whenever they settled on a new island.

It is a very different story for cacao and coffee yeast strains. While there was cross contamination within countries and even continents, there was no worldwide contamination. The rhea stayed in South America and the ostrich in Africa.

Wherever these strains come from, they work hard to make our chocolate, coffee, bread, wine, and beer. What an amazing bounty! Make some room, Dog, good ol’ Saccharomyces cerevisiae is joining you as man’s best friend.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: admixture, migration, population diversity

Lessons from Yeast: Poisoning Cancer

April 06, 2016

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In the book Dune, the mentat Thufir Hawat is captured by the evil Harkonnens and given a residual poison. He can only stay alive by getting a constant dose of the antidote. Once it is withdrawn, he will die.

A new study in the journal GENETICS by Dodgson and coworkers shows that the same sort of thing can happen to yeast that carry an extra chromosome. In this case, certain genes on the extra chromosome turn out to be like the residual poison. And a second gene turns out to be the antidote.

Once that second gene is deleted, the yeast cell dies. It has been deprived of its antidote.

This synthetic lethal phenotype isn’t just a cool finding in yeast either. Cancer cells invariably have extra and missing chromosomes. If scientists could find similar “antidote genes” in specific types of cancers and target them, then the cancer cell would die. And this would happen without damaging the other cells of the body that have a typical number of chromosomes.

The first thing these researchers did was to make separate yeast strains each with an extra chromosome I, V, VIII, IX, XI, XII, or XVI. The next step was to see what happens when every gene was deleted individually, one at a time, from each strain.

As expected, these yeast did pretty well when a gene on the extra chromosome was deleted. So, for example, a strain with an extra chromosome I tolerated a gene deleted from chromosome I. This makes sense as this just brings that gene back to its normal copy number.

But this was not the case with chromosomes VIII and XI. Here deleting genes on the extra chromosome often had a negative effect. This suggested that the screen probably had a high number of false positives and these researchers later confirmed this.

Likely reasons for the high number of false positives include the strain with the extra chromosome being W303 and the deletion strain being S288C, errors in the deletion collection itself, and what they refer to as neighboring gene effects. Basically this last one is the effect that deleting a gene has on nearby genes.

Once Dodgson and coworkers corrected for these problems, they found two broad sets of phenotypes – general and chromosome specific.

The general ones were the ones shared by most or all of the strains. These were deletions that affected the yeast no matter which chromosome they had an extra copy of.

For the most part, these genes were enriched for the Gene Ontology (GO) term vesicle-mediated transport, indicating that they have something to do with the transportation of substances in membrane-bounded vesicles. For example, deletion of MNN10, HOC1, and MNN11, genes all involved in protein transport and membrane-related processes, had a negative effect on many of the yeast strains with an extra chromosome. Consistent with this, brefeldin A, a drug that targets protein trafficking, negatively affected most of the strains.

Another gene that affected many of these strains when deleted was TPS1. This gene encodes a subunit of trehalose-6-phosphate synthase, a key enzyme for making trehalose, a molecule that helps yeast deal with stress. Perhaps not surprisingly, having an extra chromosome is stressful!

In addition to the genes that affect many strains with an extra chromosome, there were also genes that were chromosome specific. The best characterized of these was the EDE1 gene in the strain with an extra chromosome IX. Deleting EDE1 in this strain increased its doubling time by more than 80 minutes while only causing an increase of 5 minutes in the doubling time of wild type yeast. This was a severe phenotype in their assay.

They next tried to find which gene on chromosome IX might be responsible for the severe effect of deleting EDE1. Since EDE1 is known to be involved in endocytosis, they looked for genes involved in the same process. And they found one – PRK1.

The strain with a deleted EDE1 gene and an extra chromosome IX was rescued by deleting one copy of the PRK1 gene. The extra PRK1 gene was the poison and the EDE1 gene was the antidote.

If a similar pair could be found in cancers that often have the same set of extra chromosomes, then perhaps scientists could develop drugs that target an antidote gene. Now the cancer cells would die and the “normal” cells would be fine. Thanks again, yeast, for pointing us toward new ways to treat human disease.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight, Yeast and Human Disease

Tags: aneuploidy, cancer, synthetic lethal

Apply Now for the 2016 Yeast Genetics & Genomics Course

March 30, 2016

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For almost 50 years, the legendary Yeast Genetics & Genomics course has been taught each summer at Cold Spring Harbor Laboratory. (OK, the name didn’t include “Genomics” in the beginning…). The list of people who have taken the course reads like a Who’s Who of yeast research, including Nobel laureates and many of today’s leading scientists.

The application deadline is April 15th, so don’t miss your chance! Find all the details and application form here.

This year’s instructors – Grant Brown, Maitreya Dunham, and Marc Gartenberg – have designed a course (July 26 – August 15) that provides a comprehensive education in all things yeast, from classical genetics through up-to-the-minute genomics. Students will perform and interpret experiments, learning about things like:

• How to Find and Analyze Yeast Information Using SGD
• Isolation and Characterization of Mutants
• Transformation of Plasmids & Integrating DNAs
• Meiosis & Tetrad Dissection as well as mitotic recombination
• Synthetic Genetic Array Analysis
• Next-Gen. whole-genome and multiplexed DNA barcode sequencing
• Genome-based methods of analysis
• Visualization of yeast using light and fluorescence microscopy
• Exploring synthetic biology with CRISPR/CAS9-directed engineering of biosynthetic pathways

Techniques have been summarized in a completely updated course manual, which was recently published by CSHL Press.

Scientists who aren’t part of large, well-known yeast labs are especially encouraged to apply – for example, professors and instructors who want to incorporate yeast into their undergraduate genetics classrooms; scientists who want to transition from mathematical, computational, or engineering disciplines into bench science; and researchers from small labs or institutions where it would otherwise be difficult to learn the fundamentals of yeast genetics and genomics. Significant stipends (in the 30-50% range of total fees) are available to individuals expressing a need for financial support and who are selected into the course.

Besides its scientific content, the fun and camaraderie at the course is also legendary. In between all the hard work there are late-night chats at the bar and swimming at the beach. There’s a fierce competition between students at the various CSHL courses in the Plate Race, which is a relay in which teams have to carry stacks of 40 Petri dishes (used, of course). There’s also a sailboat trip, a microscopy contest, and a mysterious “Dr. Evil” lab!

Last year’s Yeast Genetics & Genomics Course was loads of fun – don’t miss out!

Categories: Announcements

Updated Genome Browser

March 27, 2016

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In an effort to provide a comprehensive view of sequence-based functional elements in Saccharomyces cerevisiae, we have upgraded our genome browser, and added new data tracks, to allow users to quickly and easily browse the information-rich yeast genome. We invite authors to work with us to integrate published data into our new JBrowse genome viewer pre- and/or post-publication. Please contact us if you are interested in participating or have questions and comments. Watch for the regular addition of new tracks to SGD’s JBrowse in the future!

Take a look at our newest video tutorial to get acquainted with JBrowse, and let us know if you have any questions or suggestions.

For more SGD Help Videos, visit our YouTube channel, and be sure to subscribe so you don’t miss anything!

Categories: Data updates, Website changes

Sometimes Simple is Better

March 23, 2016

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Diagnosing why something has gone wrong in a complicated system can be difficult. There are so many bells and whistles that you can easily get lost.

That’s why it can sometimes help to turn to a simple system and then apply what you have learned to the more complicated one. This will, of course, sound familiar to any scientists studying that marvel of a model organism, Saccharomyces cerevisiae.

For example, it is amazing what you can glean from this yeast about human brain and blood diseases. Even though, of course, baker’s yeast has neither blood nor a brain!

This becomes very clear in a study out in PLOS Genetics by Fernandez-Murray and coworkers. In this study they use yeast to help figure out why mutations in the SLC25A38 gene in people leads to something called congenital sideroblastic anemia. And even better, their work hints at a possible treatment.

People with sideroblastic anemia make too little hemoglobin in their red blood cells and have too much iron in the mitochondria close to the nucleus (perinuclear mitochondria). The current treatment for this condition is not ideal, involving lots of transfusions and iron chelation.

Sometimes people are born with this anemia and sometimes people get it later in life. One subset of the inherited version happens because a gene with an unknown function, SLC25A38, isn’t working correctly. This group of patients is the focus of this study.

Fernandez-Murray and coworkers started out by using yeast to figure out what the yeast homolog, HEM25, does in a yeast cell. When the gene was deleted, the cells made about 50% less heme than wild type yeast and adding back the human gene, SLC25A38, to this deletion strain restored heme levels. Looks like they had made a yeast model of this inherited anemia.

Previous work had suggested that SLC25A38 might be a glycine or serine transporter and the next set of experiments confirmed it as a glycine transporter in a couple of ways. In both, they took advantage of cases in which yeast can use glycine as their sole nitrogen source if the glycine can make it into the mitochondria.

In the first case, they showed that yeast cells deleted for HEM25 grew poorly on plates where glycine was available as the only nitrogen source. In the second case, they showed that cells deleted for both SER1 and HEM25 grew poorly on plates where again glycine was the only nitrogen source. This last result confirms HEM25 as a glycine transporter since yeast deleted for the SER1 phosphoserine aminotransferase can only grow in the absence of serine if they can get glycine into their mitochondria. (There isn’t space to go into it here, but they also showed that HEM25 was not a serine transporter.)

OK so now they had created a yeast cell that mimicked the effect of sideroblastic anemia and figured out why people with a mutated SLC25A38 gene had the condition. Now it was time to find a treatment.

The researchers came up with three possibilities. The first treatment was just to give the yeast extra glycine, the second was to drive glycine synthesis within the cell by adding a lot of serine, and the third was to add a downstream precursor of heme synthesis, 5-aminolevulinic acid (5-Ala).

They tested each scenario on yeast cells deleted for HEM25 and found that both glycine and 5-Ala worked to restore heme synthesis, but that added serine had no effect. Both glycine and 5-Ala returned heme levels to that seen in wild type.

Of course we aren’t yeast, so they next tested their treatment on something a bit more complicated — zebrafish. By using morpholino technology to knock down both copies of the zebrafish SLC25A38 homolog, SLC25A38a and SLC25A38b, the researchers managed to lower a zebrafish’s heme levels to about 50% of normal.

When they gave these zebrafish extra glycine or 5-Ala, their heme levels did not improve. They were still anemic!

After a bit of thought, the researchers realized that folate might be what the zebrafish were missing. In work that we didn’t have time to go over before, the researchers had shown that a folate dependent pathway was critical for getting heme levels up to normal.

Yeast could get away without added folate in these experiments because they make their own. However, zebrafish, like people, do not.

So the final step was to try to add both glycine and folate to these fish. Now the zebrafish’s heme levels returned to about 80% of normal.

These results suggest a better treatment for some people with sideroblastic anemia — added folate and glycine. And it all came from studying the problem in the simpler, bloodless S. cerevisiae. Nice work again yeast.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: anemia, human disease, model organism, zebrafish

Join SGD at The Allied Genetics Conference

March 22, 2016

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SGD will be attending The Allied Genetics Conference (TAGC) in Orlando, Florida, July 13–17, 2016! For the first time ever, the meetings of the yeast, C. elegans, ciliate, Drosophila, mouse, and zebrafish model organism communities will be united under one roof, along with a new meeting on population, evolutionary, and quantitative genetics.

Submit your abstracts now! Abstract submission closes March 23, 2016, but advance registration is available until June 29. If you want GREAT science and access to the leaders of the field, then TAGC is the place for you. SGD will be there, will you?

Categories: Conferences

New SGD Help Video: YeastMine Scenario

March 17, 2016

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If you’re not already using YeastMine to answer all your questions about the Saccharomyces cerevisiae genome and the gene products it encodes…you should be! YeastMine enables slicing and dicing of data from SGD in any way you choose. Ask questions like “Which genes can mutate to confer oxidative stress resistance, and what biological processes are they involved in?” or “Are there any undiscovered subunits of the mitochondrial ribosome?”

Take a look at our newest video tutorial to dig into YeastMine, and let us know if you have any questions or suggestions.

For more SGD Help Videos, be sure to visit our YouTube channel!

Categories: Tutorial

SGD March 2016 Newsletter

March 15, 2016

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SGD periodically sends out its newsletter to colleagues designated as contacts in SGD. This March 2016 newsletter is also available on the community wiki. If you would like to receive the SGD newsletter in the future please use the Colleague Submission/Update form to let us know.

Categories: Newsletter

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