Condensin I Reveals New Insights on Mouse Meiotic Chromosome Structure and Dynamics

Chromosome shaping and individualization are necessary requisites to warrant the correct segregation of genomes in either mitotic or meiotic cell divisions. These processes are mainly prompted in vertebrates by three multiprotein complexes termed cohesin and condensin I and II. In the present study we have analyzed by immunostaining the appearance and subcellular distribution of condensin I in mouse mitotic and meiotic chromosomes. Our results demonstrate that in either mitotically or meiotically dividing cells, condensin I is loaded onto chromosomes by prometaphase. Condensin I is detectable as a fuzzy axial structure running inside chromatids of condensed chromosomes. The distribution of condensin I along the chromosome length is not uniform, since it preferentially accumulates close to the chromosome ends. Interestingly, these round accumulations found at the condensin I axes termini colocalized with telomere complexes. Additionally, we present the relative distribution of the condensin I and cohesin complexes in metaphase I bivalents. All these new data have allowed us to propose a comprehensive model for meiotic chromosome structure

A perikinetochoric ring defined by MCAK and Aurora-B as a novel centromere domain

Mitotic Centromere-Associated Kinesin (MCAK) is a member of the kinesin-13 subfamily of kinesin-related proteins. In mitosis, this microtubule-depolymerising kinesin seems to be implicated in chromosome segregation and in the correction of improper kinetochore-microtubule interactions, and its activity is regulated by the Aurora-B kinase. However, there are no published data on its behaviour and function during mammalian meiosis. We have analysed by immunofluorescence in squashed mouse spermatocytes, the distribution and possible function of MCAK, together with Aurora-B, during both meiotic divisions. Our results demonstrate that MCAK and Aurora-B colocalise at the inner domain of metaphase I centromeres. Thus, MCAK shows a “cone”-like three-dimensional distribution beneath and surrounding the closely associated sister kinetochores. During the second meiotic division, MCAK and Aurora-B also colocalise at the inner centromere domain as a band that joins sister kinetochores, but only during prometaphase II in unattached chromosomes. During chromosome congression to the metaphase II plate, MCAK relocalises and appears as a ring below each sister kinetochore. Aurora-B also relocalises to appear as a ring surrounding and beneath kinetochores but during late metaphase II. Our results demonstrate that the redistribution of MCAK at prometaphase II/metaphase II centromeres depends on tension across the centromere and/or on the interaction of microtubules with kinetochores. We propose that the perikinetochoric rings of MCAK and Aurora-B define a novel transient centromere domain at least in mouse chromosomes during meiosis. We discuss the possible functions of MCAK at the inner centromere domain and at the perikinetochoric ring during both meiotic divisions

Sex chromosomes, synapsis, and cohesins: a complex affair

The final publication is available at http://link.springer.co

Involvement of synaptonemal complex proteins in sex chromosome segregation during marsupial male meiosis

Marsupial sex chromosomes break the rule that recombination during first meiotic prophase is necessary to ensure reductional segregation during first meiotic division. It is widely accepted that in marsupials X and Y chromosomes do not share homologous regions, and during male first meiotic prophase the synaptonemal complex is absent between them. Although these sex chromosomes do not recombine, they segregate reductionally in anaphase I. We have investigated the nature of sex chromosome association in spermatocytes of the marsupial Thylamys elegans, in order to discern the mechanisms involved in ensuring their proper segregation. We focused on the localization of the axial/lateral element protein SCP3 and the cohesin subunit STAG3. Our results show that X and Y chromosomes never appear as univalents in metaphase I, but they remain associated until they orientate and segregate to opposite poles. However, they must not be tied by a chiasma since their separation precedes the release of the sister chromatid cohesion. Instead, we show they are associated by the dense plate, a SCP3-rich structure that is organized during the first meiotic prophase and that is still present at metaphase I. Surprisingly, the dense plate incorporates SCP1, the main protein of the central element of the synaptonemal complex, from diplotene until telophase I. Once sex chromosomes are under spindle tension, they move to opposite poles losing contact with the dense plate and undergoing early segregation. Thus, the segregation of the achiasmatic T. elegans sex chromosomes seems to be ensured by the presence in metaphase I of a synaptonemal complex-derived structure. This feature, unique among vertebrates, indicates that synaptonemal complex elements may play a role in chromosome segregation

Sequential Loading of Cohesin Subunits during the First Meiotic Prophase of Grasshoppers

A previous version of this article appeared as an Early Online Release on January 2, 2007 (doi:10.1371/journal.pgen.0030028.eor).The cohesin complexes play a key role in chromosome segregation during both mitosis and meiosis. They establish sister chromatid cohesion between duplicating DNA molecules during S-phase, but they also have an important role during postreplicative double-strand break repair in mitosis, as well as during recombination between homologous chromosomes in meiosis. An additional function in meiosis is related to the sister kinetochore cohesion, so they can be pulled by microtubules to the same pole at anaphase I. Data about the dynamics of cohesin subunits during meiosis are scarce; therefore, it is of great interest to characterize how the formation of the cohesin complexes is achieved in order to understand the roles of the different subunits within them. We have investigated the spatio-temporal distribution of three different cohesin subunits in prophase I grasshopper spermatocytes. We found that structural maintenance of chromosome protein 3 (SMC3) appears as early as preleptotene, and its localization resembles the location of the unsynapsed axial elements, whereas radiation-sensitive mutant 21 (RAD21) (sister chromatid cohesion protein 1, SCC1) and stromal antigen protein 1 (SA1) (sister chromatid cohesion protein 3, SCC3) are not visualized until zygotene, since they are located in the synapsed regions of the bivalents. During pachytene, the distribution of the three cohesin subunits is very similar and all appear along the trajectories of the lateral elements of the autosomal synaptonemal complexes. However, whereas SMC3 also appears over the single and unsynapsed X chromosome, RAD21 and SA1 do not. We conclude that the loading of SMC3 and the non-SMC subunits, RAD21 and SA1, occurs in different steps throughout prophase I grasshopper meiosis. These results strongly suggest the participation of SMC3 in the initial cohesin axis formation as early as preleptotene, thus contributing to sister chromatid cohesion, with a later association of both RAD21 and SA1 subunits at zygotene to reinforce and stabilize the bivalent structure. Therefore, we speculate that more than one cohesin complex participates in the sister chromatid cohesion at prophase I.This work was supported by grants BFU2005–05668-C03–01, BFU2006–06655, BFU2005–01266, BFU2005–02431, and BFU2006–04406 from Ministerio de Educación y Ciencia, España, and grants 1001160016 and 11/BCB/013 from Universidad Autónoma de Madrid and Comunidad de Madrid. The Department of Immunology and Oncology was founded and is supported by the Spanish Council for Scientific Research (CSIC).Peer reviewe

Sequential Loading of Cohesin Subunits during the First Meiotic Prophase of Grasshoppers

The cohesin complexes play a key role in chromosome segregation during both mitosis and meiosis. They establish sister chromatid cohesion between duplicating DNA molecules during S-phase, but they also have an important role during postreplicative double-strand break repair in mitosis, as well as during recombination between homologous chromosomes in meiosis. An additional function in meiosis is related to the sister kinetochore cohesion, so they can be pulled by microtubules to the same pole at anaphase I. Data about the dynamics of cohesin subunits during meiosis are scarce; therefore, it is of great interest to characterize how the formation of the cohesin complexes is achieved in order to understand the roles of the different subunits within them. We have investigated the spatio-temporal distribution of three different cohesin subunits in prophase I grasshopper spermatocytes. We found that structural maintenance of chromosome protein 3 (SMC3) appears as early as preleptotene, and its localization resembles the location of the unsynapsed axial elements, whereas radiation-sensitive mutant 21 (RAD21) (sister chromatid cohesion protein 1, SCC1) and stromal antigen protein 1 (SA1) (sister chromatid cohesion protein 3, SCC3) are not visualized until zygotene, since they are located in the synapsed regions of the bivalents. During pachytene, the distribution of the three cohesin subunits is very similar and all appear along the trajectories of the lateral elements of the autosomal synaptonemal complexes. However, whereas SMC3 also appears over the single and unsynapsed X chromosome, RAD21 and SA1 do not. We conclude that the loading of SMC3 and the non-SMC subunits, RAD21 and SA1, occurs in different steps throughout prophase I grasshopper meiosis. These results strongly suggest the participation of SMC3 in the initial cohesin axis formation as early as preleptotene, thus contributing to sister chromatid cohesion, with a later association of both RAD21 and SA1 subunits at zygotene to reinforce and stabilize the bivalent structure. Therefore, we speculate that more than one cohesin complex participates in the sister chromatid cohesion at prophase I

Meiotic Pairing and Segregation of Achiasmate Sex Chromosomes in Eutherian Mammals: The Role of SYCP3 Protein

In most eutherian mammals, sex chromosomes synapse and recombine during male meiosis in a small region called pseudoautosomal region. However in some species sex chromosomes do not synapse, and how these chromosomes manage to ensure their proper segregation is under discussion. Here we present a study of the meiotic structure and behavior of sex chromosomes in one of these species, the Mongolian gerbil (Meriones unguiculatus). We have analyzed the location of synaptonemal complex (SC) proteins SYCP1 and SYCP3, as well as three proteins involved in the process of meiotic recombination (RAD51, MLH1, and γ-H2AX). Our results show that although X and Y chromosomes are associated at pachytene and form a sex body, their axial elements (AEs) do not contact, and they never assemble a SC central element. Furthermore, MLH1 is not detected on the AEs of the sex chromosomes, indicating the absence of reciprocal recombination. At diplotene the organization of sex chromosomes changes strikingly, their AEs associate end to end, and SYCP3 forms an intricate network that occupies the Y chromosome and the distal region of the X chromosome long arm. Both the association of sex chromosomes and the SYCP3 structure are maintained until metaphase I. In anaphase I sex chromosomes migrate to opposite poles, but SYCP3 filaments connecting both chromosomes are observed. Hence, one can assume that SYCP3 modifications detected from diplotene onwards are correlated with the maintenance of sex chromosome association. These results demonstrate that some components of the SC may participate in the segregation of achiasmate sex chromosomes in eutherian mammals

Sequential Assembly of Centromeric Proteins in Male Mouse Meiosis

The assembly of the mitotic centromere has been extensively studied in recent years, revealing the sequence and regulation of protein loading to this chromosome domain. However, few studies have analyzed centromere assembly during mammalian meiosis. This study specifically targets this approach on mouse spermatocytes. We have found that during prophase I, the proteins of the chromosomal passenger complex Borealin, INCENP, and Aurora-B load sequentially to the inner centromere before Shugoshin 2 and MCAK. The last proteins to be assembled are the outer kinetochore proteins BubR1 and CENP-E. All these proteins are not detected at the centromere during anaphase/telophase I and are then reloaded during interkinesis. The loading sequence of the analyzed proteins is similar during prophase I and interkinesis. These findings demonstrate that the interkinesis stage, regularly overlooked, is essential for centromere and kinetochore maturation and reorganization previous to the second meiotic division. We also demonstrate that Shugoshin 2 is necessary for the loading of MCAK at the inner centromere, but is dispensable for the loading of the outer kinetochore proteins BubR1 and CENP-E

A High Incidence of Meiotic Silencing of Unsynapsed Chromatin Is Not Associated with Substantial Pachytene Loss in Heterozygous Male Mice Carrying Multiple Simple Robertsonian Translocations

Meiosis is a complex type of cell division that involves homologous chromosome pairing, synapsis, recombination, and segregation. When any of these processes is altered, cellular checkpoints arrest meiosis progression and induce cell elimination. Meiotic impairment is particularly frequent in organisms bearing chromosomal translocations. When chromosomal translocations appear in heterozygosis, the chromosomes involved may not correctly complete synapsis, recombination, and/or segregation, thus promoting the activation of checkpoints that lead to the death of the meiocytes. In mammals and other organisms, the unsynapsed chromosomal regions are subject to a process called meiotic silencing of unsynapsed chromatin (MSUC). Different degrees of asynapsis could contribute to disturb the normal loading of MSUC proteins, interfering with autosome and sex chromosome gene expression and triggering a massive pachytene cell death. We report that in mice that are heterozygous for eight multiple simple Robertsonian translocations, most pachytene spermatocytes bear trivalents with unsynapsed regions that incorporate, in a stage-dependent manner, proteins involved in MSUC (e.g., γH2AX, ATR, ubiquitinated-H2A, SUMO-1, and XMR). These spermatocytes have a correct MSUC response and are not eliminated during pachytene and most of them proceed into diplotene. However, we found a high incidence of apoptotic spermatocytes at the metaphase stage. These results suggest that in Robertsonian heterozygous mice synapsis defects on most pachytene cells do not trigger a prophase-I checkpoint. Instead, meiotic impairment seems to mainly rely on the action of a checkpoint acting at the metaphase stage. We propose that a low stringency of the pachytene checkpoint could help to increase the chances that spermatocytes with synaptic defects will complete meiotic divisions and differentiate into viable gametes. This scenario, despite a reduction of fertility, allows the spreading of Robertsonian translocations, explaining the multitude of natural Robertsonian populations described in the mouse

NOR and nucleolus in the spermatogenesis of acridoid grasshoppers

6 p.-3 fig.-1 tab.By means of silver staining procedures of light microscopy the characteristics of the nucleolus and the NORs have been investigated in meiocytes of different grasshopper species. Our results show that: (1) Two is the most common number of chromosomes per haploid genome carrying active NORs although this number may vary from one up to five; (2) NOR activity is preferentially located on medium and short chromosomes but the X and the megameric chromosome are involved in nucleolar organization in a high proportion of the species studied; (3) The NOR location is normally restricted to one end in acro-telocentrics and to the short arm, near the centromere region, in metacentrics; (4) A marked correlation is observed between the number of nucleoli present in the spermatogonial cells and in the first meiotic prophase of a given species; (5) In some cases, the nucleoli are associated to chromosomes during spermatogonial premetaphases.This work was supported by the Comisión Asesora para la Investigación Científica y Técnica (Spain).Peer reviewe