ZBTB12-HERVH-LncRNA axis is a molecular barrier for dedifferentiation of human stem cells

Development is generally viewed as one-way traffic of cell state transition from primitive to developmentally advanced states. However, molecular mechanisms that ensure the unidirectional transition remain largely unknown. NanoCAGE sequencing identified an evolutionarily conserved zinc finger repressor, ZBTB12, as a molecular barrier for dedifferentiation of human pluripotent stem cells (hPSCs). Single cell RNA sequencing revealed that ZBTB12 is essential for three germ layer differentiation by blocking hPSC dedifferentiation. Mechanistically, ZBTB12 cooperates with NANOG to fine-tune the expression of human endogenous retroviruses (HERVs). Active HERVH loci drive the expression of overlapping long non-coding RNAs (lncRNAs) whose downregulation by ZBTB12 is necessary for dedifferentiation blockage and pluripotency exit. Overall, we have identified a ZBTB12-HERVH-lncRNA axis as molecular machinery that safeguards the unidirectional transition of stem cell fates. Single cell transcriptomic profiling from hESCs expressing control or ZBTB12 shRNA before and after spontaneous differentiation (8 days), using 10X genomics Chromium Next GEM system

细胞状态转变通常被认为是从原始状态向发育成熟状态单向进行的过程。然而，维持这种单向转变的分子机制在很大程度上仍未明确。纳米CAGE测序（NanoCAGE sequencing）将一个进化保守的锌指抑制因子ZBTB12鉴定为人类多能干细胞（human pluripotent stem cells, hPSCs）去分化的分子屏障。单细胞RNA测序（single cell RNA sequencing）结果显示，ZBTB12通过阻断hPSCs的去分化过程，对三胚层分化至关重要。从机制上来说，ZBTB12与NANOG协同作用，精准调控人类内源性逆转录病毒（human endogenous retroviruses, HERVs）的表达。活化的HERVH位点可驱动与其重叠的长链非编码RNA（long non-coding RNAs, lncRNAs）的表达；ZBTB12对这些lncRNAs的下调，是阻断去分化并退出多能性状态的必要条件。综上，本研究鉴定出ZBTB12-HERVH-lncRNA调控轴作为维持干细胞命运单向转变的分子机制。本研究采用10X Genomics Chromium Next GEM系统，对自发分化（8天）前后表达对照或ZBTB12短发卡RNA（short hairpin RNA, shRNA）的人类胚胎干细胞（human embryonic stem cells, hESCs）开展单细胞转录组图谱分析。