RNAseq of Zcchc8 mutant mouse embryonic heads

We identified a germline heterozygous loss-of-function mutation in ZCCHC8, an RNA exosome targeting component, in a family with autosomal dominant pulmonary fibrosis and telomerase RNA insufficiency. To understand the in vivo consequences of Zcchc8 loss, we targeted the gene locus. Zccchc8+/- showed no significant phenotypes but Zcchc8-/- mice showed severe neurodevelopmental defects and died early in adulthood by 60 days. To understand the causes underlying this neurodevelopmental defect, we performed RNAseq on embryonic mouse heads (including the brain). We found no significant changes in gene expression in Zcchc8+/- relative to Zcchc8+/+ embryos. In contrast, Zcchc8-/- mice had a skewed signature marked by upregulation of low abundant transcripts and short, single exon genes including replication-dependent histones. These genes, like telomerase RNA, had extended 3' ends consistent with ZCCHC8 playing a role in both telomerase RNA as well as histone gene post-transcriptional processing. We performed RNAseq on heads (including brains) of embryos with these genotypes: Zcchc8+/+ (n=5), Zcchc8+/- (n=3) and Zcchc8-/- (n=6) that were harvested on day E12.5.本研究在一个伴常染色体显性遗传性肺纤维化与端粒酶RNA功能不全的家系中，鉴定出ZCCHC8基因（一种RNA外泌体靶向组分）存在生殖系杂合功能丧失突变。为探究Zcchc8缺失的体内生物学效应，我们对该基因座实施了靶向编辑。Zcchc8杂合敲除（Zcchc8+/-）小鼠未表现出显著表型，而纯合敲除（Zcchc8-/-）小鼠则出现严重神经发育缺陷，并于成年早期（约60日龄）死亡。为阐明该神经发育缺陷的潜在成因，我们对小鼠胚胎头部（含脑组织）开展了RNA测序（RNAseq）。结果显示，与野生型（Zcchc8+/+）胚胎相比，Zcchc8+/-胚胎的基因表达水平无显著变化。与之形成鲜明对比的是，Zcchc8-/-小鼠的转录组特征发生显著偏移，表现为低丰度转录本以及包括复制依赖性组蛋白在内的短单外显子基因的表达上调。此类基因与端粒酶RNA类似，均存在3'端延伸现象，这与ZCCHC8同时参与端粒酶RNA与组蛋白基因的转录后加工过程的结论相符。我们对胚胎发育第12.5天（E12.5）收获的三种基因型胚胎头部（含脑组织）进行了RNA测序：野生型（Zcchc8+/+，n=5）、杂合敲除型（Zcchc8+/-，n=3）以及纯合敲除型（Zcchc8-/-，n=6）。