The quinic acid gene cluster in Neurospora: Sequence comparison and gene expression

The quinic acid (qa) cluster of Neurospora crassa is subject to two levels of control. The primary mechanism is dependent on the presence of the inducer, quinic acid: in the absence of inducer, the genes are expressed at low basal levels. The cluster is also subject to catabolite repression---the genes are repressed in the presence of a preferred carbon source, such as dextrose, even in the presence of inducer. To focus on this second level of control, I utilized a constitutive mutant of the qa gene cluster which contains a deletion of the qa-1S repressor gene. This deletion removes the primary control mechanism, ensuring that any observed effect is due to catabolite repression. Northern blot analysis indicates that transcription of the qa-y gene, which encodes a quinate permease, is repressed by the presence of dextrose even in the absence of the qa-1S repressor, demonstrating that catabolite repression of the qa cluster may be occurring by inducer exclusion. Transcription of the remainder of the qa genes does not appear to be repressed to a significant degree in the constitutive mutant by the presence of dextrose. Additionally, I used 2-dimensional protein gel electrophoresis to examine the expression of the qa proteins in samples grown under different growth conditions. Preliminary results show changes in protein expression in repressor mutants when compared to wild-type.; Gene regulation mechanisms can be revealed by comparing homologous sequences in related species. I sequenced the permease and repressor genes, along with the associated regulatory regions in the homothallic Neurospora species N. africana and N. terricola. The coding regions of both species share a high degree of homology to N. crassa: the permease genes are 93% identical and the repressor 91--92% identical. The noncoding region upstream of the repressor is less conserved, with N. africana showing 41% identity and N. terricola 59% identity to N. crassa. In contrast, the region upstream of the permease gene is highly conserved, with both species 83% identical to N. crassa. This highly conserved region could contain binding sites needed to affect catabolite repression of the permease gene.