Cloning of genes expressed early during cellulase induction in Hypocrea jecorina by a rapid subtraction hybridization approach.
The cellulase system of the filamentous fungus Hypocrea jecorina (Trichoderma reesei) is encoded by several cellobiohydrolase, endoglucanase and beta-glucosidase genes, which are co-ordinately expressed upon induction by cellulose or the disaccharide sophorose. To identify genes, which are specifically expressed under these inducing conditions and possibly related to the induction process, we applied rapid subtraction hybridization (RaSH) to sophorose induced mRNAs from the wild-type strain H. jecorina QM9414 and a mutant strain H. jecorina QM9978, which is defective in the induction of cellulase gene expression. From a total of 224 clones, 22 gene fragments representing 20 different genes were analyzed. These included one gene encoding a PAS-domain protein with similarity to the Neurospora clock modulator VIVID; one gene similar to Podospora anserina ami1 involved in nuclear migration and the genes encoding translation elongation factor 1alpha, the transcriptional activator Hap5, and myo-inositol-1-phosphate synthase; in addition, several genes were detected, whose function is unknown. Some of them did not even have potential homologues in the Neurospora or Fusarium genome databases. The differential regulation of expression of those 20 genes by sophorose in wild-type and mutant was verified by Northern blotting. Their consistent response to additional inducing conditions (cellulose) confirms their interconnection with cellulase formation.
Division of Gene Technology and Applied Biochemistry, Institute for Chemical Engineering, Vienna University of Technology, Getreidemarkt 9/1665, A-1060 Wien, Austria. mschmoll@mail.zserv.tuwien.ac.at
Monika
M
Schmoll
Susanne
S
Zeilinger
Robert L
RL
Mach
Christian P
CP
Kubicek
D015152:Blotting, Northern; D023081:CCAAT-Binding Factor / Q000235:genetics; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D005936:Glucans / Q000378:metabolism; D035901:Hypocrea / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D007296:Myo-Inositol-1-Phosphate Synthase / Q000235:genetics; D020648:Peptide Elongation Factor 1 / Q000235:genetics; D012331:RNA, Fungal / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D017422:Sequence Analysis, DNA; D017385:Sequence Homology
D023081:CCAAT-Binding Factor; D004271:DNA, Fungal; D005656:Fungal Proteins; D005936:Glucans; D020648:Peptide Elongation Factor 1; D012331:RNA, Fungal; D012333:RNA, Messenger; C423283:VVD protein, Neurospora crassa; C111072:aab-1 protein, Neurospora; D007296:Myo-Inositol-1-Phosphate Synthase; C003328:sophorose