Lee UnJin, Li Cong, Langer Christopher B, Svetec Nicolas, Zhao Li
Laboratory of Evolutionary Genetics and Genomics, The Rockefeller University, New York, NY, USA.
bioRxiv. 2025 Mar 25:2024.04.29.591771. doi: 10.1101/2024.04.29.591771.
Spermatogenesis is a key developmental process underlying the origination of newly evolved genes. However, rapid cell type-specific transcriptomic divergence of the germline has posed a significant technical barrier for comparative single-cell RNA-sequencing (scRNA-Seq) studies. By quantifying a surprisingly strong correlation between species- and cell type-specific divergence in three closely related species, we apply a new statistical procedure to identify a core set of 198 genes that are highly predictive of cell type identity while remaining robust to species-specific differences that span over 25-30 million years of evolution. We then utilize cell type classifications based on the 198-gene set to show how transcriptional divergence in cell type increases throughout spermatogenic developmental time. After validating these cross-species cell type classifications using RNA fluorescence in situ hybridization (FISH) and imaging, we then investigate the influence of genome organization on the molecular evolution of spermatogenesis vis-a-vis transcriptional bursting. We first show altering transcriptional burst size contributes to pre-meiotic transcription and altering bursting frequency contributes to post-meiotic expression. We then report global differences in autosomal vs. X chromosomal transcription may arise in a developmental stage preceding full testis organogenesis by showing evolutionarily conserved decreases in X-linked transcription bursting kinetics in all examined somatic and germline cell types. Finally, we provide evidence supporting the cultivator model of gene origination by demonstrating how the appearance of newly evolved testis-specific transcripts potentially provides short-range regulation of neighboring genes' transcriptional bursting properties during key stages of spermatogenesis.
精子发生是新进化基因起源的关键发育过程。然而，生殖系快速的细胞类型特异性转录组差异给比较单细胞RNA测序（scRNA-Seq）研究带来了重大技术障碍。通过量化三个密切相关物种中物种特异性和细胞类型特异性差异之间惊人的强相关性，我们应用一种新的统计程序来鉴定一组198个基因的核心集，这些基因对细胞类型身份具有高度预测性，同时对跨越2500万至3000万年进化的物种特异性差异具有稳健性。然后，我们利用基于198个基因集的细胞类型分类来展示细胞类型中的转录差异如何在整个精子发生发育过程中增加。在使用RNA荧光原位杂交（FISH）和成像验证这些跨物种细胞类型分类后，我们接着研究基因组组织对精子发生分子进化相对于转录爆发的影响。我们首先表明，改变转录爆发大小有助于减数分裂前的转录，而改变爆发频率有助于减数分裂后的表达。然后，我们通过展示在所有检查的体细胞和生殖细胞类型中X连锁转录爆发动力学在进化上保守的下降，报告常染色体与X染色体转录的全局差异可能在完全睾丸器官发生之前的发育阶段出现。最后，我们通过证明新进化的睾丸特异性转录本的出现如何在精子发生的关键阶段潜在地提供对邻近基因转录爆发特性的短程调节，提供了支持基因起源培育者模型的证据。
