EXAMINATION OF THE FUNCTION OF 2 KELCH PROTEINS GENERATED BY STOP CODON SUPPRESSION

The Drosophila kelch gene produces a single transcript with a UGA stop codon separating two open reading frames (ORF1 and ORF2). From the tr anscript, 76 kDa ORF1 and 160 kDa full-length (ORF1 + ORF2) proteins a re made. The expression of these two proteins is regulated in a tissue -specific manner causing the ratio of full-length to ORF1 protein to v ary in different tissues. The only detected defect for kelch mutants i s female sterility, and kelch protein is localized to the ovarian ring canals, kelch mutant ring canals are disorganized and have partly occ luded lumens, causing a failure to transport cytoplasm. ORF1 and full- length kelch proteins co-sediment with ring canals suggesting that bot h proteins are found in the ring canals. Transgenetic analysis reveals that ORF1 kelch protein is sufficient to rescue ring canal morphology and fertility. In addition, we have mutated the UGA stop codon to a U AA stop codon and to three sense codons that allow constitutive readth rough. Analysis of these mutants reveals that a full-length kelch prot ein can partially compensate for the loss of endogenous kelch, but the residue included at the stop codon is critical for function. Finally, these studies suggest that the mechanism of stop codon suppression of kelch is by tRNA suppression.