Category Archives: sordariomycetes

Genomes on the horizon at JGI

Several more fungi are on the docket for sequencing at JGI through their community sequencing program. This includes

This complements an ever growing list of fungal genome sequences which is probably topping 80+ now not including the several dozen strains of Saccharomyces that are being sequenced at Sanger Centre and a separately funded NIH project to be sequenced at WashU.

Mechanism of riboswitch controlling mRNA splicing

A exciting research paper “Control of alternative RNA splicing and gene expression by eukaryotic riboswitches” published in Nature details the mechanism of how riboswitches work in Neurospora crassa. While riboswitches have been found and studied in bacteria there has not been extensive work showing how they work in fungi. In bacteria the riboswitch acts as the direct interacting sensor that switches gene expression off through a structural change in the RNA and fit in nicely with the RNA world view.

Using N. crassa, the authors show that alternative splicing is directly regulated through the thiamine metabolism genes which contains previously identified riboswitches. As also highlighted in the accompanying commentary this is also an interesting examples of direct RNA regulation of alternative splicing rather than through peptides like SR proteins.

Clusters of genomes

As announced at the Fungal Genetics meeting, the FGI at the Broad Institute is focusing on clusters of genomes rather than single ones. Some of genome projects are already grouped.

  • Coccidioides has 3 strains already plus the outgroup Uncinocarpus and conceivable one could include Histoplasma in there. This resources will grow to 14 strains (which comprise two species) of Coccidioides contributed by FGI and one from TIGR.
  • Aspergillus currently has 8 species sequenced with several in pipeline at Broad and TIGR.
  • Fusarium group has 3 species including recently released F. oxysporium.
  • The Candida clade also have several different already sequenced genomes and of course there is the already well studied (and well utilized genome resources I’ll add) for the Saccharomyces clade.
  • There are 4 genomes (well 5 but JEC21 and B-3501 are nearly identical) of Cryptococcus.

All in all a very exciting time for comparative genomics and I’m particularly intrigued to see how people will begin to use the resources.

This work to consolidate the clusters of genomes will, I hope, be very powerful. However, I still feel we are not doing a good job translating and centralizing information from different related species into a more centralized resource. Lots of money is spent on sequencing but I don’t know that we have realized the dream of having the comparative techniques illuminate the new genomes to the point that we are learning huge new things.

It seems to me, initially there is the lure of gathering low-hanging fruit from a genome analysis (which drives the first genome(s) paper), but not always the financial support of the longer term needs of the community to feed the experimental and functional work back into the genome annotation and interpretation.  The cycle works really well for Saccharomyces cerevisiae because the curators who work with the community to insure information is deposited and that literature is gleaned to link genomic and functional information. But this is expensive in terms of funding many curators for many different projects.

It seems as we add more genomes there isn’t a very centralized effort for this type of curatorial information and so we lack the gems of high-quality annotation that is only seen in a few “model” systems.  At some point a better meta-database that builds bridges between resource and literature rich “model system” communities may help, but maybe something new will have to be created? I like thinking about this as a user-driven content via a wiki which also dynamic (and versioned!) content from automated intelligent systems to map the straight-forward things.  Tools like SCI-PHY already exist that can do this and generate robust orthology groups (or Books as the PhyloFact database organizes them) for futher analysis. The SGD wiki for yeast is a start at this, but is mostly an import of SGD data into a mediawiki framework – I wonder how this can be built upon in a more explictly comparative environment.

More Neurospora genomes

We got word last week from the JGI that our DNA for Neurospora tetrasperma and N. discreta have passed QC and library QC and are on their way to being sequenced. The center also plans to do some EST sequencing to improve gene calling abilities.

Why more Neurospora genomes? The sequencing proposal discussed these species as a model system for evolutionary and ecological genetics. It will allow us and others to test several hypotheses about the molecular evolution of things like genome defense in Neurospora and to understand more about the evolutionary history of the model organism N. crassa.

Continue reading More Neurospora genomes

That was a lot of work

I’ve never worked with Magnaporthe grisea, the fungus responsible for rice blast, one of the most devastating crop diseases, but I do know that its life cycle is complicated and that knocking out roughly 61% of the genes in the genome and evaluating the mutant phenotype to infer gene function is not trivial. In their recent letter to Nature, Jeon et al did what many of us have dreamed of doing in our fungus of interest: manipulate every gene to find those that contribute to a phenotype of interest.

In their study, the authors looked for pathogenecity genes. Interestingly, the defects in appressorium formation and condiation had the strongest correlation with defects pathogenicity, suggesting that these two developmental stages are crucial for virulence. Ultimately, the authors identify 203 loci involved in pathogenecity, the majority of which have no homologous hits in the sequence databases and have no clear enriched GO functions. Impressively, this constitutes the largest, unbiased list of pathogenecity genes identified for a single species (though so of us, I’m sure, may have a problem with the term “unbiased”).

If you’d like to play with their data, the authors have made it available in their ATMT Database.

Fungal Genetics 2007 details

I’m including a recapping as many of the talks as I remember. There were 6 concurrent sessions each afternoon so you have to miss a lot of talks. The conference was bursting at the seams as it was- at least 140 people had to be turned away beyond the 750 who attended.

If there was any theme in the conference it was “Hey we are all using these genome sequences we’ve been talking about getting”. I only found the overview talks that solely describe the genome solely a little dry as compared to those more focused on particular questions. I guess my genome palate is becoming refined.

Continue reading Fungal Genetics 2007 details

Hello, do I know you?

Blogging about Peer-Reviewed ResearchSelf and non-self recognition is important for fungi when hyphae interact fuse if they should compartmentalize and undergo apoptosis to kill the heterokaryoton or exchange nutrients. This process is part of cell defense and to limit to the movement of mycoviruses.

A paper in PLOS ONE describes the Genesis of Fungal Non-Self Repertoire. This kind of work goes on down the hall from us as well in the Glass lab among others. This recent paper describes het genes, which contain WD40 repeats and different combinations of these help control specificity. There is of course a diverse literature on this subject especially in Neurospora, and I’m not reviewing it here, but it is an imporant process in understanding how fungi interact with their environment.

Neurospora crassa

Here is an image of Neurospora crassa I took today in my first attempt at squashes. These are from strains that Dave Jacobson grew up with his constructs so I can’t take any credit other than playing with the microscope next door. Now my first attempt came out badly, so this is actually Dave’s prep as well. And these got dry so they aren’t as nices as they could be. For much nicer images, see N.B. Raju’s.

All that said, I hope these quick images give a hint at the extremely cool structures these fungi produce. These 8-chain ascospores are the result of meoisis that took place inside the perithecia (which was squeezed gently to release the rosettes [or not too gently in my case]).

N.crassa rosetteN.crassa Histone GFP

( I was previous confused about the sample and had labeled this N. tetrasperma which has 4-chained ascospores [tetra] while this sample is crassa which has 8).

Deeper and Deeper, Down the Transcriptome-hole We Fall

Your eye contains the same genetic content as your fingernail, but these two tissues look nothing alike. One significant cause of this difference is the tissue specific regulation of the genes in the genome. In some tissues in your body, a gene may be expressed (transcribed) while that same gene may be silent in another tissue type. A great deal of modern biological research explores the regulation of expression of all the genes in a genome, collectively known as the transcriptome. Such studies are, for example, aimed at understanding which genetic regulation events account for the differences between an eye and a fingernail.

However, the effectiveness of this research is predicated upon actually knowing which parts of the genome are capable of being expressed and, subsequently, regulated. Conventionally, researchers extract RNA from an organism grown in various conditions (or, as in the case of our example, various tissues from an organism) and clone and sequence the RNA to identify at least a subset of genes that are expressed (Ebbole 2004*). Such Expressed Sequence Tags (ESTs) have proven vital to our understanding of gene and gene structure annotation as they frequently provide evidence of intron splice sites. While this method has facilitated a robust understanding of gene regulation, it is expensive, time consuming, and provides a relatively low coverage of the transcriptome. If our goal is to understand everything that is expressed, then we need a superior tool.

Enter SAGE (serial analysis of gene expression) and MPSS (massively parallel signature sequencing) [Irie 2003*, Harbers 2005*]. Both methods sequence short tags of a transcript’s 3′ end. SAGE uses conventional sequencing technology while MPSS uses Solexa, Inc.’s novel bead-based hybridization technology. One of the massive advantages of these technologies is the number of sequences they provide: large EST databases are on the order of several tens of thousands, while SAGE generally provides 100,000 to 200,00 tags and MPSS can provide over a million signatures. That being said, there are still questions regarding the sensitivity of the depth of coverage of the transcriptome. It may well be that despite a lower total sequence count, ESTs provide more information about what parts of the genome are expressed.

Fortunately, Gowda et al put all three methods to work as well as an RNA microarray (which doesn’t provide sequence, but enables its inference through hybridization) in their recent study of the Magnaporthe grisea transcriptome [Gowda 2006]. M. grisea is the causative agent of rice blast, a devastating disease that results in tremendous crop yield loss. The researchers evaluated two tissues types: the non-pathogenic mycelium and the invasive, plant penetrating appressorium.

Interestingly, 40% of the MPSS tags and 55% of the SAGE tags identified represent novel genes as they had no matches in the existing M. grisea JGI EST collection. Additionally, the authors found that no one method could identify the majority of the transcripts, but that a two-way combination of array data, MPSS or SAGE could provide over 80% of the total unique transcripts all of the methods identified. One additional suprise was that roughly a quarter of the genes identified also produced an antisense RNA, possibly for siRNA regulation of the gene.

The long story short appears to be that there is, as of yet, no magic bullet of a method. To adequately cover the transcriptome, multiple techniques are required.

*These references are, unfortunately, not located in an open access journal.