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

Hwang, Grace H.; Sun, Fengyun; O’Brien, Marilyn J.; Eppig, John J.; Handel, Mary Ann; Jordan, Philip W.

doi:10.1242/dev.145607

Cited by 32 publications

(36 citation statements)

References 74 publications

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“…2D). Specifically, SMC5 for example has been shown to be important for correct segregation of homologous chromosomes during meiosis I in mouse oocytes and, in contrast to our human RNA data, that protein levels of SMC5/6 declined in mice oocytes with age (Hwang et al, 2017). Why did we observe an increase in the RNA-levels of chromosome segregation genes with age?…”

Section: Maturation Stage Is the Main Differentiator Of Oocyte Transccontrasting

confidence: 75%

Single cell analysis reveals the impact of age and maturation stage on the human oocyte transcriptome

Llonch

Barragán²,

Nieto

et al. 2020

Preprint

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Study questionTo which degree does maternal age affect the transcriptome of human oocytes at the germinal vesicle (GV) stage or at metaphase II after maturation in vitro (IVM-MII)?Summary answerWhile the oocytes’ transcriptome is predominantly determined by maturation stage, transcript levels of genes related to chromosome segregation, mitochondria and RNA processing are affected by age after in vitro maturation of denuded oocytes.What is known alreadyFemale fertility is inversely correlated with maternal age due to both a depletion of the oocyte pool and a reduction in oocyte developmental competence. Few studies have addressed the effect of maternal age on the human mature oocyte (MII) transcriptome, which is established during oocyte growth and maturation, and the pathways involved remain unclear. Here, we characterize and compare the transcriptomes of a large cohort of fully grown GV and IVM-MII oocytes from women of varying reproductive age.Study design, size, durationIn this prospective molecular study, 37 women were recruited from May 2018 to June 2019. The mean age was 28.8 years (SD=7.7, range 18-43). A total of 72 oocytes were included in the study at GV stage after ovarian stimulation, and analyzed as GV (n=40) and in vitro matured oocytes (IVM-MII; n=32).Participants/materials, setting, methodsDenuded oocytes were included either as GV at the time of ovum pick-up or as IVM-MII after in vitro maturation for 30 hours in G2™ medium, and processed for transcriptomic analysis by single-cell RNA-seq using the Smart-seq2 technology. Cluster and maturation stage marker analysis were performed using the Seurat R package. Genes with an average fold change greater than 2 and a p-value < 0.01 were considered maturation stage markers. A Pearson correlation test was used to identify genes whose expression levels changed progressively with age. Those genes presenting a correlation value (R) >= |0.3| and a p-value < 0.05 were considered significant.Main results and the role of chanceFirst, by exploration of the RNA-seq data using tSNE dimensionality reduction, we identified two clusters of cells reflecting the oocyte maturation stage (GV and IVM-MII) with 4,445 and 324 putative marker genes, respectively. Next we identified genes, for which RNA levels either progressively increased or decreased with age. This analysis was performed independently for GV and IVM-MII oocytes. Our results indicate that the transcriptome is more affected by age in IVM-MII oocytes (1,219 genes) than in GV oocytes (596 genes). In particular, we found that genes involved in chromosome segregation and RNA splicing significantly increase in transcript levels with age, while genes related to mitochondrial activity present lower transcript levels with age. Gene regulatory network analysis revealed potential upstream master regulator functions for genes whose transcript levels present positive (GPBP1, RLF, SON, TTF1) or negative (BNC1, THRB) correlation with age.Limitations, reasons for cautionIVM-MII oocytes used in this study were obtained after in vitro maturation of denuded GV oocytes, therefore, their transcriptome might not be fully representative of in vivo matured MII oocytes.The Smart-seq2 methodology used in this study detects polyadenylated transcripts only and we could therefore not assess non-polyadenylated transcripts.Wider implications of the findingsOur analysis suggests that advanced maternal age does not globally affect the oocyte transcriptome at GV or IVM-MII stages. Nonetheless, hundreds of genes displayed altered transcript levels with age, particularly in IVM-MII oocytes. Especially affected by age were genes related to chromosome segregation and mitochondrial function, pathways known to be involved in oocyte ageing. Our study thereby suggests that misregulation of chromosome segregation and mitochondrial pathways also at the RNA-level might contribute to the age-related quality decline in human oocytes.Study funding/competing interest(s)This study was funded by the AXA research fund, the European commission, intramural funding of Clinica EUGIN, the Spanish Ministry of Science, Innovation and Universities, the Catalan Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR) and by contributions of the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership and to the “Centro de Excelencia Severo Ochoa”.The authors have no conflict of interest to declare.

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Section: Maturation Stage Is the Main Differentiator Of Oocyte Transccontrasting

confidence: 75%

Single cell analysis reveals the impact of age and maturation stage on the human oocyte transcriptome

Llonch

Barragán²,

Nieto

et al. 2020

Preprint

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“…Since Nestin is expressed in progenitor cells of tissues other than neural, some degree of DNA excision was also observed in kidneys and muscles in our samples ( Figure 1B and Figure 1-figure supplement 1A) (Bernal et al, 2018;Sakairi et al, 2007). SMC5 depletion in the developing mouse cortex resulted in a significant increase in cleaved caspase 3 (CASP3)-positive cells in Smc5 cKO compared to controls in both, E13.5 and E16.5 cortices ( Figure We have previously shown that the depletion of SMC5 in mESCs causes chromosome segregation defects during mitosis (Hwang et al, 2017;Pryzhkova and Jordan, 2016). Thus, we analyzed VZ and SVZ mitotic progenitors in cortical sections of E13.5 and E16.…”

Section: Smc5 Cko Causes Neurodevelopmental and Sensorimotor Defects mentioning

confidence: 97%

SMC5/6 is required for replication fork stability and faithful chromosome segregation during neurogenesis

Atkins

et al. 2020

eLife

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Mutations of SMC5/6 components cause developmental defects, including primary microcephaly. To model neurodevelopmental defects, we engineered a mouse wherein Smc5 is conditionally knocked out (cKO) in the developing neocortex. Smc5 cKO mice exhibited neurodevelopmental defects due to neural progenitor cell (NPC) apoptosis, which led to reduction in cortical layer neurons. Smc5 cKO NPCs formed DNA bridges during mitosis and underwent chromosome missegregation. SMC5/6 depletion triggers a CHEK2-p53 DNA damage response, as concomitant deletion of the Trp53 tumor suppressor or Chek2 DNA damage checkpoint kinase rescued Smc5 cKO neurodevelopmental defects. Further assessment using Smc5 cKO and auxin-inducible degron systems demonstrated that absence of SMC5/6 leads to DNA replication stress at late-replicating regions such as pericentromeric heterochromatin regions. In summary, SMC5/6 is important for completion of DNA replication prior to entering mitosis, which ensures accurate chromosome segregation. Thus, SMC5/6 functions are critical in highly proliferative stem cells during organism development.

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“…Structural maintenance of chromosome 5/6 (SMC5/6) is an evolutionarily conserved protein complex ( 14 , 15 ) that consists of SMC5 and SMC6 along with the NSE1, -2, -3, and -4 proteins ( 16 – 18 ) and plays irreplaceable roles in DNA repair, chromosome segregation, and telomere length maintenance ( 19 – 22 ). Interestingly, in addition to its essential role in chromosome maintenance, the SMC5/6 complex was identified as a host restriction factor against HBV ( 23 – 25 ), but the detailed mechanism underlying the function of the complex in silencing DNA viruses and episomal plasmids remains unknown.…”

Section: Introductionmentioning

confidence: 99%

PJA1 Coordinates with the SMC5/6 Complex To Restrict DNA Viruses and Episomal Genes in an Interferon-Independent Manner

Zhang

et al. 2018

J Virol

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DNA viruses, including hepatitis B virus and herpes simplex virus, induce a series of immune responses in the host and lead to human public health concerns worldwide. In addition to cytokines in the cytoplasm, restriction of viral DNA in the nucleus is an important approach of host immunity. However, the mechanism of foreign DNA recognition and restriction in the cell nucleus is largely unknown. This work demonstrates that an important cellular factor (PJA1) suppresses DNA viruses and transfected plasmids independent of type I and II interferon (IFN) pathways. Instead, PJA1 interacts with the chromosome maintenance complex (SMC5/6), facilitates the complex to recognize and bind viral and episomal DNAs, and recruits DNA topoisomerases to restrict the foreign molecules. These results reveal a distinct mechanism underlying the silencing of viral and episomal invaders in the cell nuclei and suggest that PJA1 acts as a potential agent to prevent infectious and inflammatory diseases.

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

Cited by 32 publications

References 74 publications

Single cell analysis reveals the impact of age and maturation stage on the human oocyte transcriptome

Single cell analysis reveals the impact of age and maturation stage on the human oocyte transcriptome

SMC5/6 is required for replication fork stability and faithful chromosome segregation during neurogenesis

PJA1 Coordinates with the SMC5/6 Complex To Restrict DNA Viruses and Episomal Genes in an Interferon-Independent Manner

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