Molecular cloning, DNA structure, and RNA analysis of the arginase gene in Saccharomyces cerevisiae. A study of cis-dominant regulatory mutations.

Abstract

The Saccharomyces cerevisiae gene cargA + or CAR1 , encoding arginase has been cloned by recovering function in transformed yeast cells. It was used to analyse RNA and chromosomal DNA from six strains bearing cis-dominant regulatory mutations leading to constitutive arginase synthesis. The DNA from the four cargA + O- strains in which constitutive arginase synthesis was independent of the mating-type functions showed no detectable differences with the wild- typye . The cargA + O- mutations were, therefore, small alterations, possibly single base substitutions. On the other hand, the cargA + Oh-1 and cargA + Oh-2 mutations, leading to a constitutive and mating-type dependent arginase synthesis, were identified as insertions. Their size and restriction pattern strongly suggested that they were induced by the Ty1 yeast transposable element. This was confirmed by cloning and analysis of the cargA + Oh-1 mutant gene. The concentration of arginase RNA was significantly increased in the mutants, indicating that the regulation of arginase synthesis was exerted, at least in part, at the level of RNA synthesis or stability. In the cargA + Oh-2 strain the Ty1 element was located at a distance of approximately 600 base pairs from the insertion present in the cargA + Oh-1 strain. This result suggests either a surprisingly large arginase regulatory region or an indirect influence of the Ty1 element on gene expression over long distances.