Positive regulation in a eukaryote, a study of the uaY gene of Aspergillus nidulans

Abstract

In this publication we report the identification of a protein likely to be coded by uaY, a regulatory gene in the ascomycete Aspergillus nidulans. uaY is a positive control gene necessary for the expression of at least eight unlinked structural genes involved in purine uptake and degradation (Scazzocchio and Gorton 1977). The physiological effector of the uaY system is uric acid, while some of its thioanalogs serve as gratuitous inducers. Effector binding proteins were detected by binding to 2-thiouric acid after phosphocellulose column chromatography, or as uric acid binding fractions after DNA-cellulose column chromatography. Two binding peaks are present in mycelial extracts purified by either method. These are missing in a putative small deletion of the uaY gene. A “leaky” mutation, uaY 109 described in detail elsewhere (Scazzocchio et al. 1980) shows only one peak. The wild type peaks are eluted at 55 mM NaCl and at 720 mM NaCl while the peak present in uaY109 is eluted at 120 mM NaCl. This implies that at least one peak represents a protein coded by the uaY gene. The major peak was analysed by equilibrium dialysis experiments. These establish a K_diss.≃2×10^-7 and a minimum number of binding sites of 3×10^-14 moles/mg of soluble protein in a crude extract derived from protoplast lysis. An extract from a strain carrying the uaY207 deletion, purified blind, lacks any binding activity in the equilibrium dialysis cell.