CLONING AND CHARACTERIZATION OF KNR4, A YEAST GENE INVOLVED IN (1,3)-BETA-GLUCAN SYNTHESIS

k9 killer toxin from Hansenula mrakii was used to select a number of r esistant mutants from Saccharomyces cerevisiae. Preliminary biochemica l and genetic studies showed that some of them acquired structural def ects in the cell wall. One of these mutants, the knr4-1 mutant, displa ys a number of cell wall defects, including osmotic sensitivity; sensi tivity to cercosporamide, a known antifungal agent; and resistance to Zymolyase, a (1,3)-beta-glucanase. We report here the isolation and an alysis of the KNR4 gene. DNA sequence analysis revealed an uninterrupt ed open reading frame which contains five potential start codons. The longest coding template encodes a protein of 505 amino acids with a ca lculated molecular mass of 57,044 Da. A data base search revealed 100% identity with a nuclear protein, SMI1p. Disruption of the KNR4 locus does not result in cell death; however, it leads to reduced levels of both (1,3)-beta-glucan synthase activity and (1,3)-beta-glucan content in the cell wall. The gene was mapped to the right arm of chromosome VII.