What can comparative genomics tell us about species concepts in the genus Aspergillus?

Understanding the nature of species" boundaries is a fundamental question in evolutionary biology. The availability of genomes from several species of the genus Aspergillus allows us for the first time to examine the demarcation of fungal species at the whole-genome level. Here, we examine four case studies, two of which involve intraspecific comparisons, whereas the other two deal with interspecific genomic comparisons between closely related species. These four comparisons reveal significant variation in the nature of species boundaries across Aspergillus. For example, comparisons between A. fumigatus and Neosartorya fischeri (the teleomorph of A. fischerianus) and between A. oryzae and A. flavus suggest that measures of sequence similarity and species-specific genes are significantly higher for the A. fumigatus - N. fischeri pair. Importantly, the values obtained from the comparison between A. oryzae and A. flavus are remarkably similar to those obtained from an intra-specific comparison of A. fumigatus strains, giving support to the proposal that A. oryzae represents a distinct ecotype of A. flavus and not a distinct species. We argue that genomic data can aid Aspergillus taxonomy by serving as a source of novel and unprecedented amounts of comparative data, as a resource for the development of additional diagnostic tools, and finally as a knowledge database about the biological differences between strains and species.

Whole genome comparison of the A. fumigatus family.

The availability of the genome sequences of multiple Aspergillus spp. presents the research community with an unprecedented opportunity for discovery. The genomes of Neosartorya fischeri and Aspergillus clavatus have been sequenced in order to extend our knowledge of Aspergillus fumigatus, the primary cause of invasive aspergillosis. Through comparative genomic analysis, we hope to elucidate both obvious and subtle differences between genomes, developing new hypotheses that can be tested in the laboratory. A preliminary examination of the genomes and their predicted proteomes reveals extensive conservation between protein sequences and significant synteny, or conserved gene order. Comparative genomic analysis at the level of these closely related aspergilli should provide important insight into the evolutionary forces at play and their effect on gene content, regulation and expression.

Endogenous targets of transcriptional gene silencing in Arabidopsis.

Transcriptional gene silencing (TGS) frequently inactivates foreign genes integrated into plant genomes but very likely also suppresses an unknown subset of chromosomal information. Accordingly, RNA analysis of mutants impaired in silencing should uncover endogenous targets of this epigenetic regulation. We compared transcripts from wild-type Arabidopsis carrying a silent transgene with RNA from an isogenic transgene-expressing TGS mutant. Two cDNA clones were identified representing endogenous RNA expressed only in the mutant. The synthesis of these RNAs was found to be released in several mutants affected in TGS, implying that TGS in general and not a particular mutation controls the transcriptional activity of their templates. Detailed analysis revealed that the two clones are part of longer transcripts termed TSI (for transcriptionally silent information). Two major classes of related TSI transcripts were found in a mutant cDNA library. They are synthesized from repeats present in heterochromatic pericentromeric regions of Arabidopsis chromosomes. These repeats share sequence homology with the 3' terminal part of the putative retrotransposon Athila. However, the transcriptional activation does not include the transposon itself and does not promote its movement. There is no evidence for a general release of silencing from retroelements. Thus, foreign genes in plants encounter the epigenetic control normally directed, at least in part, toward a subset of pericentromeric repeats.

Disruption of the plant gene MOM releases transcriptional silencing of methylated genes.

Epigenetic modifications change transcription patterns in multicellular organisms to achieve tissue-specific gene expression and inactivate alien DNA such as transposons or transgenes. In plants and animals, DNA methylation is involved in heritability and flexibility of epigenetic states, although its function is far from clear. We have isolated an Arabidopsis gene, MOM, whose product is required for the maintenance of transcriptional gene silencing. Mutation of this gene or depletion of its transcript by expression of antisense RNA reactivates transcription from several previously silent, heavily methylated loci. Despite this, the dense methylation at these reactivated loci is maintained even after nine generations, indicating that transcriptional activity and methylation pattern are inherited independently. The predicted MOM gene product is a nuclear protein of 2,001 amino acids containing a region similar to part of the ATPase region of the SWI2/SNF2 family, members of which are involved in chromatin remodelling. MOM is the first known molecular component that is essential for transcriptional gene silencing and does not affect methylation pattern. Thus, it may act downstream of methylation in epigenetic regulation, or be part of a new pathway that does not require methylation marks.

High-efficiency transformation of Arabidopsis thaliana with a selectable marker gene regulated by the T-DNA 1' promoter.

Two selectable marker genes harbouring the bar coding region but differing in their promoters were compared in an Arabidopsis thaliana transformation assay using in planta infiltration with Agrobacterium tumefaciens. Surprisingly, in four Arabidopsis ecotypes examined, the 1' promoter from the right T-DNA was superior to the most commonly used 35S promoter of cauliflower mosaic virus (CaMV). The ecotype Wassilewskija gave the highest transformation frequencies, with an average of between 5.3 and 6.3% of the seedlings subjected to the selection. This is approximately 30-fold higher than previously reported results. Analysis of T-DNA integration patterns in single transformed plants or pooled populations revealed independent T-DNA integration events in each case. Results show that the 1' promoter is an attractive alternative to the 35S promoter for the generation of T-DNA insertion lines. The 1' promoter may be especially beneficial for the secondary transformation of transgenic strains containing the 35S promoter to exclude homology-mediated gene silencing.