Department of Cell, Developmental and Cancer Biology, Oregon Health and Science University School of Medicine, Portland, Oregon 97239, USA.
Division of Reproductive and Developmental Sciences, Oregon National Primate Research Center, Beaverton, Oregon 97006, USA.
Genome Res. 2019 Mar;29(3):367-382. doi: 10.1101/gr.239830.118. Epub 2019 Jan 25.
Aneuploidy that arises during meiosis and/or mitosis is a major contributor to early embryo loss. We previously showed that human preimplantation embryos encapsulate missegregated chromosomes into micronuclei while undergoing cellular fragmentation and that fragments can contain chromosomal material, but the source of this DNA was unknown. Here, we leveraged the use of a nonhuman primate model and single-cell DNA-sequencing (scDNA-seq) to examine the chromosomal content of 471 individual samples comprising 254 blastomeres, 42 polar bodies, and 175 cellular fragments from a large number ( = 50) of disassembled rhesus cleavage-stage embryos. Our analysis revealed that the aneuploidy and micronucleation frequency is conserved between humans and macaques, and that fragments encapsulate whole and/or partial chromosomes lost from blastomeres. Single-cell/fragment genotyping showed that these chromosome-containing cellular fragments (CCFs) can be maternally or paternally derived and display double-stranded DNA breaks. DNA breakage was further indicated by reciprocal subchromosomal losses/gains between blastomeres and large segmental errors primarily detected at the terminal ends of chromosomes. By combining time-lapse imaging with scDNA-seq, we determined that multipolar divisions at the zygote or two-cell stage were associated with CCFs and generated a random mixture of chromosomally normal and abnormal blastomeres with uniparental or biparental origins. Despite frequent chromosome missegregation at the cleavage-stage, we show that CCFs and nondividing aneuploid blastomeres showing extensive DNA damage are prevented from incorporation into blastocysts. These findings suggest that embryos respond to chromosomal errors by encapsulation into micronuclei, elimination via cellular fragmentation, and selection against highly aneuploid blastomeres to overcome chromosome instability during preimplantation development.
减数分裂和/或有丝分裂过程中出现的非整倍体是导致早期胚胎丢失的主要原因。我们之前曾表明，人类胚胎在经历细胞碎片化时会将错误分离的染色体包裹到微核中，而且这些碎片可能含有染色体物质，但这些 DNA 的来源尚不清楚。在这里，我们利用非人类灵长类动物模型和单细胞 DNA 测序 (scDNA-seq) 来检查 254 个卵裂球、42 个极体和 175 个细胞碎片的 471 个个体样本的染色体含量，这些样本来自大量（= 50）拆开的恒河猴卵裂期胚胎。我们的分析表明，人类和猕猴之间的非整倍体和微核形成频率是保守的，而且这些碎片包裹着从卵裂球丢失的完整和/或部分染色体。单细胞/碎片基因分型表明，这些含有染色体的细胞碎片 (CCF) 可以来自母体或父体，并显示双链 DNA 断裂。卵裂球之间的相互亚染色体丢失/增益和主要在染色体末端检测到的大节段错误进一步表明了 DNA 断裂。通过将延时成像与 scDNA-seq 相结合，我们确定了合子或 2 细胞期的多极分裂与 CCF 有关，并产生了具有单亲或双亲来源的正常和异常卵裂球的随机混合物。尽管在卵裂期经常发生染色体错误分离，但我们发现 CCF 和表现出广泛 DNA 损伤的非分裂非整倍体卵裂球被阻止整合到囊胚中。这些发现表明，胚胎通过包裹到微核中、通过细胞碎片化消除以及选择具有高度非整倍体的卵裂球来应对染色体错误，从而克服胚胎植入前发育过程中的染色体不稳定性。
