Molecular cloning, identification and transcriptional analysis of genes involved in acetate utilization in Neurospora crassa.

Four Neurospora crassa genomic clones have been selected as hybridizing much more strongly to labelled mRNA isolated from acetate-grown mycelium than to mRNA from sucrose-grown mycelium. Hybridization of restriction fragments with acetate-specific mRNA or cDNA has been used to delimit the transcribed region(s) of each clone. The transcription of all four clones is strongly induced by transfer of growing mycelium from sucrose to acetate as sole carbon source. In wild-type mycelium, mRNAs corresponding to the four clones reach maximum levels after four hours of induction. They accumulate more rapidly and reach higher levels in an acetate non-utilizing mutant, acu-7, which has been found to overproduce enzymes of the glyoxylate cycle and to have a partial block in the TCA cycle. Molecular transformation of a Neurospora acu-5 mutant and of an Aspergillus nidulans acuE mutant by DNA of clone 2 and clone 1, respectively, strongly suggests that clone 2 codes for acetyl-coenzyme A synthetase and that clone 1 codes for malate synthase. The transcribed segments of clones 1 and 2 each hybridize to corresponding clones from Aspergillus nidulans (R. A. Sandeman and M. J. Hynes, personal communication).