MONITORING THE GCN4 PROTEIN-MEDIATED RESPONSE IN THE YEAST SACCHAROMYCES-CEREVISIAE

In Saccharomyces cerevisiae the GCN4 gene encodes the transcriptional activator of the ''general control'' system of amino acid bioynthesis, a network of at least 12 different biosynthetic pathways. We characte rized the consequences of the general control response upon the signal ''amino acid starvation'' induced by the histidine analogue 3-aminotr iazole with respect to Gcn4p levels in more detail, Therefore, we esta blished test systems to monitor the time course of different parameter s, including GCN4 mRNA, Gcn4 protein, Gcn4p DNA binding activity, as w ell as Gcn4p transactivation ability. me observed a biphasic response of Gcn4p activity in the cell. At first, translation of GCN4 mRNA is i nduced within 20 min after switch to starvation conditions. However, a n additional increase in GCN4 transcript steady state level was observ ed, leading to an additional second phase of GCN4 expression after 3-4 h of starvation. The DNA binding activity of Gcn4p, as web as the abi lity to activate transcription of target genes, correlate with the amo unt of Gcn4 protein in the cell, suggesting that under the tested cond itions there is no additional regulation of DNA binding or transactiva tion ability of Gcn4p, respectively.