Comparative transcriptome and secretome analysis of wood decay fungi Postia placenta and Phanerochaete chrysosporium

Abstract

Cellulose degradation by brown rot fungi, such as Postia placenta, is poorly understood relative to the phylogenetically related white rot basidiomycete, Phanerochaete chrysosporium. To elucidate the number, structure, and regulation of genes involved in lignocellulosic cell wall attack, secretome and transcriptome analyses were performed on both wood decay fungi cultured for 5 days in media containing ball-milled aspen or glucose as the sole carbon source. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), a total of 67 and 79 proteins were identified in the extracellular fluids of P. placenta and P. chrysosporium cultures, respectively. Viewed together with transcript profiles, P. chrysosporium employs an array of extracellular glycosyl hydrolases to simultaneously attack cellulose and hemicelluloses. In contrast, under these same conditions, P. placenta secretes an array of hemicellulases but few potential cellulases. The two species display distinct expression patterns for oxidoreductase-encoding genes. In P. placenta, these patterns are consistent with an extracellular Fenton system and include the upregulation of genes involved in iron acquisition, in the synthesis of low-molecular-weight quinones, and possibly in redox cycling reactions.

FIG. 1.

Heat map showing hierarchical clustering of Phanerochaete chrysosporium genes exhibiting highly significant (P < 0.001) accumulation of transcripts in aspen-grown versus glucose-grown cultures. Numbers within boxes indicate the fold difference in transcripts for aspen versus glucose cultures. Only those P. chrysosporium genes exceeding a 4-fold difference are shown. Underlined P. chrysosporium models correspond to genes with >50 cDNA tags in medium containing powdered oak (43). On the right, a heat map shows transcript levels of the most closely related P. placenta sequences or, in the absence of a clear homolog, labeled as “None.” Protein IDs followed by asterisks correspond to peptides unambiguously identified by LC-MS/MS in filtrates from aspen-grown cultures. The log₂-based scales below maps are calibrated to the data shown above. Boldface denotes models in need of editing: GH10 xylanase model Ppl113670 has been superceded by an annotated preferred version, Ppl134783, which shows less pronounced upregulation (1.10) with a log₂ signal of 11.3. GH27 α-galactosidase model Ppl128150 has been superceded by annotated preferred version Ppl13470, which shows more pronounced upregulation (3.08) with a log₂ signal of 14.4 (Table 3; see also Table S1 in the supplemental material for detailed data).

FIG. 2.

Heat map showing hierarchical clustering of P. placenta genes exhibiting highly significant (P < 0.001) accumulation of transcripts in aspen-grown versus glucose-grown cultures. Numbers within boxes indicate the fold difference in transcripts in aspen versus glucose cultures. Only those P. placenta genes exceeding a 4-fold difference are shown. On right is a heat map showing transcript levels of the most closely related P. chrysosporium sequences or, in the absence of a clear homolog, labeled as “None.” Protein IDs followed by asterisks correspond to peptides unambiguously identified by MS in filtrates from aspen-grown cultures. Scales below maps show log₂-based signals. Underlined P. chrysosporium models correspond to genes with >50 cDNA tags in medium containing powdered oak (43).

FIG. 3.

Transcript levels of P. placenta genes encoding copper radical oxidases (Ppl130305, Ppl56703, and Ppl64380), GMC oxidoreductases with distant similarity to glucose oxidase (GOX; Ppl108489 and Ppl128830), an intact laccase (Ppl89382), a truncated laccase model (Ppl46931), and the highly expressed FAD-dependent oxidoreductase Ppl114192 (Table 4). Gray bars and the left axis represent log₂ microarray signals. Blackened bars and the right axis represent cDNA as determined by competitive RT-PCR. Asterisks indicate protein models whose peptides were identified in concentrated filtrates of aspen-grown cultures.