No abstract

The silent mating-type loci of Saccharomyces cerevisiae, HML and HMR, are flanked by transcriptional silencers that have ARS activity (i.e., they function as replication origins when in plasmids). To test whether these ARS elements are chromosomal origins, we mapped origins near HML (close to the left telomere of chromosome III). Our results indicate that the HML-associated ARS elements either do not function as chromosomal replication origins or do so at a frequency below our detection level, suggesting that replication from a silencer-associated origin in each S phase is not essential for the maintenance of transcriptional repression at HML. Our results also imply that the ability of a DNA fragment to function as an ARS element in a plasmid does not ensure its ability to function as an efficient chromosomal replication origin. Telomere proximity is not responsible for inactivating these ARS elements, because they are not detectably functional as chromosomal origins even in genetically modified strains in which they are far from the telomere.The complex process of chromosomal DNA replication in eukaryotes is poorly understood, partly because origins of DNA replication are not well characterized. In Saccharomyces cerevisiae, autonomously replicating sequences (ARS elements) promote efficient plasmid replication and thus permit plasmids containing them to transform yeast cells at high frequency without integration into the genome. ARS elements can be classified as strong or weak, depending on the mitotic stabilities of plasmids containing them. Strong ARS elements promote high mitotic stabilities, and weak ARS elements promote lower mitotic stabilities. The properties of ARS elements suggest that they may serve as replication origins (7,47 see references 40 and 40a for the locations of these ARS elements on chromosome III)-and a weak ARS in tandemly repeated ribosomal DNA (31) are all active as chromosomal replication origins. In ribosomal DNA, however, only 5 to 30% of available ARS elements are active as origins in each cell cycle (31). Whether all the ARS elements in nonrepeated chromosomal DNA are active as replication origins was, until recently, unknown. The experiments described in this paper reveal that some ARS elements in unique DNA do not detectably function as origins in their normal chromosomal environment.In S. cerevisiae, there are two mating types, a and a. Information specifying mating types is present at three * Corresponding author. t Present address: Department of Zoology, Kutir Mahavidyalaya, Chakkey, Jaunpur-222146, U.P., India.locations on chromosome III, the HML, MAT, and HMR loci (Fig. 1). Under normal conditions, only the information at the MAT locus (either a or a) is expressed. Haploid yeast cells can switch mating type by replacing the MAT information with a copy of either the information present at HML (usually a) or the information present at HMR (usually a) (reviewed by Herskowitz [22]). The HML and HMR loci are located near the left and right telomeres of chromosome III, and infor...

Two dimensional gel electrophoretic techniques were used to locate all functional DNA replication origins in a 22.5 kb stretch of yeast chromosome III. Only one origin was detected, and that origin is located within several hundred bp of an ARS element.

The silent mating-type loci of Saccharomyces cerevisiae, HML and HMR, are flanked by transcriptional silencers that have ARS activity (i.e., they function as replication origins when in plasmids). To test whether these ARS elements are chromosomal origins, we mapped origins near HML (close to the left telomere of chromosome III). Our results indicate that the HML-associated ARS elements either do not function as chromosomal replication origins or do so at a frequency below our detection level, suggesting that replication from a silencer-associated origin in each S phase is not essential for the maintenance of transcriptional repression at HML. Our results also imply that the ability of a DNA fragment to function as an ARS element in a plasmid does not ensure its ability to function as an efficient chromosomal replication origin. Telomere proximity is not responsible for inactivating these ARS elements, because they are not detectably functional as chromosomal origins even in genetically modified strains in which they are far from the telomere.