SMC (structural maintenance of chromosomes) structural protein family and their role in chromatin reorganization

Structural chromatin proteins of the SMC (Structural Maintenance of Chromosomes) family play an important role in structural DNA reorganization in pro- and eukaryotes. Eukaryotic SMC proteins are the core components of the cohesin and condensin complexes. The cohesin complex is responsible for sister chromatid and homolog cohesion in mitosis and meiosis. The condensin complex uses ATP energy to induce positive coiled-coils in DNA, which results in compaction of the latter and formation of mitotic chromosome scaffold. In addition, the SMC proteins constitute recombination and recombination repair complexes. In hermaphrodites of nematode Caenorhabditis elegans, the SMC protein-containing complex controls dosage compensation and inactivation of the X chromosome genes.     

SMC complexes and topoisomerase II work together so that sister chromatids can work apart

The pairing of sister chromatids in interphase facilitates error-free homologous recombination (HR). Sister chromatids are held together by cohesin, one of three Structural Maintenance of Chromosomes (SMC) complexes. In mitosis, chromosome condensation is controlled by another SMC complex, condensin, and the type II topoisomerase (Top2). In prophase, cohesin is stripped from chromosome arms, but remains at centromeres until anaphase, whereupon it is removed via proteolytic cleavage. The third SMC complex, Smc5/6, is generally described as a regulator of HR-mediated DNA repair. However, cohesin and condensin are also required for DNA repair, and HR genes are not essential for cell viability, but the SMC complexes are. Smc5/6 null mutants die in mitosis, and in fission yeast, Smc5/6 hypomorphs show lethal mitoses following genotoxic stress, or when combined with a Top2 mutant, top2-191. We found these mitotic defects are due to retention of cohesin on chromosome arms. We also show that Top2 functions in the cohesin cycle, and accumulating data suggests this is not related to its decatenation activity. Thus the SMC complexes and Top2 functionally interact, and any DNA repair function ascribed to Smc5/6 is likely a reflection of a more fundamental role in the regulation of chromosome structure.     

Condensin and cohesin display different arm conformations with characteristic hinge angles

Structural maintenance of chromosomes (SMC) proteins play central roles in higher-order chromosome dynamics from bacteria to humans. In eukaryotes, two different SMC protein complexes, condensin and cohesin, regulate chromosome condensation and sister chromatid cohesion, respectively. Each of the complexes consists of a heterodimeric pair of SMC subunits and two or three non-SMC subunits. Previous studies have shown that a bacterial SMC homodimer has a symmetrical structure in which two long coiled-coil arms are connected by a flexible hinge. A catalytic domain with DNA- and ATP-binding activities is located at the distal end of each arm. We report here the visualization of vertebrate condensin and cohesin by electron microscopy. Both complexes display the two-armed structure characteristic of SMC proteins, but their conformations are remarkably different. The hinge of condensin is closed and the coiled-coil arms are placed close together. In contrast, the hinge of cohesin is wide open and the coiled-coils are spread apart from each other. The non-SMC subunits of both condensin and cohesin form a globular complex bound to the catalytic domains of the SMC heterodimers. We propose that the "closed" conformation of condensin and the "open" conformation of cohesin are important structural properties that contribute to their specialized biochemical and physiological functions.     

Condensin but not cohesin SMC heterodimer induces DNA reannealing through protein-protein assembly

Condensin and cohesin are chromosomal protein complexes required for chromosome condensation and sister chromatid cohesion, respectively. They commonly contain the SMC (structural maintenance of chromosomes) subunits consisting of a long coiled-coil with the terminal globular domains and the central hinge. Condensin and cohesin holo-complexes contain three and two non-SMC subunits, respectively. In this study, DNA interaction with cohesin and condensin complexes purified from fission yeast was investigated. The DNA reannealing activity is strong for condensin SMC heterodimer but weak for holo-condensin, whereas no annealing activity is found for cohesin heterodimer SMC and Rad21-bound heterotrimer complexes. One set of globular domains of the same condensin SMC is essential for the DNA reannealing activity. In addition, the coiled-coil and hinge region of another SMC are needed. Atomic force microscopy discloses the molecular events of DNA reannealing. SMC assembly that occurs on reannealing DNA seems to be a necessary intermediary step. SMC is eliminated from the completed double-stranded DNA. The ability of heterodimeric SMC to reanneal DNA may be regulated in vivo possibly through the non-SMC heterotrimeric complex.     

Combing Chromosomal DNA Mediated by the SMC Complex: Structure and Mechanisms

Genome maintenance requires various nucleoid‐associated factors in prokaryotes. Among them, the SMC (Structural Maintenance of Chromosomes) protein has been thought to play a static role in the organization and segregation of the chromosome during cell division. However, recent studies have shown that the bacterial SMC is required to align left and right arms of the emerging chromosome and that the protein dynamically travels from origin to Ter region. A rod form of the SMC complex mediates DNA bridging and has been recognized as a machinery responsible for DNA loop extrusion, like eukaryotic condensin or cohesin complexes, which act as chromosome organizers. Attention is now turning to how the prototype of the complex is loaded on the entry site and translocated on chromosomal DNA, explaining its overall conformational changes at atomic levels. Here, we review and highlight recent findings concerning the prokaryotic SMC complex and discuss possible mechanisms from the viewpoint of protein architecture.     

The Smc5-Smc6 complex is required to remove chromosome junctions in meiosis

Meiosis, a specialized cell division with a single cycle of DNA replication round and two consecutive rounds of nuclear segregation, allows for the exchange of genetic material between parental chromosomes and the formation of haploid gametes. The structural maintenance of chromosome (SMC) proteins aid manipulation of chromosome structures inside cells. Eukaryotic SMC complexes include cohesin, condensin and the Smc5-Smc6 complex. Meiotic roles have been discovered for cohesin and condensin. However, although Smc5-Smc6 is known to be required for successful meiotic divisions, the meiotic functions of the complex are not well understood. Here we show that the Smc5-Smc6 complex localizes to specific chromosome regions during meiotic prophase I. We report that meiotic cells lacking Smc5-Smc6 undergo catastrophic meiotic divisions as a consequence of unresolved linkages between chromosomes. Surprisingly, meiotic segregation defects are not rescued by abrogation of Spo11-induced meiotic recombination, indicating that at least some chromosome linkages in smc5-smc6 mutants originate from other cellular processes. These results demonstrate that, as in mitosis, Smc5-Smc6 is required to ensure proper chromosome segregation during meiosis by preventing aberrant recombination intermediates between homologous chromosomes.     

Condensin architecture and interaction with DNA: regulatory non-SMC subunits bind to the head of SMC heterodimer

Condensin and cohesin are two protein complexes that act as the central mediators of chromosome condensation and sister chromatid cohesion, respectively. The basic underlying mechanism of action of these complexes remained enigmatic. Direct visualization of condensin and cohesin was expected to provide hints to their mechanisms. They are composed of heterodimers of distinct structural maintenance of chromosome (SMC) proteins and other non-SMC subunits. Here, we report the first observation of the architecture of condensin and its interaction with DNA by atomic force microscopy (AFM). The purified condensin SMC heterodimer shows a head-tail structure with a single head composed of globular domains and a tail with the coiled-coil region. Unexpectedly, the condensin non-SMC trimers associate with the head of SMC heterodimers, producing a larger head with the tail. The heteropentamer is bound to DNA in a distributive fashion, whereas condensin SMC heterodimers interact with DNA as aggregates within a large DNA-protein assembly. Thus, non-SMC trimers may regulate the ATPase activity of condensin by directly interacting with the globular domains of SMC heterodimer and alter the mode of DNA interaction. A model for the action of heteropentamer is presented.     

Smc5/6 Coordinates Formation and Resolution of Joint Molecules with Chromosome Morphology to Ensure Meiotic Divisions

During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and the regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of a subset of joint-molecule intermediates. In this regard, Smc5/6 functions independently of the major crossover pathway defined by the MutLγ complex. Furthermore, we show that Smc5/6 is required for stable chromosomal localization of the XPF-family endonuclease, Mus81-Mms4^Eme1. Our data suggest that the Smc5/6 complex is required for specific recombination and chromosomal processes throughout meiosis and that in its absence, attempts at cell division with unresolved joint molecules and residual cohesin lead to severe recombination-induced meiotic catastrophe.     

In vivo dissection of the chromosome condensation machinery: reversibility of condensation distinguishes contributions of condensin and cohesin

The machinery mediating chromosome condensation is poorly understood. To begin to dissect the in vivo function(s) of individual components, we monitored mitotic chromosome structure in mutants of condensin, cohesin, histone H3, and topoisomerase II (topo II). In budding yeast, both condensation establishment and maintenance require all of the condensin subunits, but not topo II activity or phospho-histone H3. Structural maintenance of chromosome (SMC) protein 2, as well as each of the three non-SMC proteins (Ycg1p, Ycs4p, and Brn1p), was required for chromatin binding of the condensin complex in vivo. Using reversible condensin alleles, we show that chromosome condensation does not involve an irreversible modification of condensin or chromosomes. Finally, we provide the first evidence of a mechanistic link between condensin and cohesin function. A model discussing the functional interplay between cohesin and condensin is presented.     

Structure and DNA binding activity of the mouse condensin hinge domain highlight common and diverse features of SMC proteins

Structural Maintenance of Chromosomes (SMC) proteins are vital for a wide range of processes including chromosome structure and dynamics, gene regulation and DNA repair. Eukaryotes have three SMC complexes, consisting of heterodimeric pairs of six different SMC proteins along with several specific regulatory subunits. In addition to their other functions, all three SMC complexes play distinct roles in DNA repair. Cohesin (SMC1–SMC3) is involved in DNA double-strand break repair, condensin (SMC2–SMC4) participates in single-strand break (SSB) repair, and the SMC5–SMC6 complex functions in various DNA repair pathways. SMC proteins consist of N- and C-terminal domains that fold back onto each other to create an ATPase ‘head’ domain, connected to a central ‘hinge’ domain via long coiled-coils. The hinge domain mediates dimerization of SMC proteins and binds DNA, but it is not clear to what purpose this activity serves. We studied the structure and function of the condensin hinge domain from mouse. While the SMC hinge domain structure is largely conserved from prokaryotes to eukaryotes, its function seems to have diversified throughout the course of evolution. The condensin hinge domain preferentially binds single-stranded DNA. We propose that this activity plays a role in the SSB repair function of the condensin complex.     

Functional interplay between cohesin and Smc5/6 complexes

Chromosomes are subjected to massive reengineering as they are replicated, transcribed, repaired, condensed, and segregated into daughter cells. Among the engineers are three large protein complexes collectively known as the structural maintenance of chromosome (SMC) complexes: cohesin, condensin, and Smc5/6. As their names suggest, cohesin controls sister chromatid cohesion, condensin controls chromosome condensation, and while precise functions for Smc5/6 have remained somewhat elusive, most reports have focused on the control of recombinational DNA repair. Here, we focus on cohesin and Smc5/6 function. It is becoming increasingly clear that the functional repertoires of these complexes are greater than sister chromatid cohesion and recombination. These SMC complexes are emerging as interrelated and cooperating factors that control chromosome dynamics throughout interphase. However, they also release their embrace of sister chromatids to enable their segregation at anaphase, resetting the dynamic cycle of SMC-chromosome interactions.     

Identification of a novel non-structural maintenance of chromosomes (SMC) component of the SMC5-SMC6 complex involved in DNA repair

Structural maintenance of chromosomes (SMC) proteins play central roles in chromosome organization and dynamics. They have been classified into six subtypes, termed SMC1 to SMC6, and function as heterodimer components of large protein complexes that also include several non-SMC proteins. The SMC2-SMC4 and SMC1-SMC3 complexes are also known as condensin and cohesin, respectively, but the recently identified SMC5 and SMC6 complex is less well characterized. Here, we report that NSE1 from Saccharomyces cerevisiae encodes a novel non-SMC component of the SMC5(Yol034wp)-SMC6(Rhc18p) complex corresponding to the 2-3-MDa molecular mass. Nse1p is essential for cell proliferation and localizes primarily in the nucleus. nse1 mutants are highly sensitive to DNA-damaging treatments and exhibit abnormal cellular morphologies, suggesting aberrant mitosis as a terminal morphological phenotype. These results are consistent with the reported features of the Schizosaccharomyces pombe SMC6 gene, rad18, which is thought to be involved in recombinational DNA repair. We conclude that Nse1p and the SMC5-SMC6 heterodimer together form a high molecular mass complex that is conserved in eukaryotes and required for both DNA repair and proliferation.     

The role of SMC proteins in the responses to DNA damage

The SMC proteins form the cores of three protein complexes in eukaryotes, cohesin, condensin and the Smc5-6 complex. Cohesin holds sister chromatids together after DNA replication and is involved in both the repair of double-strand breaks by homologous recombination and the intra-S-phase checkpoint. Condensin assists in the condensation of chromosomes at mitosis and also has a role in checkpoint control pathways. The Smc5-6 complex is involved in a variety of DNA repair and damage response pathways by as yet unknown mechanisms, but is also associated with repair by homologous recombination.     

How DNA loop extrusion mediated by cohesin enables V(D)J recombination

‘Structural maintenance of chromosomes’ (SMC) complexes are required for the folding of genomic DNA into loops. Theoretical considerations and single-molecule experiments performed with the SMC complexes cohesin and condensin indicate that DNA folding occurs via loop extrusion. Recent work indicates that this process is essential for the assembly of antigen receptor genes by V(D)J recombination in developing B and T cells of the vertebrate immune system. Here, I review how recent studies of the mouse immunoglobulin heavy chain locus Igh have provided evidence for this hypothesis and how the formation of chromatin loops by cohesin and regulation of this process by CTCF and Wapl might ensure that all variable gene segments in this locus (V_H segments) participate in recombination with a re-arranged DJ_H segment, to ensure generation of a maximally diverse repertoire of B-cell receptors and antibodies.     

Three-dimensional topology of the SMC2/SMC4 subcomplex from chicken condensin I revealed by cross-linking and molecular modelling

SMC proteins are essential components of three protein complexes that are important for chromosome structure and function. The cohesin complex holds replicated sister chromatids together, whereas the condensin complex has an essential role in mitotic chromosome architecture. Both are involved in interphase genome organization. SMC-containing complexes are large (more than 650 kDa for condensin) and contain long anti-parallel coiled-coils. They are thus difficult subjects for conventional crystallographic and electron cryomicroscopic studies. Here, we have used amino acid-selective cross-linking and mass spectrometry combined with structure prediction to develop a full-length molecular draft three-dimensional structure of the SMC2/SMC4 dimeric backbone of chicken condensin. We assembled homology-based molecular models of the globular heads and hinges with the lengthy coiled-coils modelled in fragments, using numerous high-confidence cross-links and accounting for potential irregularities. Our experiments reveal that isolated condensin complexes can exist with their coiled-coil segments closely apposed to one another along their lengths and define the relative spatial alignment of the two anti-parallel coils. The centres of the coiled-coils can also approach one another closely in situ in mitotic chromosomes. In addition to revealing structural information, our cross-linking data suggest that both H2A and H4 may have roles in condensin interactions with chromatin.     

At the heart of the chromosome: SMC proteins in action

Structural maintenance of chromosomes (SMC) proteins are ubiquitous in organisms from bacteria to humans, and function as core components of the condensin and cohesin complexes in eukaryotes. SMC proteins adopt a V-shaped structure with two long arms, each of which has an ATP-binding head domain at the distal end. It is important to understand how these uniquely designed protein machines interact with DNA strands and how such interactions are modulated by the ATP-binding and -hydrolysis cycle. An emerging idea is that SMC proteins use a diverse array of intramolecular and intermolecular protein-protein interactions to actively fold, tether and manipulate DNA strands.     

Crystal structure of the MukB hinge domain with coiled‐coil stretches and its functional implications

The structural maintenance of chromosomes (SMC) family proteins are commonly found in the multiprotein complexes involved in chromosome organization, including chromosome condensation and sister chromatid cohesion. These proteins are characterized by forming a V‐shaped homo‐ or heterodimeric structure with two long coiled‐coil arms having two ATPase head domains at the distal ends. The hinge domain, located in the middle of the coiled coil, forms the dimer interface. In addition to being the dimerization module, SMC hinges appear to play other roles, including the gateway function for DNA entry into the cohesin complex. Herein, we report the homodimeric structure of the hinge domain of Escherichia coli MukB, which forms a prokaryotic condensin complex with two non‐SMC subunits, MukE and MukF. In contrast with SMC hinge of Thermotoga maritima which has a sizable central hole at the dimer interface, MukB hinge forms a constricted dimer interface lacking a hole. Under our assay conditions, MukB hinge does not interact with DNA in accordance with the absence of a notable positively charged surface patch. The function of MukB hinge appears to be limited to dimerization of two copies of MukB molecules. Proteins 2010. © 2009 Wiley‐Liss, Inc.     

H2A.Z-Dependent Regulation of Cohesin Dynamics on Chromosome Arms

Structural maintenance of chromosomes (SMC) complexes and DNA topoisomerases are major determinants of chromosome structure and dynamics. The cohesin complex embraces sister chromatids throughout interphase, but during mitosis most cohesin is stripped from chromosome arms by early prophase, while the remaining cohesin at kinetochores is cleaved at anaphase. This two-step removal of cohesin is required for sister chromatids to separate. The cohesin-related Smc5/6 complex has been studied mostly as a determinant of DNA repair via homologous recombination. However, chromosome segregation fails in Smc5/6 null mutants or cells treated with small interfering RNAs. This also occurs in Smc5/6 hypomorphs in the fission yeast Schizosaccharomyces pombe following genotoxic and replication stress, or topoisomerase II dysfunction, and these mitotic defects are due to the postanaphase retention of cohesin on chromosome arms. Here we show that mitotic and repair roles for Smc5/6 are genetically separable in S. pombe. Further, we identified the histone variant H2A.Z as a critical factor to modulate cohesin dynamics, and cells lacking H2A.Z suppress the mitotic defects conferred by Smc5/6 dysfunction. Together, H2A.Z and the SMC complexes ensure genome integrity through accurate chromosome segregation.