Genetic Analysis of the Roles of Yeast ARS Binding Factor I

Abstract

Yeast autonomously replicating sequences (ARSs) were originally identified as segments of DNA that permitted autonomous replication of colinear DNA as plasmids in yeast (Struhl et al., 1979). Direct evidence that ARSs function as origins of replication on plasmids, and more important, that a subset of ARSs function as origins of replication in the chromosome, comes from 2-D gel analysis of replication intermediates (Brewer and Fangman, 1987; Huberman et al., 1987, 1988). ARS1, the ARS with which we have been working for many years, is an active origin within the chromosome. Sequence comparisons between the 20 or so different ARSs that have been identified and the effect of mutations on plasmid stability have revealed two underlying similarities: A high A/T content compared to chromosomal DNA and the presence of a conserved core consensus sequence: 5’ (A/T)TTTA(T/C)(A/G)TTT(A/T(T/C/G) 3’ (see Campbell and Newlon, 1991 for review). Point mutations, small deletions and small substitutions abolish ARS function. Many ARSs contain multiple close matches to this sequence, though spacing and orientation varies. The core consensus sequence is a potential initiator protein binding site. DNase I footprinting patterns of the chromosomal copy of ARS1 showed that the core might be protected from digestion (Lohr and Torchia, 1988).