Sister chromatid cohesion ensures the faithful segregation of chromosomes in mitosis and in both meiotic divisions. Meiosis-specific components of the cohesin complex, including the recently described SMC1 isoform SMC1 beta, were suggested to be required for meiotic sister chromatid cohesion and DNA recombination. Here we show that SMC1 beta-deficient mice of both sexes are sterile. Male meiosis is blocked in pachytene; female meiosis is highly error-prone but continues until metaphase II. Prophase axial elements (AEs) are markedly shortened, chromatin extends further from the AEs, chromosome synapsis is incomplete, and sister chromatid cohesion in chromosome arms and at centromeres is lost prematurely. In addition, crossover-associated recombination foci are absent or reduced, and meiosis-specific perinuclear telomere arrangements are impaired. Thus, SMC1 beta has a key role in meiotic cohesion, the assembly of AEs, synapsis, recombination, and chromosome movements.

n meiotic prophase, the sister chromatids of each chromosome develop a common axial element (AE) that is integrated into the synaptonemal complex (SC). We analyzed the incorporation of sister chromatid cohesion proteins (cohesins) and other AE components into AEs. Meiotic cohesin REC8 appeared shortly before premeiotic S phase in the nucleus and formed AE-like structures (REC8-AEs) from premeiotic S phase on. Subsequently, meiotic cohesin SMC1 ␤ , cohesin SMC3, and AE proteins SCP2 and SCP3 formed dots along REC8-AEs, which extended and fused until they lined REC8-AEs along their length. In metaphase I, SMC1 ␤ , SMC3, SCP2, and SCP3 disappeared from the I chromosome arms and accumulated around the centromeres, where they stayed until anaphase II. In striking contrast, REC8 persisted along the chromosome arms until anaphase I and near the centromeres until anaphase II. We propose that REC8 provides a basis for AE formation and that the first steps in AE assembly do not require SMC1 ␤ , SMC3, SCP2, and SCP3. Furthermore, SMC1 ␤ , SMC3, SCP2, and SCP3 cannot provide arm cohesion during metaphase I. We propose that REC8 then provides cohesion. RAD51 and/or DMC1 coimmunoprecipitates with REC8, suggesting that REC8 may also provide a basis for assembly of recombination complexes.

Structural maintenance of chromosomes (SMC) proteins fulfill pivotal roles in chromosome dynamics. In yeast, the SMC1-SMC3 heterodimer is required for meiotic sister chromatid cohesion and DNA recombination. Little is known, however, about mammalian SMC proteins in meiotic cells. We have identified a novel SMC protein (SMC1␤), which-except for a unique, basic, DNA binding C-terminal motif-is highly homologous to SMC1 (which may now be called SMC1␣) and is not present in the yeast genome. SMC1␤ is specifically expressed in testes and coimmunoprecipitates with SMC3 from testis nuclear extracts, but not from a variety of somatic cells. This establishes for mammalian cells the concept of cell-type-and tissue-specific SMC protein isoforms. Analysis of testis sections and chromosome spreads of various stages of meiosis revealed localization of SMC1␤ along the axial elements of synaptonemal complexes in prophase I. Most SMC1␤ dissociates from the chromosome arms in late-pachytene-diplotene cells. However, SMC1␤, but not SMC1␣, remains chromatin associated at the centromeres up to metaphase II. Thus, SMC1␤ and not SMC1␣ is likely involved in maintaining cohesion between sister centromeres until anaphase II.The eukaryotic, evolutionarily highly conserved SMC (Structural Maintenance of Chromosomes) proteins are involved in several key DNA and chromatin dynamic processes (for recent reviews, see references 11, 21, 26, 27, 31, 48, 60, and 62). The best-documented processes are chromosome condensation and sister chromatid cohesion. Evidence is also accumulating for a function in DNA recombination and repair. A fourth role of SMC proteins is in gene dosage compensation in Caenorhabditis elegans. The phylogenetic tree comprises five subfamilies (32): SMC1 to SMC4 and an ancestral family that includes the recently defined SMC5 and SMC6 groups with the Rad18 and Spr18 proteins of Schizosaccharomyces pombe (16), which act in recombinational repair.SMC proteins share a characteristic design. Coiled-coil domains are flanked by globular N-and C-terminal domains and are divided in the central region by a flexible hinge domain of about 150 aa. The N-and C-terminal domains of about 100 to 150 aa are highly conserved and carry important motifs. The N-terminal domain includes an NTP binding motif (Walker A box [68]), which has been shown to bind the ATP analogue azido-ATP (1). The C-terminal domain contains a DA box (68). The C-terminal and second coiled-coil domains, but not the N terminus, bind DNA (1, 2). It has been proposed that the antiparallel, heterodimeric SMC1-SMC3 protein with an N and C terminus at each end may connect two DNA molecules, such as sister chromatids, and may directly contribute to their alignment in cohesion and to recombination between sister chromatids (2, 26, 62).In eukaryotes the SMC1-SMC3 or SMC2-SMC4 heterodimers form large multiprotein complexes. One of these complexes is condensin, which, besides the SMC2-SMC4 heterodimer, contains several non-SMC subunits. Condensin is necessary for mitotic chromosome c...

SUMMARY Pluripotent stem cell lines can be derived from blastocyst embryos, which yield embryonic stem cell lines (ES cells), as well as the post-implantation epiblast, which gives rise to epiblast stem cell lines (EpiSCs). Remarkably, ES cells and EpiSCs display profound differences in the combination of growth factors that maintain their pluripotent state. Molecular and functional differences between these two stem cell types demonstrate that the tissue of origin and/or the growth factor milieu may be important determinants of the stem cell identity. We explored how developmental stage of the tissue of origin and culture growth factor conditions affect the stem cell pluripotent state. Our findings reveal that novel stem cell lines can be generated from blastocyst embryos with unique functional and molecular properties. We demonstrate that the culture growth factor environment and cell-cell interaction play a critical role in defining several unique and stable stem cell ground states.

Double-strand DNA breaks (DSBs) pose a major threat to living cells, and several mechanisms for repairing these lesions have evolved. Eukaryotes can process DSBs by homologous recombination (HR) or non-homologous end joining (NHEJ). NHEJ connects DNA ends irrespective of their sequence, and it predominates in mitotic cells, particularly during G1 (ref. 3). HR requires interaction of the broken DNA molecule with an intact homologous copy, and allows restoration of the original DNA sequence. HR is active during G2 of the mitotic cycle and predominates during meiosis, when the cell creates DSBs (ref. 4), which must be repaired by HR to ensure proper chromosome segregation. How the cell controls the choice between the two repair pathways is not understood. We demonstrate here a physical interaction between mammalian Ku70, which is essential for NHEJ (ref. 5), and Mre11, which functions both in NHEJ and meiotic HR (Refs 2,6). Moreover, we show that irradiated cells deficient for Ku70 are incapable of targeting Mre11 to subnuclear foci that may represent DNA-repair complexes. Nevertheless, Ku70 and Mre11 were differentially expressed during meiosis. In the mouse testis, Mre11 and Ku70 co-localized in nuclei of somatic cells and in the XY bivalent. In early meiotic prophase, however, when meiotic recombination is most probably initiated, Mre11 was abundant, whereas Ku70 was not detectable. We propose that Ku70 acts as a switch between the two DSB repair pathways. When present, Ku70 destines DSBs for NHEJ by binding to DNA ends and attracting other factors for NHEJ, including Mre11; when absent, it allows participation of DNA ends and Mre11 in the meiotic HR pathway.