The origin recognition complex and Sir4 protein recruit Sir1p to yeast silent chromatin through independent interactions requiring a common Sir1p domain.
Sir1p is one of four SIR (silent information regulator) proteins required for silencing the cryptic mating-type locus HMRa 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 HMRa 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 HMRa silencing and Sir1p binding to HMRa in chromatin. These data support a model in which Sir1p's association with HMRa 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.
Bose
Melissa E
ME
Department of Biomolecular Chemistry, 587 MSC, University of Wisconsin Medical School, 1300 University Avenue, Madison, WI 43706-1532, USA.
McConnell
Kristopher H
KH
Gardner-Aukema
Kelly A
KA
Müller
Ulrika
U
Weinreich
Michael
M
Keck
James L
JL
Fox
Catherine A
CA
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics; D020868:Gene Silencing; D005800:Genes, Fungal; D049770:Genes, Mating Type, Fungal; D008954:Models, Biological; D008969:Molecular Sequence Data; D051738:Origin Recognition Complex; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics
D002843:Chromatin; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D051738:Origin Recognition Complex; C084121:SIR1 protein, S cerevisiae; C061573:SIR4 protein, S cerevisiae; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae