Analysis and in vivo disruption of the gene coding for calmodulin in Schizosaccharomyces pombe

Abstract

Calmodulin is a low molecular weight calcium-binding protein that modulates many enzyme systems in eukaryotes. We have cloned the gene encoding calmodulin from the fission yeast, Schizosaccharomyces pombe, by using synthetic oligonucleotide probes that correspond to three distinct regions of Tetrahymena calmodulin. A 1.6-kilobase (kb) DNA fragment that hybridized to all of them contains a gene whose deduced product possesses 74% amino acid homology with bovine calmodulin. This gene, which is unique in the S. pombe genome and is named cam1, encodes 149 amino acids excluding the first methionine and is transcribed into mRNA of 1.2-kb length. It has an intron that apparently starts immediately after the initiation codon and is 126 bp long. S. pombe calmodulin exhibits more homology to vertebrate calmodulin than to that of the budding yeast, Saccharomyces cerevisiae. Gene disruption experiments revealed that cam1 gene function is essential for vegetative growth of S. pombe. Spores bearing disrupted cam1 halt growth soon after germination and rarely carry out the first cell division, indicating that calmodulin does not exist in excess in those cells.