HOMEOSTATIC REGULATION OF COPPER UPTAKE IN YEAST VIA DIRECT BINDING OF MAC1 PROTEIN TO UPSTREAM REGULATORY SEQUENCES OF FRE1 AND CTR1

Copper deprivation of Saccharomyces cerevisiae induces transcription o f the FRE1 and CTR1 genes, FRE1 encodes a surface reductase capable of reducing and mobilizing copper chelates outside the cell, and CTR1 en codes a protein mediating copper uptake at the plasma membrane, In thi s paper, the protein encoded by MAC1 is identified as the factor media ting this homeostatic control. A novel dominant allele of MAC1, MAC1(u p2), is mutated in a Cys-rich domain that may function in copper sensi ng (a G to A change of nucleotide 812 resulting in a Cys-271 to Tyr su bstitution), This mutant is functionally similar to the MAC1(up1) alle le in which His-279 in the same domain has been replaced by Gin. Both mutations confer constitutive copper-independent expression of FRE1 an d CTR1. A sequence including the palindrome TTTGCTCA,, TGAGCAAA, appea ring within the 5'-flanking region of the CTR1 promoter, is necessary and sufficient for the copper- and MAC1-dependent CTR1 transcriptional regulation. An identical sequence appears as a direct repeat in the F RE1 promoter, The data indicate that the signal resulting from copper deprivation is transduced via the Cys-rich motif of MAC1 encompassing residues 264-279, MAC1 then binds directly and specifically to the CTR 1 and FRE1 promoter elements, inducing transcription of those target g enes, This model defines the homeostatic mechanism by which yeast regu lates the cell acquisition of copper in response to copper scarcity or excess.