Single-oocyte transcriptome analysis reveals aging-associated effects influenced by life stage and calorie restriction

Chromosome segregation errors in oocytes lead to the production of aneuploid eggs, which are the leading cause of pregnancy loss and of several congenital diseases such as Down syndrome. The frequency of chromosome segregation errors in oocytes increases with maternal age, especially at a late stage of reproductive life. How aging at various life stages affects oocytes differently remains poorly understood. In this study, we describe aging-associated changes in the transcriptome profile of mouse oocytes throughout reproductive life. Our single-oocyte comprehensive RNA sequencing using RamDA-seq revealed that oocytes undergo transcriptome changes at a late reproductive stage, whereas their surrounding cumulus cells exhibit transcriptome changes at an earlier stage. Calorie restriction, a paradigm that reportedly prevents aging-associated egg aneuploidy, promotes a transcriptome shift in oocytes with the up-regulation of genes involved in chromosome segregation. This shift is accompanied by the improved maintenance of chromosomal cohesin, the loss of which is a hallmark of oocyte aging and causes chromosome segregation errors. These findings have implications for understanding how oocytes undergo aging-associated functional decline throughout their reproductive life in a context-dependent manner. Single oocytes and their surrounding cumulus cells were analyzed for each age/diet class in two independent experiments: 2 months (Young), 4months, 9 months (Middle), 14 months (Old), 9 months (Calorie Restriction) with hyperovulation, and 2 months without hyperovulation.卵母细胞（oocyte）的染色体分离错误会导致非整倍体卵子的产生，这是流产及唐氏综合征等多种先天性疾病的首要诱因。卵母细胞的染色体分离错误频率随母龄增长而升高，尤其在生殖寿命晚期阶段更为显著。目前学界对生命不同阶段的衰老如何差异化影响卵母细胞的机制仍知之甚少。 本研究刻画了小鼠卵母细胞在整个生殖寿命中与衰老相关的转录组谱（transcriptome profile）变化。我们采用RamDA-seq技术开展的单细胞全面RNA测序结果显示，卵母细胞在生殖寿命晚期才会出现转录组改变，而其周围的卵丘细胞（cumulus cell）则在更早阶段即发生转录组变化。 热量限制（calorie restriction）作为一种被报道可延缓衰老相关卵子非整倍体的干预范式，可促进卵母细胞发生转录组转变，上调与染色体分离相关的基因表达。该转变同时伴随染色体黏连蛋白（cohesin）维持功能的改善，而染色体黏连蛋白的丢失是卵母细胞衰老的标志性特征，亦是引发染色体分离错误的诱因。 上述研究结果有助于理解卵母细胞在整个生殖寿命中如何以情境依赖的方式发生衰老相关的功能衰退。本研究在两项独立实验中，针对各年龄/饮食组别分析了单个卵母细胞及其周围的卵丘细胞，所涉组别包括：2月龄（年轻组）、4月龄、9月龄（中年组）、14月龄（老年组）、经超数排卵处理的9月龄热量限制组，以及未进行超数排卵的2月龄组。