Transcriptional Metabolic Reprograming Drives Meiotic Fate Decision in Mammalian Germ Cells

Nitrogen starvation drives meiosis in yeasts1, while retinoic acid (RA) is required for mammalian meiosis through the activation of its germline-specific target, stimulated by retinoic acid gene 8 (Stra8).Here, by using single-cell transcriptome analysis of wild-type and Stra8-deficient testes, our data revealed that the expression of major metabolic genes are downregulated during germ cell entry into meiosis. Recently, we collected mouse testicular cells from wild-type and Stra8-deficient mice at postnatal day 12 (P12) and P21 for single cell RNA-sequencing (scRNA-seq). Upon scRNA-seq, we obtained a total of 23,470 cells from these samples. Using stringent quality control, 22,839 cells were selected for further analysis. All cells, including both wild-type and Stra8-deficient testicular cells, were pooled together to perform clustering analysis, which revealed 6 major cell clusters based on their distinct gene expression patterns. We annotated these six clusters by using known marker genes, including spermatogonia (SPG; Stra8; 3,941 cells), spermatocyte (SPC; Syce1; 4,924 cells), spermatid (STD; Acrv1; 710 cells), Sertoli cell (SC; Sox9; 9,488 cells), Myeloid (MYD; Acta2; 3,531 cells), and Leydig Cells (LY; Star; 245 cells). There is an increase in the percentage of spermatogonia in Stra8-deficient testes at both P12 (28% versus 14% in wild-type) and P21 (17% versus 10% in wild-type). At P21, while spermatocytes are abundant and spermatid are formed in wild-type testes, there are only a few spermatocytes (0.2% versus 73% in wild-type), and spermatids are absent in Stra8-deficient testes. Signature genes are enriched for GO terms for distinct biological function and metabolic pathways in different cell types. Single cell RNA sequencing of wild-type and stra8 deficient mouse testis were generated by deep sequencing using the 10x Genomics Chromium Single Cell Gene Expression Solution酵母菌的减数分裂受氮饥饿调控[1]，而视黄酸（retinoic acid, RA）通过激活其生殖系特异性靶基因——视黄酸诱导基因8（stimulated by retinoic acid gene 8, Stra8），是哺乳动物减数分裂过程所必需的。本研究通过对野生型与Stra8敲除小鼠睾丸开展单细胞转录组分析，发现生殖细胞进入减数分裂阶段时，核心代谢基因的表达均出现下调。近期，我们分别于出生后第12天（P12）与第21天（P21）采集野生型及Stra8敲除小鼠的睾丸细胞，进行单细胞RNA测序（single cell RNA-sequencing, scRNA-seq）。经测序后，本研究从这批样本中共获取23470个细胞；经过严格的质量质控，最终筛选出22839个细胞用于后续分析。我们将所有野生型与Stra8敲除睾丸细胞合并后进行聚类分析，依据各细胞独特的基因表达模式，共鉴定出6个主要细胞类群。我们借助已知标记基因对这6个类群完成注释：精原细胞（spermatogonia, SPG；标记基因Stra8；共3941个细胞）、精母细胞（spermatocyte, SPC；标记基因Syce1；共4924个细胞）、精子细胞（spermatid, STD；标记基因Acrv1；共710个细胞）、支持细胞（Sertoli cell, SC；标记基因Sox9；共9488个细胞）、髓系细胞（Myeloid, MYD；标记基因Acta2；共3531个细胞）以及睾丸间质细胞（Leydig Cells, LY；标记基因Star；共245个细胞）。在P12与P21两个时间点，Stra8敲除小鼠睾丸内的精原细胞占比均显著升高：P12时敲除组占比28%，野生型组为14%；P21时敲除组占比17%，野生型组为10%。至P21时，野生型小鼠睾丸内精母细胞丰富且已生成精子细胞；而Stra8敲除小鼠睾丸中仅存在极少量精母细胞（占比0.2%，野生型组为73%），且完全缺失精子细胞。不同细胞类群的特征基因均显著富集于对应生物学功能与代谢通路的基因本体（Gene Ontology, GO）术语中。本研究所涉及的野生型与Stra8敲除小鼠睾丸单细胞RNA测序，均通过10x Genomics Chromium单细胞基因表达解决方案完成深度测序。