Functional analysis of HO gene in delayed homothallism in Saccharomyces cerevisiae wy2

Saccharomyces cerevisiae wy2 exhibits a novel life cycle, with delayed homo thallism caused by a defective HO gene. In this strain, gradual diploidizat ion occurs during successive subcultures. Three amino acids of wy2 HO were different from those of wild-type (wt) HO, which included a nonsense mutati on (TAG) from Trp-292 and two amino acid changes of His-475 to Leu and Glu- 530 to Lys. The ho gene of heterothallic strain CG379 was also sequenced in this study. Four amino acids of ho were different from those of HO. Among different amino acids in wy2 HO and ho, the alteration of His-475 to Leu wa s common between them. His-475 in HO was previously suggested to be involve d in the DNA binding. We constructed a variety of chimeric HO genes by exch anging the corresponding restriction fragments generated from the wt HO, wy 2 HO and ho genes. These results and the site-directed mutagenesis studies allowed us to draw the following conclusions: (a) Gly-223 is essential for HO activity; (b) mutation of His-475 to Leu significantly reduces the HO ac tivity; (c) amber mutation (TAG) in wy2 HO can be suppressed inefficiently. Copyright (C) 1999 John Wiley & Sons, Ltd.