Abstract
Cohesin is a ring-shaped protein complex that is capable of embracing DNA. Most of the ring circumference is comprised of the anti-parallel intramolecular coiled coils of the Smc1 and Smc3 proteins, which connect globular head and hinge domains. Smc coiled coil arms contain multiple acetylated and ubiquitylated lysines. To investigate the role of these modifications, we substituted lysines for arginines to mimic the unmodified state and uncovered genetic interaction between the Smc arms. Using scanning force microscopy, we show that wild-type Smc arms associate with each other when the complex is not on DNA. Deacetylation of the Smc1/Smc3 dimers promotes arms' dissociation. Smc arginine mutants display loose packing of the Smc arms and, although they dimerize at the hinges, fail to connect the heads and associate with the DNA. Our findings highlight the importance of a "collapsed ring," or "rod," conformation of cohesin for its loading on the chromosomes.
Copyright © 2016 Elsevier Inc. All rights reserved.
MeSH terms
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Acetylation
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Amino Acid Substitution
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Animals
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Arginine / metabolism
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Baculoviridae / genetics
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Baculoviridae / metabolism
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Cell Cycle Proteins / chemistry*
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Chromatids / chemistry
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Chromatids / metabolism
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Chromatids / ultrastructure
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Chromosomal Proteins, Non-Histone / chemistry*
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Chromosomal Proteins, Non-Histone / genetics
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Chromosomal Proteins, Non-Histone / metabolism
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Chromosomes, Fungal / chemistry
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Chromosomes, Fungal / metabolism
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Chromosomes, Fungal / ultrastructure
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Cloning, Molecular
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Cohesins
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DNA, Fungal / chemistry*
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DNA, Fungal / genetics
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DNA, Fungal / metabolism
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Gene Expression
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Gene Expression Regulation, Fungal
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Lysine / metabolism*
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Protein Conformation, alpha-Helical
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Protein Interaction Domains and Motifs
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Protein Processing, Post-Translational*
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / chemistry*
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Sf9 Cells
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Signal Transduction
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Spodoptera
Substances
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Cell Cycle Proteins
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Chromosomal Proteins, Non-Histone
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DNA, Fungal
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Recombinant Proteins
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SMC3 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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structural maintenance of chromosome protein 1
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Arginine
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Lysine