cAnaerobic gut fungi represent a distinct early-branching fungal phylum (Neocallimastigomycota) and reside in the rumen, hindgut, and feces of ruminant and nonruminant herbivores. The genome of an anaerobic fungal isolate, Orpinomyces sp. strain C1A, was sequenced using a combination of Illumina and PacBio single-molecule real-time (SMRT) technologies. The large genome (100.95 Mb, 16,347 genes) displayed extremely low G؉C content (17.0%), large noncoding intergenic regions (73.1%), proliferation of microsatellite repeats (4.9%), and multiple gene duplications. Comparative genomic analysis identified multiple genes and pathways that are absent in Dikarya genomes but present in early-branching fungal lineages and/or nonfungal Opisthokonta. These included genes for posttranslational fucosylation, the production of specific intramembrane proteases and extracellular protease inhibitors, the formation of a complete axoneme and intraflagellar trafficking machinery, and a near-complete focal adhesion machinery. Analysis of the lignocellulolytic machinery in the C1A genome revealed an extremely rich repertoire, with evidence of horizontal gene acquisition from multiple bacterial lineages. Experimental analysis indicated that strain C1A is a remarkable biomass degrader, capable of simultaneous saccharification and fermentation of the cellulosic and hemicellulosic fractions in multiple untreated grasses and crop residues examined, with the process significantly enhanced by mild pretreatments. This capability, acquired during its separate evolutionary trajectory in the rumen, along with its resilience and invasiveness compared to prokaryotic anaerobes, renders anaerobic fungi promising agents for consolidated bioprocessing schemes in biofuels production.

No abstract

SummaryXylan is the tnain carbohydrate found in the henlicellulosic fraction of plant tissues and accounls for one third of all renewable organic carbon aV'lilable on earlh.Xylanase, the rnajor cOluponent of an enzyillatic consortiUl11, acts in nature by depo(ynlerizing xylan JTIolecules into ITIonomeric penlosan units that are used by bacterial and fungal populations as a primary carbon source. Xylanase producers have been isolated fr0I11 all ecological niches where plant Inaterial is deposited, and nlicfoorganislTIs often contain lTIultiple loci encoding overlapping xylanolytic funct ions. The nUlllcrical excess of genes and the extensive sharing of structural features \vilhin f3-glycanase fcunilies suggests that extensive gene dupIicalion and conversion events have occurred during xyJanase evolution. Hydrolysis of J3-glycosidic linkages is sponsored by ,1 general acid catalytic reaclion conlnlon (0 all glycanases, whereas subslrale recognition is specified by subsites that interaCl \vith adjacent glycosyJ units. Under natural conditions xylanases are inducible by the products of their own action and subject to carbon catabolile repression. Bleaching paper pulps with xylanases is the first successful cOlnrnercial application for these cnzYlnes. The recovery of cellulosic textile fibers is the next logical application and bioconvcrsion of biomass into fuels and chemicals~renlains the ultitnate target Recent developInents have sho\vll that Jnetabolic path\\1ays can be transferred frOln one organiSI1110 another and proteins can be Jnodified to gain confonnational stability. suggesting that naturally occurring systclns can be clistOJn engineered to the situation in the fennentation tank. Thus, biotechnologies developed to tranSfOrIll bionulss into Inarketable products that gradually substitute Inaterials derived froIII non-rene\vablc resources are becolning cOJTIluercially worthwhile.

Summary Free‐living cells monitor extracellular ‘osmotic strength’ and respond metabolically to offset unfavourable osmotic intracellular solute concentrations. Here, we report the reconstruction of the Aspergillus nidulans salt stress‐controlling MAP kinase pathway, based on homology analysis with known yeast genes. In A. nidulans, salt stress HOG genes, such as pbsA, hogA, ptpA and msnA, are upregulated when exposed to high concentrations of salt and, in a hogA deletion mutant (SIK1), the accumulation of pbsA is strongly reduced, suggesting a salt‐specific feedback induction mechanism. Growth of SIK1 appears to be unchanged in unstressed cells, but hyphal extension rates are reduced by as much as 60% in the presence of salt. Microscopic observation revealed abnormal hyperbranched hyphal tips, disproportionate accumulation of nuclei and absence of septa. Thus, the inability to maintain turgor pressure depresses cell expansion and results in slower volume increases. In addition, SIK1 fails to partition the apical cell; thus, nuclei are not likely to arrest mitosis in interphase as in normal cells, but continue to divide, accumulating to high levels.

The Aspergillus nidulans brlA locus controls conidiophore development in conjunction with the products of several other regulatory loci. In this paper, we show that the brlA locus consists of overlapping transcription units, designated alpha and beta, with alpha transcription initiating within beta intronic sequences. The predicted BrlA polypeptides differ by 23 amino acid residues at their N‐termini. Targeted mutations specifically eliminating either the alpha or beta transcript led to developmental abnormalities similar to those produced by previously identified hypomorphic mutants, showing that both transcripts have essential functions for normal development. However, provision of additional doses of alpha in a beta‐ strain or of beta in an alpha‐ strain remediated the developmental defects, indicating that the polypeptides have redundant functions. It is likely that differential regulation of alpha and beta expression in the wild type is important for the initiation and temporal regulation of development.

The aim of the study was to demonstrate that the bZIP-type transcription factor AtfA regulates different types of stress responses in Aspergillus nidulans similarly to Atf1, the orthologous 'all-purpose' transcription factor of Schizosaccharomyces pombe. Heterologous expression of atfA in a S. pombe Deltaatf1 mutant restored the osmotic stress tolerance of fission yeast in surface cultures to the same level as recorded in complementation studies with the atf1 gene, and a partial complementation of the osmotic and oxidative-stress-sensitive phenotypes was also achieved in submerged cultures. AtfA is therefore a true functional ortholog of fission yeast's Atf1. As demonstrated by RT-PCR experiments, elements of both oxidative (e.g. catalase B) and osmotic (e.g. glycerol-3-phosphate dehydrogenase B) stress defense systems were transcriptionally regulated by AtfA in a stress-type-specific manner. Deletion of atfA resulted in oxidative-stress-sensitive phenotypes while the high-osmolarity stress sensitivity of the fungus was not affected significantly. In A. nidulans, the glutathione/glutathione disulfide redox status of the cells as well as apoptotic cell death and autolysis seemed to be controlled by regulatory elements other than AtfA. In conclusion, the orchestrations of stress responses in the aspergilli and in fission yeast share several common features, but further studies are needed to answer the important question of whether a fission yeast-like core environmental stress response also operates in the euascomycete genus Aspergillus.

SummaryCellular differentiation relies on precise and controlled means of gene expression that act on several levels to ensure a flexible and defined spatiotemporal expression of a given gene product. In our aim to identify transcripts enriched during fruiting body formation of the homothallic ascomycete Aspergillus (Emericella) nidulans, the grrA gene could be identified in a negative subtraction hybridization screening procedure. It encodes a protein similar to fungal F-box proteins, which function as substrate receptors for ubiquitin ligases, and that is highly related to the Saccharomyces cerevisiae regulatory protein Grr1p. Expression studies confirmed induction of grrA transcription and expression of its gene product during cleistothecial development of A. nidulans. Functional complementation of a yeast grr1D mutant was achieved by overexpression of the grrA coding sequence. A grrAD deletion mutant resembles the wild-type in hyphal growth, asexual sporulation, Hülle cell formation or development of asci-containing cleistothecia, but is unable to produce mature ascospores due to a block in meiosis as demonstrated by cytological staining of cleistothecial contents. Our results specify a particular involvement of the E3 ubiquitin ligase SCF GrrA in meiosis and sexual spore formation of an ascomyceteous fungus and shed light on the diverse functions of ubiquitin-proteasome-mediated protein degradation in eukaryotic development.

No abstract