The Origin Recognition Complex and Sir4 Protein Recruit Sir1p to Yeast Silent Chromatin through Independent Interactions Requiring a Common Sir1p Domain

Abstract

ABSTRACT Sir1p is one of four SIR (silent information regulator) proteins required for silencing the cryptic mating-type locus HMR a in the budding yeast Saccharomyces cerevisiae . A Sir1p interaction with Orc1p, the largest subunit of the origin recognition complex (ORC), is critical for Sir1p's ability to bind HMR a and function in the formation of silent chromatin. Here we show that a discrete domain within Sir1p, the ORC interaction region (OIR), was necessary and sufficient for a Sir1p-ORC interaction. The OIR contains the originally defined silencer recognition-defective region as well as additional amino acids. In addition, a Sir1p-Sir4p interaction required a larger region of Sir1p that included the OIR. Amino acid substitutions causing defects in either a Sir1p-Orc1p or a Sir1p-Sir4p interaction reduced HMR a silencing and Sir1p binding to HMR a in chromatin. These data support a model in which Sir1p's association with HMR a is mediated by separable Sir1p-ORC and Sir1p-Sir4p interactions requiring a common Sir1p domain, and they indicate that a Sir1p-ORC interaction is restricted to silencers, at least in part, through interactions with Sir4p.