Regulation of cellulolytic genes by ClrB-2 in Penicillium oxalicum

Cellulolytic fungi have evolved sophisticated systems to adapt their natural habitat. Penicillium oxalicum has been proved to be a promising cellulase producer. Regulation of cellulase production occurs at multiple levels of cell responses. Cellulolytic transcription factors (ClrB, ClrB-2, CreA, XlnR, and AmyR) were screened and identified to be involved in cellulose deconstruction by P. oxalicum. ClrB-2 encodes a conserved fungal Zn₂Cys₆ binuclear cluster domain and shares 37% identity with CLR-2 in N. crassa and a 45% identity with P. oxalicum ClrB. It was found that the cellulase expression level was activated by ClrB-2 and overexpression of clrB-2 facilitates the synergistic induction expression of cellulase genes in P. oxalicum. The transcriptional expression of ClrB-2 is responsive to ClrB, XlnR, CreA and AmyR, which implied that ClrB-2 may mediate the cascade transcriptional regulation for cellulolytic genes by the factors. ClrB-2 and ClrB additively regulated cellulolytic gene expression. Nonetheless, simultaneous overexpression of ClrB and lack of Bgl2 caused strong synergistic effects on induction expression for cellulase genes under non-inducing conditions, the further deletion of clrB-2 markedly decreased transcription levels for cellulase genes, suggesting that ClrB-2 is one of the most interesting target in P. oxalicum cellulolytic regulatory networks, which also signified that ClrB-2 and ClrB were tightly involved in cellulase expression regulatory during energy abstinence. These results demonstrate these differential regulatory roles for ClrB-2 in cellulase expression under inducing/non-inducing conditions, and provide a basis for the rational design of cellulase super producers.