Conditional mutation of Smc5 in mouse embryonic stem cells perturbs condensin localization and mitotic progression

Abstract

Correct duplication of stem cell genetic material and its appropriate segregation into daughter cells are requisites for tissue, organ and organism homeostasis. Disruption of stem cell genomic integrity can lead to developmental abnormalities and cancer. Roles of the Smc5/6 structural maintenance of chromosomes complex in pluripotent stem cell genome maintenance have not been investigated, despite its important roles in DNA synthesis, DNA repair and chromosome segregation as evaluated in other model systems. Using mouse embryonic stem cells (mESCs) with a conditional knockout allele of Smc5, we showed that Smc5 protein depletion resulted in destabilization of the Smc5/6 complex, accumulation of cells in G2 phase of the cell cycle and apoptosis. Detailed assessment of mitotic mESCs revealed abnormal condensin distribution and perturbed chromosome segregation, accompanied by irregular spindle morphology, lagging chromosomes and DNA bridges. Mutation of Smc5 resulted in retention of Aurora B kinase and enrichment of condensin on chromosome arms. Furthermore, we observed reduced levels of Polo-like kinase 1 at kinetochores during mitosis. Our study reveals crucial requirements of the Smc5/6 complex during cell cycle progression and for stem cell genome maintenance.

SMC5/6 is required for the formation of segregation-competent bivalent chromosomes during meiosis I in mouse oocytes

“…Therefore, it was hypothesized that mutation of Smc5 would affect TOP2A localization in mouse oocytes. However, no defects in TOP2A localization were observed, which corresponds to what has been reported for Smc5 cKO in mouse embryonic stem cells (Pryzhkova and Jordan, 2016). Studies of yeast SMC5/6 have shown that the complex is linked with TopoIIdependent catenation/decatenation functions (Jeppsson et al, 2014;Kanno et al, 2015;Kegel et al, 2011).…”

“…S5). This phenotype is reminiscent of the mitotic catastrophe observed in Smc5 cKO mouse embryonic stem cells (Pryzhkova and Jordan, 2016). Fig.…”

“…RNAi depletion of SMC5 and SMC6 in human RPE-1 cells resulted in defective axial localization of condensin (Gallego-Paez et al, 2013). Abnormal condensin localization was also observed using Smc5 cKO mouse embryonic stem cells (Pryzhkova and Jordan, 2016). Furthermore, mutation of smc-5 in C. elegans leads to abnormal distribution of condensin along bivalents during meiosis I (Hong et al, 2016).…”

“…Mice harboring Smc5 with a floxed exon 4 (designated Smc5 flox ) and deleted exon 4 (designated Smc5 del ) have been previously described (Pryzhkova and Jordan, 2016). Heterozygous Smc5 del mice were bred to mice harboring the Zp3-Cre transgene [C57BL/6-Tg(Zp3-cre)93Knw/J], which resulted in progeny heterozygous for the Smc5 del allele and hemizygous for the Zp3-Cre transgene (Smc5 +/del , Zp3-Cre tg/0).…”

In‐frame deletion of SMC5 related with the phenotype of primordial dwarfism, chromosomal instability and insulin resistance

“…Furthermore, the impaired adipogenesis was revealed to be caused by G2/M phase arrest during the process of MCE. Accordingly, in mouse embryonic stem cells with a conditional KO of Smc5, Pryzhkova et al 47 found that the downregulation of Smc5 results in cells accumulating in the G2 phase. Moreover, mitotic defects are apparent only when the function of SMC5/6 is compromised during the S/G2 phases, suggesting its crucial role in the sister chromatid disjunction.…”