Fig. 1

S. cerevisiae gcd7-201 gcn2∆ mutant strain confers Slg⁺ and Gcd⁺ phenotype when transformed with high copy (hc) pEG(KG)/EMC4 plasmid. GCD7 gcn2∆, gcd7-201 gcn2∆ harboring hc/EMC4, or empty hc vector pEG(KG) were streaked in parallel on SC medium lacking uracil, but either containing (a) raffinose or (b) galactose. Uracil-based plasmid hc/EMC4 was evicted on SC medium containing (c) FOA, and were further streaked on (d) SC-Ura medium. (e) Spotting of GCD7 gcn2∆, gcd7-201 gcn2∆ harboring hc/EMC4, or empty hc vector pEG(KG) was done on SC medium containing 2% galactose and lacking uracil or SC medium containing 2% galactose, 30 mM 3-AT, and lacking uracil. Plates were incubated at 30 °C for 2 days. (f) Western analysis of GST-Emc4p expression in gcd7-201 gcn2∆ strain with anti-GST antibody. The whole cell protein extracts (20 μg) were prepared from uninduced and 2% galactose-induced cultures of the strain harboring hc/EMC4. Samples were then separated on 10% SDS gel followed by Western blotting using anti-GST for GST-Emc4 and anti-Gcd6 antibodies (Loading control). UI, uninduced; I, induced