Dominance of Metallothionein in Metal Ion Buffering in Yeast Capable of Synthesis of (γEC)_nG Isopeptides

The relationship of yeast metallothionein (MT) and (γEC)_nG isopeptides (phytochelatins) in metal ion buffering was assessed. The effect of constitutive expression of yeast metallothionein (MT) genes on accumulation of metal-(γEC)_nG isopeptide (phytochelatin) complexes was analyzed in Candida glabrata and Schizosaccharomyces pombe cultures incubated in the presence of cadmium salts. Constitutive expression of the Saccharomyces cerevisiae MT (CUP1) gene inhibited the accumulation of metal-phytochelatin complexes in both C. glabrata and S. pombe. Intracellular Cd(II) sequestration occurred by formation of CdMT complexes. Phytochelatin (γEC)_nG complexes appear to function in metal buffering in cells when MT genes are not present or expressed. A third condition in which metal-(γEC)_nG complexes are observed is when constitutively expressed MT does not accumulate. We observed that C. glabrata lacking the AMT1 gene necessary for copper induction of the MT genes expressed MTII constitutively, but this expression does not lead to CdMTII accumulation. Only Cd-(γEC)_nG complexes accumulate. Likewise, metal exposed cultures of S. cerevisiae (cup1) transformed with C. glabrata MTII under the constitutive ADH1 promoter resulted in constitutive expression of MTII and accumulation of CuMTII complexes but no CdMTII complexes. The inability of constitutively expressed C. glabrata MTII to buffer Cd(II) ions may arise in part from an inherent kinetic lability of CdMTII complexes. Incubation of ZnMTII with a metallochromic chelator, 4-(2-pyridylazo)resorcinol resulted in greater Zn(II) loss than Zn(II) complexes with CUP1 MT or C. glabrata MTI. C. glabrata MTII appears to be the first MT described which forms an unstable Cd(II) complex.