Single-cell transcriptomics and translatomics dual-omics analysis reveals the potential mechanism of HFD-induced maternal RNA degradation abnormalities
This study explores the impact of high-fat diet (HFD)-induced obesity on maternal RNA degradation and its implications for oocyte quality and development in mice, using a novel transcriptome and translatome sequencing (T&T-seq) strategy. By combining RiboLace for translatome profiling and Smart-seq for transcriptome analysis, the research investigates how HFD disrupts RNA degradation pathways during oocyte maturation, particularly from the germinal vesicle (GV) to metaphase II (MII) stages. The findings reveal that HFD impairs the degradation of key transcripts such as BTG4 and CNOT7, and alters the translation efficiency of meiotic regulators, which are crucial for proper meiotic progression. This study provides new insights into how maternal dietary factors influence oocyte quality at the molecular level, with potential implications for fertility and embryonic development. Overall design: We employed T&T-seq (translatome and transcriptome sequencing) to comprehensively analyze gene expression profiles in GV oocytes and MII oocytes from both high-fat diet (HFD) and normal diet (ND) mice.To achieve absolute quantification of transcript abundance, we performed spike-in calibrated single-cell transcriptomics on GV and MII oocytes from both dietary groups.Furthermore, to investigate the functional role of translation initiation in oocyte gene regulation, we conducted EIF2B1 knockdown in GV-stage oocytes using siRNA-mediated gene silencing. Both EIF2B1-KD oocytes and negative control (NC) oocytes were subjected to spike-in incorporated single-cell transcriptomic analysis to precisely quantify transcriptome changes resulting from translational perturbation.