New & Noteworthy

Yeast, Place your Bets

May 25, 2012

Life is a balancing act.  An organism needs to grow and divide as fast as possible in its current environment.  But it also needs to be able to survive when the environment changes.

One way nature has come up with to deal with this balancing act is called bet hedging.  Basically some members in a population grow well in one set of circumstances and another set grows well in another.

Now this makes obvious sense when looking at members of a species that vary genetically.  Where it gets interesting is when bet hedging happens in a clonal population.

The idea is that even though they share the same DNA, there are epigenetic differences that cause subtle variations in gene expression levels between individuals.  These differences in gene expression patterns result in altered survival rates under different circumstances.

This phenomenon has been difficult to study because researchers need to focus on individuals and not populations.  Growth curves in liter flasks are of little use.

But now Levy and coworkers have come up with a new high throughput assay that allows them to look at how a few individuals are growing.  This has allowed them to quantitate how different individuals grow in a population and why the slower growers and/or the elderly are better able to survive stress.

The assay uses time-lapse bright-field microscopy to look at tens of thousands of microcolonies all at once.  What they find is a wide range of growth rates.  Somewhere between 1.3-10% of microcolonies grow at less than half the rate of the population as a whole (the number depends on the strain). 

The researchers identified multiple genes that impacted the range of growth rates within a population without necessarily affecting the overall growth rate.  In other words, this phenomenon isn’t simply due to chance–there are key genetic factors that help determine the amount of individual to individual variation in a population.

Levy and coworkers focused on Tsl1p, a component of the trimeric complex that synthesizes trehalose.  What they found was that those cells that made more Tsl1p divided less often and so grew more slowly.  Remember again, this is in a clonal population.

Trehalose is thought to help preserve proper protein folding under stress.  So the idea is that some subset of individuals is primed for stress but that in turn, this preparation makes them grow more slowly.  And this is just what the researchers found.

When they subjected colonies to heat stress, those that made lots of trehalose were more likely to survive.  But the survivors didn’t stay slow growing for long.  After multiple generations, the population returned to the original growth rate with the original individual to individual variation.  The phenotype was reversible.

Finally the researchers discovered that older yeast cells tended to make more trehalose and so survived stress better.  It may be that as a yeast cell gets older, it makes more Tsl1p which helps to set up the range of growth rates among individuals.  This may be one way individual to individual variation has evolved in yeast.

Bet hedging is obviously a great way to ensure the survival of a clonal population. Under ideal conditions, the fast growers can grow like mad, spreading themselves far and wide.  But when conditions become more hostile, a few slower, tougher individuals can survive to keep the population alive.

Video showing that slow growers survive heat shock and then revert to fast growers.

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

Alternatives to Whale Puke

May 11, 2012

Imagine scouring the beaches of the world for balls of whale vomit.  People are willing to do this because finding one is like finding a huge gold nugget.  Perfume companies will pay around $10,000 per pound for this ambergris (which is the more scientific name for the stuff). 

Of course perfume companies would rather have a more reliable and less expensive source for their ambergris.  And it wouldn’t hurt to find one that was a little less ethically troubling than using products from an endangered animal.

Until recently their best bet was a chemical called cis-abienol from balsam firs.  While less murky ethically, this chemical is tricky to harvest and even trickier to synthesize in the lab.  The perfume industry could definitely use an alternative.  And researchers in the Bohlmann lab may be on their way to finding it.

In a recent study from this lab, Zerbe and coworkers used “…metabolite profiling, tissue-specific deep transcriptome sequencing and functional (i.e. biochemical) genomics…” to identify the key enzyme for making cis-abienol in balsam firs.  They next plan to put the gene into yeast and have the yeast crank out this chemical.

Finding the gene was not trivial.  The first thing they did was to identify that most of the cis-abienol was made in the bark and phloem of the balsam fir.  They then sequenced the transcriptome from these sources and looked for likely candidate genes.  For this last step, they used a curated library of 146 known terpene synthases.

They found 4 candidate genes (AbdiTPS1-4) and successfully cloned three of them.  They then went on to express these proteins in E. coli, purify them, and found that AbdiTPS4 catalyzed the synthesis of cis-abienol in vitro. They renamed the gene AbCAS at that point.

According to Dr. Bohlmann, the lab is now testing conditions for getting the enzyme to work in yeast.  If they can pull it off, the perfume industry will finally get a cheap and easy alternative to whale vomit.  Let’s see if they pass any savings down to the consumer…

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

Take SGD with you wherever you go!

May 7, 2012

SGD has just released an app for the iPhone, iPad and iPod touch ap(p)tly called “YeastGenome”, containing the latest Saccharomyces cerevisiae information from the database, available now as a free download on iTunes. Search by gene names, gene descriptions or simply browse for quick access to Gene Ontology annotations, mutant phenotypes and protein and genetic interaction data for your favorite genes – all at your fingertips!

Use YeastGenome to:

• Search using gene name or keywords
• Browse by genomic feature types
• Save your favorite genomic features
• Quickly see fundamental genomic feature information
• Find Gene Ontology terms, Phenotypes, Interactions, and References associated with genomic features

… with or without an internet connection!

How many ATP-dependent RNA helicases is the S. cerevisiae genome known to encode? Which proteins have a zinc finger motif? Now you can answer these questions and more with the YeastGenome app, whether you’re in line at the supermarket or having lunch with your colleagues or attending a seminar with no wifi-access! Get your friends and colleagues as fired up about The Awesome Power of Yeast as you are – use YeastGenome to email information about genomic features to collaborators and spread the word!

Please read our FAQ or visit iTunes for more information.

When Heat is Not the Burning Issue

May 4, 2012

When is a temperature sensitive mutation not a temperature sensitive mutation? When the process being studied is affected by temperature too.

Mutations conferring temperature sensitivity – that is, a phenotype that only appears at higher- or lower-than-normal growth temperature due to the loss or alteration of function of a gene product at that temperature – have for decades been an invaluable tool in dissecting many biological processes in yeast and other model organisms. Now there is reason to question whether some temperature sensitive phenotypes might actually be the result of a more complex interaction between multiple genes.

This synthetic genetic interaction is easy to mistake for the effect of a single mutation. A scientist starts out with a mutation that weakens a gene in a certain process. Unfortunately for the scientist, the process being studied is itself weakened by higher temperatures. The effect of the higher temperature combines with the effect of the mutation to shut down the process. The mutation looks temperature sensitive even when it isn’t.

And this does not appear to be merely a theoretical concern. As Paschini and coworkers show in a new study out in GENETICS, something similar may have happened with key mutations used to study telomere function in yeast.

These researchers looked at several mutations but we’ll focus on the work they did with cdc13-1. A key experiment that had been previously done with this mutation dealt with the effects of the loss of cdc13 function in the absence of RAD9.

Basically, researchers had found that prolonged incubation of a cdc13-1/Δrad9 strain at 36° severely compromised viability. These results were used to infer CDC13 function based on its loss at 36°. However, Paschini and coworkers provide compelling data that cdc13-1 behaves equally poorly at 23° and 36°.

First off, they showed that prolonged incubation of wild type yeast at the temperatures used in these studies (36°) resulted in shorter telomeres. They found very little effect on telomere length at 32°.

Next they showed that biochemically, cdc13-1 didn’t behave like a temperature sensitive mutation. Strains with cdc13-1 produced around 4-fold less protein at both 23° and 36° and the protein that was made bound telomeres equally well at both temperatures.

They argue from these two pieces of data that the loss of viability comes from a combination of the compromised cdc13-1 mutation and the effects of higher temperature on telomere function. Something is going on in the rad9 experiment but it is not due to an increased loss of CDC13 function at higher temperatures. There is some other factor involved that is being inhibited.

Of course it could be that cdc13-1 still confers temperature sensitivity, but that they didn’t have the right biochemical assay to see it. To address this issue, they generated five new mutations in cdc13 that behaved more like traditional temperature sensitive mutations.

They focused on one, cdc13-S611L, that was compromised for protein production at temperatures of 32° and above. They then repeated the rad9 double mutant experiment at 32° and 36°. They found that viability was compromised only at 36° even though Cdc13p was equally absent at both 32° and 36°. These results suggest that the loss of viability at 36° is not only the result of cdc13-1.

If this and other results hold up, scientists will need to rethink what previous experiments meant and they may need to modify their models. This should also get other researchers thinking about their temperature sensitive mutations.  It is important to confirm biochemically that a mutation indeed makes a specific gene product temperature sensitive. Because sometimes even if it quacks, it isn’t a duck…

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