Regulation of expression of GLT1, the gene encoding glutamate synthase in Saccharomyces cerevisiae.

Saccharomyces cerevisiae glutamate synthase (GOGAT) is an oligomeric enzyme composed of three 199-kDa identical subunits encoded by GLT1. In this work, we analyzed GLT1transcriptional regulation. GLT1-lacZ fusions were prepared and GLT1 expression was determined in a GDH1wild-type strain and in a gdh1 mutant derivative grown in the presence of various nitrogen sources. Null mutants impaired inGCN4, GLN3, GAT1/NIL1, orUGA43/DAL80 were transformed with a GLT1-lacZfusion to determine whether the above-mentioned transcriptional factors had a role in GLT1 expression. A collection of increasingly larger 5? deletion derivatives of the GLT1 promoter was constructed to identify DNA sequences that could be involved inGLT1 transcriptional regulation. The effect of the lack ofGCN4, GLN3, or GAT1/NIL1 was also tested in the pertinent 5? deletion derivatives. Our results indicate that (i) GLT1 expression is negatively modulated by glutamate-mediated repression and positively regulated by Gln3p- and Gcn4p-dependent transcriptional activation; (ii) twocis-acting elements, a CGGN15CCG palindrome and an imperfect poly(dA-dT), are present and could play a role inGLT1 transcriptional activation; and (iii) GLT1expression is moderately regulated by GCN4 under amino acid deprivation. Our results suggest that in a wild-type strain grown on ammonium, GOGAT constitutes an ancillary pathway for glutamate biosynthesis.