Cellotriose and Cellotetraose as Inducers of the Genes Encoding Cellobiohydrolases in the Basidiomycete Phanerochaete chrysosporium

Abstract

The wood decay basidiomycete Phanerochaete chrysosporium produces a variety of cellobiohydrolases belonging to glycoside hydrolase (GH) families 6 and 7 in the presence of cellulose. However, no inducer of the production of these enzymes has yet been identified. Here, we quantitatively compared the transcript levels of the genes encoding GH family 6 cellobiohydrolase (cel6A) and GH family 7 cellobiohydrolase isozymes (cel7A to cel7F/G) in cultures containing glucose, cellulose, and cellooligosaccharides by real-time quantitative PCR, in order to evaluate the transcription-inducing effect of soluble sugars. Upregulation of transcript levels in the presence of cellulose compared to glucose was observed for cel7B, cel7C, cel7D, cel7F/G, and cel6A at all time points during cultivation. In particular, the transcription of cel7C and cel7D was strongly induced by cellotriose or cellotetraose. The highest level of cel7C transcripts was observed in the presence of cellotetraose, whereas the highest level of cel7D transcripts was found in the presence of cellotriose, amounting to 2.7 ؋ 10 6 and 1.7 ؋ 10 6 copies per 10 5 actin gene transcripts, respectively. These numbers of cel7C and cel7D transcripts were higher than those in the presence of cellulose. In contrast, cellobiose had a weaker transcription-inducing effect than either cellotriose or cellotetraose for cel7C and had little effect in the case of cel7D. These results indicate that cellotriose and cellotetraose, but not cellobiose, are possible natural cellobiohydrolase gene transcription inducers derived from cellulose.

Induction of lignocellulose-degrading enzymes in Neurospora crassa by cellodextrins

“…Our results indicate that deletion of the predicted major β-glucosidase genes enabled N. crassa to induce both transcription and secretion of the complete repertoire of cellulases upon exposure to cellobiose. In P. chrysosporium, cellobiose does not induce cellulase gene expression (10), although cellotriose or cellotetraose were able to do so. We hypothesize that in P. chrysosporium, as in N. crassa, any inducing affect of cellobiose is masked by its degradation to glucose by endogenous β-glucosidases, leading to catabolite repression.…”

“…Historically, studies examining cellulases in filamentous fungi have used transcription as a readout for enzyme induction (10,36). To determine whether the transcriptional response of the Δ3βG and Δ3βGΔcre strains in response to cellobiose corresponds to an increase in functional protein, we assessed secreted proteins and cellulase activity of the Δ3βG and Δ3βGΔcre strains in response to induction with either cellobiose or Avicel (SI Materials and Methods), as compared to WTcultures.…”

“…Cellobiose, the major soluble end product of cellulases, moderately induces cellulase gene expression and activity in Hypocrea jecorina (Trichoderma reesei) (5-7) and Aspergillus species (8), which are commonly used fungi for high-level enzyme production (9). However, cellobiose is unable to induce cellulase gene expression in the more distantly related fungus Phanerochaete chrysosporium, which instead responds to cellotriose or cellotetraose (10). Notably, the oligosaccharide sophorose, which can be generated by the transglycosylation of cellobiose by an extracellular β-glucosidase, acts as a potent inducer of cellulases in T. reesei (11-13), although differences in both gene expression and protein production are apparent in cellulose-versus sophoroseinduced cultures (14).…”

Time-Dependent Profiles of Transcripts Encoding Lignocellulose-Modifying Enzymes of the White Rot Fungus Phanerochaete carnosa Grown on Multiple Wood Substrates

“…Transcripts for actin and GAPDH have traditionally been used as standards in studies of P. chrysosporium (6,14,17). However, previous transcriptomic analysis of P. carnosa suggests that the abundance of these transcripts varies during the growth of this organism on different substrates, and that the level of transcripts encoding a chitin synthase (Chs-257626) is more stable (Fig.…”

Transcriptional Response of the Cellobiose Dehydrogenase Gene to Cello- and Xylooligosaccharides in the Basidiomycete Phanerochaete chrysosporium

“…Cellobiose dehydrogenase (CDH) activity of the culture filtrates was assayed using cellobiose and cytochrome c as described previously (17). For transcriptional analysis in cultures with purified xylooligosac-charides, P. chrysosporium was precultivated for 3 days and the mycelium was transferred into resting medium as previously described (18). After resting cultivation for 6 h, 100 M glucose, cellobiose, cellotriose, cellotetraose, cellopentaose, xylose, xylobiose, xylotriose, or xylotetraose was added to the medium.…”