From the XXVII North American Testis Workshop: The Function of SMC and Other Cohesin Proteins in Meiosis

To develop into haploid gametes, germ cells must undergo meiosis, a 2-step cell division process that is characterized by a sophisticated design in chromosome architecture and highly specific dynamic behavior of chromosomes. After premeiotic duplication, homologous chromosomes, each with 2 connected sister chromatids, pair and synapse, exchange genetic material, and then align on the metaphase plate to be segregated in anaphase of the first meiotic division. In the second step, the remaining pairs of sister chromatids are separated, and 4 daughter cells, haploid gametes each with 1 chromatid per chromosome, are generated. An elaborate set of proteins promotes and regulates these structures and processes. Complexes built on the structural maintenance of chromosome (SMC) protein family take center stage in these events. In recent years, important new data were gathered on these complexes and their partners, and hypotheses were developed that carry us significantly further on our road toward an understanding of the meiotic process. This review summarizes the most significant developments in this field. The purpose of meiosis is to create haploid germ cells from diploid parental cells (Figure 1). Chromosomes are duplicated during premeiotic S-phase, which generates cells with 4 chromatids of each type of chromosome—2 maternal and 2 paternal—similar to the mitotic S phase. In contrast to mitosis, meiotic DNA replication is followed by 2 successive rounds of nuclear divisions called meiosis I and II, which result in haploid gametes containing 1 copy of each chromosome. At meiosis I, the reductional division, the homologous chromosomes move toward opposite poles, while the sister chromatids of each homolog remain connected, generating cells with 2 chromatids. This is followed by a second mitosislike equational division in which sister chromatids separate from each other and the haploid cells are generated. Errors in meiosis result in the production of aneuploid zygotes with devastating consequences. Aneuploidy is a key factor in ∼35% of spontaneous pregnancy losses and the most commonly recognized cause of mental retardation (Hassold and Hunt, 2001; Page and Hawley, 2003). . Chromosome dynamics in meiosis. Simplified scheme of both meiotic divisions with an emphasis on synaptonemal complex formation, recombination, and sister chromatid cohesion. During prophase of meiosis I, various chromatin rearrangements take place, which are, like the reductional division itself, unique to meiosis (Roeder, 1997; van Heemst and Heyting, 2000; Page and Hawley, 2003; Petronczki et al, 2003). Homologous chromosomes, each consisting of 2 sister chromatids, find each other and pair to form the “bivalent,” a structure containing all 4 sister chromatids. Maternal and paternal chromosomes pair and recombine to generate new combinations of alleles. Recombination occurs between 1 sister chromatid of each homolog and also serves to keep the homologs physically connected until metaphase I. The pairing between homologous chromosomes in prophase I is stabilized by tight axial associations called synapsis and is possible within the synaptonemal complex (SC), a proteinaceous ladderlike structure that consists of 2 axial elements (AEs) connected all along their length by transverse filaments (TF). In mammals, 3 proteins were identified as components of the SC: SYCP1 (named SCP1 in the rat) is a component of the TF, and SYCP2 (SCP2) and SYCP3 (SCP3) are constituents of the AE (reviewed in Moens et al, 1998). Thus, the 4 sister chromatids are specifically structured and behave uniquely during meiosis. SMC protein complexes are central in determining these features. Unless otherwise noted, this review focuses on mammalian (mouse) cohesins and their functions. Generally, cohesion is essential to hold sister chromatids together until they are separated at nuclear division. During the last few years, protein components of the mitotic cohesin complex were identified (for recent reviews, see Jessberger, 2002, 2003; Hagström and Meyer, 2003). In mammalian somatic cells, the cohesin complex consists of at least 4 different subunits, 2 of them belong to the SMC family of proteins (Figure 2). SMC proteins are highly dynamic proteins that modulate chromosome structures. Six members of the family, usually called SMC1 to SMC6 fall into 2 subgroups. One member of a subgroup heterodimerizes with 1 SMC of the other subgroup (Figure 2B). These dimers associate with other proteins to form large complexes and act in various chromosome-related processes (Table). For example, the SMC2-SMC4 heterodimer is part of the condensin complex, which contributes to chromosome condensation. The SMC1-SMC3 heterodimer constitutes the core of the cohesin complex. Cohesin is an essential multiprotein complex required for sister chromatid cohesion (ie, the ordered arrangement of the newly replicated daughter DNA duplices in S, G2, and early M phase of the cell cycle; Figure 2C). In the mitotic cohesin complex, the heterodimer associates with 2 non-SMC subunits: the Rad21 (Scc1) protein, a member of the kleisin protein family (Schleiffer et al, 2003), and an Scc3-type protein, of which there are several variants. In mitotic cells, these are either the SA1 or the SA2 proteins. Homologs of all these proteins were identified in many organisms, and they appear to be essential. If defective, chromosomes are not segregated properly during mitosis, aneuploidy is generated, and the cells die. There is also impaired DNA recombinational repair, which under normal circumstances after S phase usually uses the aligned sister chromatids. The meiotic cohesin significantly differs from its mitotic counterparts and is specifically adapted to accommodate the unique structural and dynamic needs of meiotic chromosomes. . Structural maintenance of chromosome (SMC) proteins. (A) Monomeric SMC; molecular mass between 130 and 170 kd. (B) Heterodimer of 2 SMC proteins (eg, SMC1 and SMC3) in their most frequent V-shaped configuration. Note that dimerization occurs through interaction of hinge domains. N- and C-terminal domains of the same SMC molecules interact. (C) Model for a ringlike cohesin embracing the 2 sister chromatids (according to Gruber et al, 2003). Although the protein composition of the mitotic cohesin complex is well understood, it is not yet entirely clear how cohesin interacts with the 2 sister chromatids. Recent data derived from the crystal structure of bacterial SMC hinge domains (Haering et al, 2002) and supported by biochemical protein interaction studies on yeast cohesin subunits (Gruber et al, 2003) suggest that cohesin forms a large proteinaceous ring within which sister chromatids could be entrapped after DNA replication (Figure 2C). Embracing of DNA strands by the cohesin ring creates a link between sister chromatids, which might be more topological than chemical (ie, through protein-DNA binding). In vivo evidence for this elegant model still needs to be substantiated. The meiotic cohesin, however, differs significantly from its mitotic counterparts. In fact, not just 1 cohesin but several cohesin complexes are found in meiotic cells (Table). All are built around the SMC1-SMC3 heterodimer. In mammalian meiotic cells, the SMC1 protein exists in 2 variants: the canonical SMC1, called SMC1α, and a meiosis-specific variant, SMC1β (Revenkova et al, 2001). SMC1β has only been found in vertebrate cells. There is no published evidence for SMC3 variants, but the RAD21 protein coexists with a meiosis-specific paralog named REC8. REC8 is highly conserved through evolution, and orthologs from yeast, plants, and vertebrates are described (DeVeaux et al, 1992; Klein et al, 1999; Watanabe and Nurse, 1999; Cai et al, 2003; Lee et al, 2003). The SA1 and SA2 proteins coexist with their meiosis-specific variant STAG3 (Pezzi et al, 2000; Prieto et al, 2001). In somatic cells, cohesion between sister chromatids is established during S phase, and only the fully assembled cohesin complex can provide cohesion. Data obtained by studying the yeast Schizosaccharomyces pombe suggested that at least Rec8 associates with chromosomes during the last premeiotic round of replication, just before entry into meiosis (Watanabe and Nurse, 1999). In mice, the equivalent might be spermatogonial cells that are about to enter preleptotene. Although REC8 was not observed in spermatogonia, immunofluorescence studies showed its presence in preleptotene cells (Eijpe et al, 2003). Interestingly, SMC1β was not found at this early stage of meiosis but appeared later in leptotene cells (Figure 3). Therefore, REC8, possibly within a complex with SMC1α, might provide an initial basis for sites of meiotic cohesion, onto which the complex assembles. In preleptotene REC8 forms AE-like filamentous chromatin structures, which are, however, still quite diffuse. SMC1α exists in preleptotene cells; thus, other cohesin components presumably are present in those cells as well, including SMC3 (Figure 3). In leptotene, zygotene, and pachytene, all known cohesin components localize to the AEs, although with different characteristics. Unlike SMC1β or SMC3, the SMC1α protein, which is detectable only until the end of prophase I, does not localize as uniformly throughout the chromosomal axes; rather, it appears concentrated in certain areas, displaying a dotlike pattern (Eijpe et al, 2000). This, together with its absence in later meiosis, led to the suggestion that SMC1β rather than SMC1α is primarily responsible for sister chromatid cohesion during meiosis (Revenkova et al, 2001). . Presence of cohesin components at different stages throughout meiosis. Localization on chromosome arms or at centromeres (if known) is indicated, as are controversial data. Some data suggest physical interactions between components of the SC and cohesin proteins (Eijpe et al, 2000; Lee et al 2003), but the assembly of filamentous cohesin structures on meiotic chromatin does not depend on SC formation. Mice lacking the SYCP3 protein, a component of the AE, still assemble filamentous cohesin structures, although the SC is absent (Pelttari et al, 2001). Thus a “cohesin core” exists that is believed to layer itself on top of and parallel to the AE. This cohesin core might, together with SC proteins, regulate the structure of meiotic chromosomes (eg, their compaction). In early meiosis, there are at least 2 types of cohesin complexes, probably more, but the situation is not yet clear. There is certainly an SMC1α- and at least 1 SMC1β-based complex. Both SMC1 variants dimerize with SMC3 but might associate with different non-SMC proteins. Recent coimmunoprecipitation experiments indicate the of 2 complexes in the of complexes to at least 3 (Revenkova et al, One SMC1α complex to the mitotic complex et al, et al, 2003; et al, et al, the other the meiotic STAG3 and REC8 proteins (Revenkova et al, in no coimmunoprecipitation of SMC1α with REC8 was observed et al, 2003). also on the chromosomal and of One that a mammalian complex of SMC3, and an at the centromeres until the metaphase to anaphase et al, that RAD21 is from the centromeres in I and is not present at meiosis centromeres et al, In yeast, Rec8 was found to associate either with a STAG3 or with an of the mitotic SA1 or SA2 cohesin subunits et al, 2003). Thus it appears as many of the combinations of the various cohesin or in germ cells. The composition of each complex and their chromosomal and to sister chromatid cohesion, AE formation, and meiotic are a of and be further is important to between sister chromatid cohesion in chromosome arms and cohesion. Although cohesion along with is in metaphase I, cohesion is until metaphase to keep the sister chromatids connected for their toward a in anaphase I. in anaphase are sister chromatids separated to their to opposite and of the haploid cells. REC8 might not only provide an early basis for cohesin assembly but also to be in cohesion throughout meiosis. In mammalian cells, SMC1α, and SMC3 are found all along the prophase chromosomes, possibly to cohesion along the chromosomal gathered from of a in SMC1β to an essential of SMC1β in sister chromatid cohesion in (Revenkova et al, the enter into but can be into metaphase by they all sister chromatid cohesion, cells with cohesion. SMC1β and SMC3 from the chromosome arms in prophase I, REC8 along the chromosome arms until metaphase I (Eijpe et al, 2003; Lee et al, 2003). At least in S the 2 different subunits with which Rec8 associates Rec8 the complex to chromosome and the complex to the centromeres et al, 2003). the most for the SMC1β complex is in cohesion the protein, along with REC8 and SMC3, with the centromeres until anaphase (Revenkova et al, 2001). In mice, in of develop further but in metaphase II, in which the absence of sister chromatid cohesion to of chromatids. are (Revenkova et al, If the SMC1β complex for cohesion in around pachytene, then other cohesin complexes cohesion early in meiosis. The complexes provide this SMC1α is and along with in prophase to metaphase I et al, cohesion is RAD21 also from chromosome arms to at the centromeres until the metaphase to anaphase to REC8, SMC3, and SMC1β et al, This the of 2 complexes are at the with REC8 and 1 with RAD21 as a kleisin other is still of Rec8 sister chromatid in yeast et al, not to be to cohesion. recent data on different of REC8 and RAD21 in the suggest that these proteins different et al, REC8 to sites that the it could of to the for of homologs in meiosis I, to the for yeast Rec8 et al, 2003). to et al RAD21 to the centromeres in meiosis and could be in of the to the for of sister chromatids. In et al suggest the presence of RAD21 only at meiosis I centromeres and its to rather than In an Prieto et al that within an complex only to chromosomes in prophase I and in metaphase I chromosomes in early RAD21 and SA2 were only to chromosomes or in the (Eijpe et al, 2000). This with the by et al in which the presence of RAD21 was along the chromosome arms in parallel with from on the of cohesins in cohesion until meiosis is by et al these not SMC1, SMC3, REC8, and RAD21 at the centromeres of meiosis II, they suggest that cohesins might not be required at all for cohesion in meiosis This is in contrast to studies in and S pombe et al, 1999; Watanabe and Nurse, and to on the presence of SMC3, REC8, and RAD21 at meiosis centromeres in mammalian (Eijpe et al, 2003; et al, 2001; Lee et al, 2003; et al, is also to with the of all cohesion, including cohesion in (Revenkova et al, These are on different to et al, other as these different might they on a meiotic for RAD21 and REC8 and on the of a of different complexes in mammalian The to this rather complex and and still quite of the of cohesins is the SMC1β cohesin complex was also to be essential for a of other in meiosis (Revenkova et al, or in this protein to form and of in a unique their chromosomes are about than those of (Figure Thus, it appears as this cohesin the length of meiotic chromosomes. might by the of chromatin into the (eg, at the of chromatin that from the If chromatin in be by in of chromatin the this the chromatin in the are about more from the than those of that the of DNA in the is the same in both and a for SMC1β in determining the of chromatin in chromatin in the filamentous core in early meiosis is The chromosomes in several other not properly to the nuclear as they in (ie, of the at on the nuclear for early prophase is synapsis of homologs is in the Both could be a of chromosome For example, the chromosomes might to the nuclear of and those might in the homolog for sister chromatid cohesion in prophase at both the chromosome arms and the it clear that the meiosis-specific SMC1β protein essential in meiotic chromosome dynamics and structure (Revenkova et al, . of the meiosis-specific variant of a structural maintenance of chromosome protein, the length of prophase I chromosomes to about chromosome of or were by immunofluorescence for the synaptonemal complex component SCP1 and the cohesin SMC3 has the vertebrate an SMC1 protein other meiosis with just In and presumably other the SMC1α is on the chromosome and might be during early meiosis. on chromosome in chromosome in could a variant during to of an SMC1 protein all throughout meiosis. SMC1β might later then early in meiosis, as by the described of sister chromatid cohesion at the centromeres and of cohesion along the arms is through of cohesins from the arms and The as to the that this of cohesion. During in yeast, sister chromatids are separated after of by a that to the same family as (reviewed in 2001; and 2003). after from its called which is at the metaphase to anaphase In however, the of cohesin is by a as chromosome in a with the and at the metaphase to anaphase of cohesin during the first stage in prophase a which RAD21 et al, 2000; et al, In meiotic yeast cells, from chromosome arms at the of anaphase I, but until the metaphase to anaphase (Watanabe and Nurse, as observed in (Eijpe et al, 2003). of yeast from chromosome arms is a result of by et al, 2000). of Rec8 along chromosome arms and at the centromeres by is also required in yeast for of homologs and of sister chromatids in meiosis I and II, et al, 2003). in the chromosomes to separate during meiosis I of of et al, or of for et al, 2000). In it that of homologs and of sister chromatids are in Both the and the are for from meiosis I to in et al, 2001; et al, 2001). in yeast and is required for the metaphase I to anaphase I The suggest that either on the of the or might the different that cohesion in mammalian meiosis those for yeast and In the meiosis I to meiosis also on of in chromosome in meiosis I et al, 2003). If REC8 from chromosomal arms at the metaphase I to anaphase I how are REC8 and cohesin at the it must until anaphase the basis of studies in it has been that regulates the of chromosome cohesion during both meiotic probably by of et al, In along the chromosome during meiosis, present at the and for sister chromatid cohesion during both meiotic divisions et al, 2003). Therefore, it is possible that of cohesin complexes on arms and centromeres could result in 2-step cohesion in the first and second meiotic to centromeres could also act as cohesion. The meiosis-specific in yeast et al, 2002) and the protein of et al, been suggested as The recent of the meiosis-specific protein and its as a of the Rec8 cohesin in and more into the molecular responsible for cohesion during meiosis I et al, et al, The of Rec8 is and meiotic cohesion to at centromeres after meiosis I in cells. was suggested that as a for it physically from exchange of DNA between the maternal and paternal chromosomes is essential for and is a of The of to a protein-DNA structure called which also to physically the homologs in prophase I that they can be properly aligned in metaphase for no chromosome is is for an important of cohesins in meiotic DNA An early for a from studies in S et al, or Rec8 are In of by not only in chromosome synapsis at pachytene, but also of DNA et al, to a DNA of In the of SMC1β and SMC3 to between chromosomes, which are in and and are to sites of suggested that these cohesins in meiotic in as well (Revenkova et al, 2001; et al, 2003). the cohesin REC8 appears to be absent from these chromosomal the of there exists a SMC complex that to and possibly on data obtained through of the the of a of cohesins in meiotic (Revenkova et al, of can be by the proteins and in The absence of SMC1β or in and significantly their in Interestingly, proteins that appear at early during meiotic (ie, at are appear by the SMC1β of can be observed also in metaphase I in which homologs fall of together their Thus, the SMC1β complex in DNA after its to of could this cohesin complex possibly might, for example, be in the assembly of proteins required for of the early in the DNA as cohesin might the that from and the of chromosomes. there is evidence for of cohesins in DNA repair, in the of through homologous between sister chromatids meiotic SMC proteins could also be in DNA in This not only be for the SMC1-SMC3 heterodimer in the of cohesin or a complex, but also for the SMC the which is known to act in DNA in mitotic cells and associates with other proteins, probably has chromosomal and is highly in the et al, 2001; et al, 2003). a recent et al, that in yeast an complex with proteins and is important for meiotic chromosome and Chromosome during cell division from chromatin to a metaphase chromosome (for see and 2003) and is required for of sister chromatids in mitosis, or of homologs and later of sister chromatids in meiotic anaphase I and II, The axial and of chromosomes is with of chromosomes and sister chromatids. is known about proteins and processes that specifically to meiotic chromosome but it is to the most important of mitotic chromosome condensation. In a protein complex called condensin was and characterized as a protein component of mitotic chromosomes et al, 3 non-SMC proteins, the highly conserved condensin 2 SMC a heterodimer of and in condensin subunits, all of which are essential in S to in chromosome of yeast or of chromosome including a more from prophase to and the of chromosome in anaphase and from impaired sister chromatid et al, et al, 2000; et al, 2001). Interestingly, chromosome is but still in the of or in that condensin is not for chromosome but rather for of the sister chromatids et al, a of the of condensin was in a cell et al, 2003). chromosome was but not sister chromatids to and the of was were also for cells in et al, 2003). is important for between sister chromatids, its or to be could be a for the in chromosome a second condensin complex, built on the same and proteins, that associates with an entirely different set of other proteins et al, 2003). Both condensin complexes to the of the of chromosomes et al, 2003). they the both complexes are by the in the cell from these recent studies that although important for a chromosome structure of chromosome is not essential for chromosome in vertebrate cells. condensin structures, how it promotes of mitotic chromosomes is not yet In condensin the of certain DNA and 1997; et al, 1999). This might to the of condensin in the The of condensin in meiosis is not well of condensin in S were by the of and characterized meiotic in condensin In the absence of condensin subunits in meiosis I, the homologs and chromosomes but both pairing and were significantly than in the of the SC was highly a of condensin in of meiosis-specific chromosome proteins. In anaphase I and II, the a of between chromosomes. this was on in also in of recombination, this chromosome was to the Thus the physical between chromosomes, which in condensin appear to be by or of The of and in the of et al, 2002) of the of condensin in meiotic cells. I to but at anaphase of meiosis II, the chromosomes to chromatin between the second and the maternal as in the other condensin is required for sister chromatid but it condensin has no on the first meiotic division. One the of a second complex in meiosis I. more is known about or in meiosis or processes. In the 2 highly homologous in both cause impaired and and et al, 2003). of the non-SMC condensin in was and the protein was to during et al, 2003). of by of not of the chromosomes in the but chromosomes and spontaneous of sister chromatid which occurs in the last few years, evidence has been that important of cohesin and SMC protein complexes in meiosis. SMC complexes are essential for chromosome in both meiotic the chromosome structure with and in prophase I, sister chromatid cohesion in meiosis I and II, chromosome as the to the nuclear and formation, and of DNA (ie, of these processes are essential for chromosome If defective, of chromosomes could and it is not to that of an SMC complex in meiotic cells constitutes of the for the frequent meiotic as the (Hassold and Hunt, 2001).