Defect in hematopoiesis and embryonic lethality at midgestation of Vps13a/Vps13c double knockout mice

VPS13 is the founding member of a family of proteins that mediate lipid transfer at intracellular membrane contact sites by a bridge-like mechanism. Mammalian genomes comprise 4 VPS13 genes encoding proteins with distinct localizations and function. The gene duplication resulting in VPS13A and VPS13C is the most recent in evolution and, accordingly, these two proteins are the most similar to each other. However, they have distinct subcellular localizations and their loss of function mutations in humans are compatible with life but result in two different age-dependent neurodegenerative diseases, chorea-acanthocytosis and Parkinsons disease, respectively. Thus, it remains unclear whether these two proteins have overlapping functions. Here, we show that while Vps13a KO and Vps13c KO mice are viable, embryonic development of Vps13a/Vps13c double knockout (DKO) mice is arrested at midgestation. Prior to death, DKO embryos were smaller than controls, were anemic and had a smaller liver, the key erythropoietic site at this developmental stage. Further analyses of erythroid precursor cells showed that their differentiation was impaired and that this defect was accompanied by activation of innate immunity as revealed by upregulation of interferon stimulated genes (ISGs). Additionally, the RIG-I and MDA5 components of dsRNA triggered innate immunity were found upregulated in the DKO fetal liver. Activation of innate immunity may result from loss of integrity of the membranes of intracellular organelles, such as mitochondria and autophagic lysosomes, due to the absence of these lipid transport proteins. The surprising and striking synthetic effect resulting for the combined loss of VPS13A and VPS13C suggests that despite of the different localization of these two proteins, the lipid fluxes that they mediate are partially redundant. RNAseq profiling of isolated S0 cells (erythroid precursors) from E12.5 WT and Vps13a/Vps13c DKO fetal liversVPS13是一类通过桥接样机制在细胞内膜接触位点介导脂质转运的蛋白质家族的首个成员。哺乳动物基因组包含4个VPS13基因，分别编码具有不同定位与功能的蛋白质。进化过程中最晚发生的基因复制事件产生了VPS13A与VPS13C，因此这两种蛋白质的同源性最高。然而二者的亚细胞定位各不相同，且人类中二者的功能丧失突变虽不致命，但分别会引发两种不同的年龄相关性神经退行性疾病：舞蹈症棘红细胞增多症与帕金森病。因此，目前仍不清楚这两种蛋白质是否存在功能重叠。本研究发现，尽管Vps13a基因敲除（Knockout, KO）与Vps13c基因敲除小鼠均可存活，但Vps13a/Vps13c双基因敲除（Double Knockout, DKO）小鼠的胚胎发育会在妊娠中期停滞。在死亡前，DKO胚胎的体积小于对照组胚胎，且存在贫血症状，肝脏体积也更小——而肝脏是该发育阶段关键的红细胞生成位点。对红系前体细胞的进一步分析显示，其分化过程受到损伤，且该缺陷伴随先天免疫激活，这一点可通过干扰素刺激基因（Interferon Stimulated Genes, ISGs）的上调得到证实。此外，在DKO胚胎的肝脏中，双链RNA（double-stranded RNA, dsRNA）触发先天免疫的关键组分RIG-I与MDA5的表达均出现上调。先天免疫的激活可能源于：由于缺乏这些脂质转运蛋白，细胞内细胞器（如线粒体与自噬溶酶体）的膜完整性遭到破坏。VPS13A与VPS13C联合缺失所产生的出人意料且显著的协同表型提示：尽管二者的亚细胞定位存在差异，但它们所介导的脂质转运通量存在部分功能冗余。对E12.5天野生型（Wild Type, WT）与Vps13a/Vps13c DKO胚胎肝脏中分离得到的S0细胞（红系前体细胞）进行RNA测序（RNA-seq）分析