The Sir4 C-terminal coiled coil is required for telomeric and mating type silencing in Saccharomyces cerevisiae.
Saccharomyces cerevisiae Sir4p plays important roles in silent chromatin at telomeric and silent mating type loci. The C terminus of Sir4p (Sir4CT) is critical for its functions in vivo because over-expression or deletion of Sir4CT fragments disrupts normal telomeric structure and abolishes the telomere position effect. The 2.5A resolution X-ray crystal structure of an Sir4CT fragment (Sir4p 1217-1358) reveals a 72 residue homodimeric, parallel coiled coil, burying an extensive 3600A(2) of surface area. The crystal structure is consistent with results of protein cross-linking and analytical ultracentrifugation results demonstrating that Sir4CT exists as a dimer in solution. Disruption of the coiled coil in vivo by point mutagenesis results in total derepression of telomeric and HML silent mating marker genes, suggesting that coiled coil dimerization is essential for Sir4p-mediated silencing. In addition to the coiled coil dimerization interface (Sir4CC interface), a crystallographic interface between pairs of coiled coils is significantly hydrophobic and buries 1228A(2) of surface area (interface II). Remarkably, interface II mutants are deficient in telomeric silencing but not in mating type silencing in vivo. However, point mutants of interface II do not affect the oligomerization state of Sir4CT in solution. These results are consistent with the hypothesis that interface II mimics a protein interface between Sir4p and one of its protein partners that is essential for telomeric silencing but not mating type silencing.
Murphy
Guinevere A
GA
Department of Chemistry and Biochemistry, University of Colorado at Boulder, Boulder, CO 80309-0215, USA.
Spedale
Erik J
EJ
Powell
Sean T
ST
Pillus
Lorraine
L
Schultz
Steve C
SC
Chen
Lin
L
D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D020868:Gene Silencing; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D016615:Telomere / Q000378:metabolism*
C061573:SIR4 protein, S cerevisiae; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae