Impact of a Uty gene trap on the regulation of genes and the abundance of corresponding transcripts in adult mouse somatic (non-reproductive) cells (nucRNA-Seq)

In addition to sperm-related genes, the male-specific chromosome Y (chrY) contains a class of ubiquitously expressed and evolutionary conserved dosage-sensitive regulator genes, which include the neighboring Uty, Ddx3y and (in mice) Eif2s3y genes. However, the impact of targeted mutations of these genes on the functions of adult non-reproductive somatic cells is still unknown due to lack of experimental evidence. We thus compared adult male mice carrying a gene trap within their Uty gene (UtyGT) to wild-type (WT) isogenic controls, and performed deep sequencing of RNA and genome-wide profiling of chromatin in extracts from either cardiac tissue, cardiomyocyte-specific nuclei and purified cardiomyocytes. The impact of UtyGT on gene transcription was mostly limited to its effects on genes surrounding the locus of insertion, i.e. Uty, Ddx3y, long non-coding RNAs (lncRNAs) contained within their introns, Eif2s3y, as well as on the autosomal Malat1 lncRNA. Notwithstanding, UtyGT also caused coordinate changes in the abundance of hundreds of mRNA transcripts related to particular cell functions, including RNA processing and translation. The results altogether indicated that tightly co-regulated chrY genes had nonetheless more widespread effects on the autosomal transcriptome in adult somatic cells, most likely due to mechanisms other than just transcriptional regulation of protein-coding genes. Frozen left ventricles were obtained from either UtyGt(XS0378)Wtsi (which carries a gene trap within the 4th intron of the Uty gene) or their wild-type counterparts from the isogenic strain C57BL/6.Y<129P2> adult mice. For each strain, the procedure was repeated on three different days in order to obtain triplicate samples.除与精子发生相关的基因外，雄性特异性染色体Y（chrY）还包含一类普遍表达且进化保守的剂量敏感性调控基因，其中包括相邻的Uty、Ddx3y以及（小鼠中）Eif2s3y基因。然而，由于缺乏实验证据，这些基因的靶向突变对成年非生殖体细胞功能的影响仍未明确。为此，我们将携带Uty基因内基因陷阱（gene trap）的成年雄性小鼠（UtyGT）与同基因背景的野生型（wild-type, WT）对照小鼠进行对比，并分别对心脏组织、心肌细胞特异性细胞核以及纯化心肌细胞的提取物开展了RNA深度测序与全基因组染色质谱分析。UtyGT对基因转录的影响主要局限于其对插入位点周围基因的调控作用，即Uty、Ddx3y、位于其内含子内的长链非编码RNA（long non-coding RNAs, lncRNAs）、Eif2s3y，以及常染色体Malat1长链非编码RNA。尽管如此，UtyGT还会引发数百个与特定细胞功能（包括RNA加工与翻译）相关的mRNA转录本丰度的协同变化。综合所有结果表明，紧密共调控的chrY基因对成年体细胞的常染色体转录组具有更广泛的影响，这一效应大概率并非仅通过对蛋白编码基因的转录调控实现。实验所用冰冻左心室组织分别来自携带Uty基因第4内含子内基因陷阱的UtyGt(XS0378)Wtsi小鼠，以及同基因背景C57BL/6.Y<129P2>品系的野生型成年小鼠。针对每个品系，我们均重复实验三次以获取生物学重复样本。