INTERACTIONS OF THE YEAST CENTROMERE AND PROMOTER FACTOR, CPF1P, WITHTHE CYTOCHROME C(1) UPSTREAM REGION AND FUNCTIONAL IMPLICATIONS ON REGULATED GENE-EXPRESSION

The upstream activation site (UAS) of the cytochrome c(1) gene, CYT1, contains sequences for DNA-binding of several transcription factors, A mong them are the heme-dependent protein, Hap1p, and the multiprotein complex, Hap2/3/4/5, which mediate transcriptional induction under aer obic conditions and after exhaustion of glucose, respectively, The mul tiple interactions of nuclear proteins with the UAS region of CYT1 obs erved in electrophoretic mobility shift experiments are influenced by carbon source and oxygen tension, but are independent of both regulato rs, Hap1p and Hap2/3/4/5. All protein-DNA complexes obtained are solel y due to the association of the centromere and promoter factor 1 (Cpf1 p) with the centromere determining element (CDE I)-like motif at the 5 ' boundary of the UAS(CYT1) This motif overlaps with a consensus seque nce for the binding of the general factor Abf1p, Functional analyses a fter the separate introduction of point mutations into both elements r eveal no role for the latter protein and only a minor role for Cpf1p i n the regulated expression of CYT1/lacZ chimaeric proteins, However, i n cpf1-mutants, induction of CMI reaches higher steady state levels an d adaptation to aerobic conditions occurs faster than in wild-type, Th us, Cpf1p seems to reduce CYT1 promoter activity under partly inducing conditions, e.g. when only one of the activators, Hap1p or the Hap2 c omplex, exerts its function.