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      The Condensin Complex Governs Chromosome Condensation and Mitotic Transmission of Rdna

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          Abstract

          We have characterized five genes encoding condensin components in Saccharomyces cerevisiae. All genes are essential for cell viability and encode proteins that form a complex in vivo. We characterized new mutant alleles of the genes encoding the core subunits of this complex, smc2-8 and smc4-1. Both SMC2 and SMC4 are essential for chromosome transmission in anaphase. Mutations in these genes cause defects in establishing condensation of unique (chromosome VIII arm) and repetitive (rDNA) regions of the genome but do not impair sister chromatid cohesion. In vivo localization of Smc4p fused to green fluorescent protein showed that, unexpectedly, in S. cerevisiae the condensin complex concentrates in the rDNA region at the G2/M phase of the cell cycle. rDNA segregation in mitosis is delayed and/or stalled in smc2 and smc4 mutants, compared with separation of pericentromeric and distal arm regions. Mitotic transmission of chromosome III carrying the rDNA translocation is impaired in smc2 and smc4 mutants. Thus, the condensin complex in S. cerevisiae has a specialized function in mitotic segregation of the rDNA locus. Chromatin immunoprecipitation (ChIP) analysis revealed that condensin is physically associated with rDNA in vivo. Thus, the rDNA array is the first identified set of DNA sequences specifically bound by condensin in vivo. The biological role of higher-order chromosome structure in S. cerevisiae is discussed.

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          Comprehensive identification of cell cycle-regulated genes of the yeast Saccharomyces cerevisiae by microarray hybridization.

          We sought to create a comprehensive catalog of yeast genes whose transcript levels vary periodically within the cell cycle. To this end, we used DNA microarrays and samples from yeast cultures synchronized by three independent methods: alpha factor arrest, elutriation, and arrest of a cdc15 temperature-sensitive mutant. Using periodicity and correlation algorithms, we identified 800 genes that meet an objective minimum criterion for cell cycle regulation. In separate experiments, designed to examine the effects of inducing either the G1 cyclin Cln3p or the B-type cyclin Clb2p, we found that the mRNA levels of more than half of these 800 genes respond to one or both of these cyclins. Furthermore, we analyzed our set of cell cycle-regulated genes for known and new promoter elements and show that several known elements (or variations thereof) contain information predictive of cell cycle regulation. A full description and complete data sets are available at http://cellcycle-www.stanford.edu
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            Cohesins: chromosomal proteins that prevent premature separation of sister chromatids.

            Cohesion between sister chromatids opposes the splitting force exerted by microtubules, and loss of this cohesion is responsible for the subsequent separation of sister chromatids during anaphase. We describe three chromosmal proteins that prevent premature separation of sister chromatids in yeast. Two, Smc1p and Smc3p, are members of the SMC family, which are putative ATPases with coiled-coil domains. A third protein, which we call Scc1p, binds to chromosomes during S phase, dissociates from them at the metaphase-to-anaphase transition, and is degraded by the anaphase promoting complex. Association of Scc1p with chromatin depends on Smc1p. Proteins homologous to Scc1p exist in a variety of eukaryotic organisms including humans. A common cohesion apparatus might be used by all eukaryotic cells during both mitosis and meiosis.
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              New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites.

              We describe the production of new alleles of the LEU2, URA3 and TRP1 genes of Saccharomyces cerevisiae by in vitro mutagenesis. Each new allele, which lacks restriction enzyme recognition sequences found in the pUC19 multicloning site, was used to construct a unique series of yeast-Escherichia coli shuttle vectors derived from the plasmid pUC19. For each gene a 2 mu vector (YEplac), an ARS1 CEN4 vector (YCplac) and an integrative vector (YIplac) was constructed. The features of these vectors include (i) small size; (ii) unique recognition site for each restriction enzyme found in the pUC19 multicloning site; (iii) screening for plasmids containing inserts by color assay; (iv) high plasmid yield; (v) efficient transformation of S. cerevisiae. These vectors should allow greater flexibility with regard to DNA restriction fragment manipulation and subcloning.
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                Author and article information

                Contributors
                Journal
                J Cell Biol
                The Journal of Cell Biology
                The Rockefeller University Press
                0021-9525
                1540-8140
                15 May 2000
                : 149
                : 4
                : 811-824
                Affiliations
                [a ]Unit of Chromosome Structure and Function, National Institutes of Health, National Institute of Child Health and Human Development, Laboratory of Molecular Embryology, Bethesda, Maryland 20892-5430
                Article
                9905069
                10.1083/jcb.149.4.811
                2174567
                10811823
                687749b5-1383-4714-9a06-9abf1b02aba3
                © 2000 The Rockefeller University Press
                History
                : 18 May 1999
                : 22 March 2000
                : 29 March 2000
                Categories
                Original Article

                Cell biology
                chromosome condensation,condensin,chromosome segregation,smc,chromatin
                Cell biology
                chromosome condensation, condensin, chromosome segregation, smc, chromatin

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