Genome Wide Screening The Cellulase Pro-Duction Of C₂H₂ Transcription Factor Family Genes Knock Out Of Neurospora Crassa

Fungal lignocellulose degrading enzymes genes could be induced by the soluble in-ducing molecules, and fungal cellulase gene expression is tightly controlled by transcrip-tional activators and repressers. Therefore, identification and characterization of transcrip-tional factors involved in cellulase gene expression could be great useful for improving the production of lignocellulolytic enzymes.In order to discover the novel transcriptional factor genes involved in cellulase gene expression,57 mutants of C2H2 zinc finger transcription factors family were screened with 2% crystalline cellulose as the sole carbon by measuring protein, endo-beta-1,4-glucanase activity, cellobiohydrolase activity, beta-glucosidase activity and biomass. Accoding to screening results, protein and endo-beta-1,4-glucanase activity were significantly increased by 25%-75% in 7 mutants, while â–³cea-1 and â–³cea-2 were significantly reduced by 65%-80% at protein and endo-beta-1,4-glucanase activity. The further investigations in present studies focused on cea-1 and cea-2 genes. Firstly, the complemented transformants were con-structed, and were found phenotypically indistinguishable from wild type strain. Thus, the deficiency of Acea-1 and â–³cea-2 in cellulase gene expression was indeed due to the disrup-tion of cea-1 and cea-2. And as expected, these two transcriptional factors were found to be localized to the nucleus. Furthermore, the transcriptome profiling analysis of â–³cea-1 and Acea-2 in response to Avicel showed that deletion of these two transcriptional factors caused lots of cellulase genes significantly down-regulated. And other transcriptional fac-tors invovled in lignocellulolytic gene expression were also regulated by cea-1 and cea-2, such as cellulase genes expression activators Clr-1, Clr-2 and carbon catabolite repressor CRE-1. Over-expression of cea-1 and cea-2 would dramatically improve protein produc-tion and cellulase activity.In conclusion, these findings suggest that these two novel transcription factors play an important role in cellulase enzymes expression. These preliminary observations could deepen the understandig of the complex network, and are useful to provide new strategies for constructing high cellulase producing strains.