Brown-rot fungi such as Postia placenta are common inhabitants of forest ecosystems and are also largely responsible for the destructive decay of wooden structures. Rapid depolymerization of cellulose is a distinguishing feature of brown-rot, but the biochemical mechanisms and underlying genetics are poorly understood. Systematic examination of the P. placenta genome, transcriptome, and secretome revealed unique extracellular enzyme systems, including an unusual repertoire of extracellular glycoside hydrolases. Genes encoding exocellobiohydrolases and cellulose-binding domains, typical of cellulolytic microbes, are absent in this efficient cellulose-degrading fungus. When P. placenta was grown in medium containing cellulose as sole carbon source, transcripts corresponding to many hemicellulases and to a single putative ␤-1-4 endoglucanase were expressed at high levels relative to glucose-grown cultures. These transcript profiles were confirmed by direct identification of peptides by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Also upregulated during growth on cellulose medium were putative iron reductases, quinone reductase, and structurally divergent oxidases potentially involved in extracellular generation of Fe(II) and H2O2. These observations are consistent with a biodegradative role for Fenton chemistry in which Fe(II) and H2O2 react to form hydroxyl radicals, highly reactive oxidants capable of depolymerizing cellulose. The P. placenta genome resources provide unparalleled opportunities for investigating such unusual mechanisms of cellulose conversion. More broadly, the genome offers insight into the diversification of lignocellulose degrading mechanisms in fungi. Comparisons with the closely related white-rot fungus Phanerochaete chrysosporium support an evolutionary shift from white-rot to brown-rot during which the capacity for efficient depolymerization of lignin was lost.cellulose ͉ fenton ͉ lignin ͉ cellulase ͉ brown-rot

Efficient lignin depolymerization is unique to the wood decay basidiomycetes, collectively referred to as white rot fungi. Phanerochaete chrysosporium simultaneously degrades lignin and cellulose, whereas the closely related species, Ceriporiopsis subvermispora, also depolymerizes lignin but may do so with relatively little cellulose degradation. To investigate the basis for selective ligninolysis, we conducted comparative genome analysis of C. subvermispora and P. chrysosporium . Genes encoding manganese peroxidase numbered 13 and five in C. subvermispora and P. chrysosporium , respectively. In addition, the C. subvermispora genome contains at least seven genes predicted to encode laccases, whereas the P. chrysosporium genome contains none. We also observed expansion of the number of C. subvermispora desaturase-encoding genes putatively involved in lipid metabolism. Microarray-based transcriptome analysis showed substantial up-regulation of several desaturase and MnP genes in wood-containing medium. MS identified MnP proteins in C. subvermispora culture filtrates, but none in P. chrysosporium cultures. These results support the importance of MnP and a lignin degradation mechanism whereby cleavage of the dominant nonphenolic structures is mediated by lipid peroxidation products. Two C. subvermispora genes were predicted to encode peroxidases structurally similar to P. chrysosporium lignin peroxidase and, following heterologous expression in Escherichia coli , the enzymes were shown to oxidize high redox potential substrates, but not Mn 2+ . Apart from oxidative lignin degradation, we also examined cellulolytic and hemicellulolytic systems in both fungi. In summary, the C. subvermispora genetic inventory and expression patterns exhibit increased oxidoreductase potential and diminished cellulolytic capability relative to P. chrysosporium .

Identification of specific genes and enzymes involved in conversion of lignocellulosics from an expanding number of potential feedstocks is of growing interest to bioenergy process development. The basidiomycetous wood decay fungi Phanerochaete chrysosporium and Postia placenta are promising in this regard because they are able to utilize a wide range of simple and complex carbon compounds. However, systematic comparative studies with different woody substrates have not been reported. To address this issue, we examined gene expression of these fungi colonizing aspen (Populus grandidentata) and pine (Pinus strobus). Transcript levels of genes encoding extracellular glycoside hydrolases, thought to be important for hydrolytic cleavage of hemicelluloses and cellulose, showed little difference for P. placenta colonizing pine versus aspen as the sole carbon source. However, 164 genes exhibited significant differences in transcript accumulation for these substrates. Among these, 15 cytochrome P450s were upregulated in pine relative to aspen. Of 72 P. placenta extracellular proteins identified unambiguously by mass spectrometry, 52 were detected while colonizing both substrates and 10 were identified in pine but not aspen cultures. Most of the 178 P. chrysosporium glycoside hydrolase genes showed similar transcript levels on both substrates, but 13 accumulated >2-fold higher levels on aspen than on pine. Of 118 confidently identified proteins, 31 were identified in both substrates and 57 were identified in pine but not aspen cultures. Thus, P. placenta and P. chrysosporium gene expression patterns are influenced substantially by wood species. Such adaptations to the carbon source may also reflect fundamental differences in the mechanisms by which these fungi attack plant cell walls.

No abstract

Lignin peroxidases (LiP) ofPhanerochaete chrysosporium are encoded by a family of six closely related genes.Five LiP genes have been localized to the same dimorphic chromosome. In this investigation, relative transcript levels of the LiP genes were determined. Transcripts of the LiPA, LiPB, and 0282 genes were at similar levels in both carbon-and nitrogen-limited cultures. In contrast, transcription of the GLG5, V4, and GLG4 genes was dramatically altered by culture conditions. Under carbon-limited conditions, GLG4 transcripts were, by far, the most abundant. Southern blot analyses of clamped homogeneous field gels were used to map the GLG4 gene to a dimorphic chromosome separate from the other LiP genes.