Inhibition of homologous recombination by a cohesin-associated clamp complex recruited to the rDNA recombination enhancer.
Silencing within the yeast ribosomal DNA (rDNA) repeats protects the integrity of this highly repetitive array by inhibiting hyperrecombination and repressing transcription from foreign promoters. Using affinity purification combined with highly sensitive mixture mass spectrometry, we have analyzed the protein interaction network involved in suppressing homologous recombination within the rDNA locus. We show that the Net1 and Sir2 subunits of the RENT (regulator of nucleolar silencing and telophase exit) silencing complex, and Fob1, which recruits RENT to the nontranscribed spacer I (NTS1) region of rDNA, are physically associated with Tof2. In addition to RENT components and Fob1, Tof2 copurified with a two-subunit complex composed of Lrs4 and Csm1. Tof2, Lrs4, and Csm1 are recruited to the NTS1 region by Fob1 and are specifically required for silencing at this rDNA region. Moreover, Lrs4 and Csm1 act synergistically with Sir2 to suppress unequal crossover at the rDNA and are released from the nucleolus during anaphase. Together with previous observations showing that Csm1 physically associates with cohesin, these findings suggest a possible model in which RENT, Tof2, and Lrs4/Csm1 physically clamp rDNA to the cohesin ring, thereby restricting the movement of rDNA sister chromatids relative to each other to inhibit unequal exchange.
Huang
Julie
J
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Brito
Ilana L
IL
Villén
Judit
J
Gygi
Steven P
SP
Amon
Angelika
A
Moazed
Danesh
D
D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000502:physiology; D002466:Cell Nucleolus / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry*; D004275:DNA, Ribosomal / Q000737:chemistry*; D021903:DNA, Ribosomal Spacer; D004268:DNA-Binding Proteins / Q000502:physiology; D020868:Gene Silencing; D008954:Models, Biological; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000737:chemistry* / Q000502:physiology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011995:Recombination, Genetic; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology; D012854:Sister Chromatid Exchange; D000097722:Cohesins
D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004275:DNA, Ribosomal; D021903:DNA, Ribosomal Spacer; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D000097722:Cohesins; C105579:FOB1 protein, S cerevisiae; C474336:LRS4 protein, S cerevisiae