ROLE OF THE SACCHAROMYCES-CEREVISIAE GENERAL REGULATORY FACTOR CP1 INMETHIONINE BIOSYNTHETIC GENE-TRANSCRIPTION

Saccharomyces cerevisiae general regulatory factor CP1 (encoded by the gene CEP1) is required for optimal chromosome segregation and methion ine prototrophy. MET16-CYC1-lacZ reporter constructs were used to show that MET16 5'-flanking DNA contains a CP1-dependent upstream activati on sequence (UAS). Activity of the UAS required an intact CP1-binding site, and the effects of cis-acting mutations on CP1 binding and UAS a ctivity correlated. In most respects, MET16-CYC1-lacZ reporter gene ex pression mirrored that of chromosomal MET16; however, the endogenous g ene was found to be activated in response to amino acid starvation (ge neral control). The latter mechanism was both GCN4 and CP1 dependent. MET25 was also found to be activated by GCN4, albeit weakly. More impo rtantly, MET25 transcription was strongly CP1 dependent in gcn4 backgr ounds. The modulation of MET gene expression by GCN4 can explain discr epancies in the literature regarding CPI dependence of MET gene transc ription. Lastly, micrococcal nuclease digestion and indirect end label ing were used to analyze the chromatin structure of the MET16 locus in wild-type and cep1 cells. The results indicated that CPI plays no maj or role in configuring chromatin structure in this region, although lo calized CP1-specific differences in nuclease sensitivity were detected .