REACTIVATION OF THE ATCASE DOMAIN OF THE URA2 GENE-COMPLEX - A POSITIVE SELECTION METHOD FOR TY INSERTIONS AND CHROMOSOMAL REARRANGEMENTS IN SACCHAROMYCES-CEREVISIAE

Genetic rearrangements such as deletions or duplications of DNA sequen ces are rarely detected in the yeast Saccharomyces cerevisiae. We have developed a screening system using the URA2 gene coding for the bi-fu nctional CPSase-ATCase (carbamyl phosphate synthetase - aspartate tran scarbamylase) to select positively for these kinds of events. Nonsense mutations in the CPSase region cause a complete loss of the ATCase ac tivity because of their strong polar effect. Thirty-seven ATCase(+) re vertants were isolated from a strain containing three nonsense mutatio ns in the proximal CPSase region. Genetic and structural analysis of t he URA2 locus in these strains allowed us to characterize two major cl asses of revertants. In the first, an entire copy of a Ty transposon w as found to be inserted in the CPSase coding domain. This event, which represents a new form of Ty-mediated gene activation was further anal ysed by mapping the Ty integration site in 26 strains. In a second cla ss of revertants, we observed chromosomal rearrangements and, in parti cular, duplication of the ATCase region and its integration in a new c hromosomal environment in which this sequence becomes active.