Downregulated lncRNA RCPCD promotes differentiation of embryonic stem cells into cardiac pacemaker-like cells by suppressing HCN4 promoter methylation.
Downregulated lncRNA RCPCD promotes differentiation of embryonic stem cells into cardiac pacemaker-like cells by suppressing HCN4 promoter methylation.
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下调lncRNA RCPCD通过抑制HCN4启动子甲基化促进胚胎干细胞分化为心脏起搏器样细胞
DOI：
10.1038/s41419-021-03949-5
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发表时间：
2021-07-02
影响因子：
9
通讯作者：
Gu X
中科院分区：
文献类型：
--
作者：
Zhu Y;You J;Wei W;Gu J;Xu C;Gu X
长非编码RNA(LncRNA)在胚胎干细胞(ESCs)研究中受到越来越多的关注。然而，lncRNA在胚胎干细胞分化为起搏样细胞中的作用尚不清楚。因此，本研究旨在探讨lncRNA在胚胎干细胞向起搏样细胞分化过程中的作用和机制。培养胚胎干细胞，并诱导其向起搏样细胞分化。RNA测序鉴定胚胎干细胞分化为起搏样细胞过程中的差异表达。采用细胞形态观察、流式细胞术、实时定量聚合酶链式反应、免疫印迹和免疫荧光等方法检测胚胎干细胞向起搏样细胞分化的情况。LncRNA和基因通过转染腺病毒在分化过程中过度表达或被敲除。荧光原位杂交(FISH)检测分化的ESCs中的lncRNA定位。采用荧光素酶报告基因分析、甲基化特异性聚合酶链式反应、染色质免疫共沉淀和RNA免疫共沉淀等方法研究了lncRNA调控HCN4基因表达的机制。挽救实验证实，lncRNA通过HCN4调节ESCs向起搏样细胞的分化。我们成功地培养了胚胎干细胞，并将其诱导分化为起搏样细胞。在胚胎干细胞向起搏样细胞分化的过程中，lncRNA RCPCD的表达显著降低。RCPCD过表达抑制了ESCs向起搏样细胞的分化。RCPCD通过通过DNMT1、DNMT2和DNMT3增加启动子区域的HCN4甲基化来抑制HCN4的表达。RCPCD通过抑制HCN4的表达抑制ESCs向起搏样细胞的分化。我们的结果证实了lncRNA RCPCD在胚胎干细胞分化为起搏器样细胞过程中的作用和机制，为基于细胞的生物起搏器的发展奠定了基础。
Long non-coding RNA (lncRNA) is receiving increasing attention in embryonic stem cells (ESCs) research. However, the roles of lncRNA in the differentiation of ESCs into pacemaker-like cells are still unclear. Therefore, the present study aims to explore the roles and mechanisms of lncRNA in the differentiation of ESCs into pacemaker-like cells. ESCs were cultured and induced differentiation to pacemaker-like cells. RNA sequencing was used to identify the differential expression lncRNAs during the differentiation of ESCs into pacemaker-like cells. Cell morphology observation, flow cytometry, quantitative real-time polymerase chain reaction, western blot, and immunofluorescence were used to detect the differentiation of ESCs into pacemaker-like cells. LncRNA and genes overexpression or knockdown through transfected adenovirus in the differentiation process. The fluorescence in situ hybridization (FISH) detected the lncRNA location in the differentiated ESCs. Luciferase reporter gene assay, methylation-specific PCR, chromatin immunoprecipitation assay, and RNA immunoprecipitation assay were performed to reveal the mechanism of lncRNA-regulating HCN4 expression. Rescue experiments were used to confirm that lncRNA regulates the differentiation of ESCs into pacemaker-like cells through HCN4. We cultured the ESCs and induced the differentiation of ESCs into pacemaker-like cells successfully. The expression of lncRNA RCPCD was significantly decreased in the differentiation of ESCs into pacemaker-like cells. Overexpression of RCPCD inhibited the differentiation of ESCs into pacemaker-like cells. RCPCD inhibited the expression of HCN4 by increasing HCN4 methylation at the promoter region through DNMT1, DNMT2, and DNMT3. RCPCD inhibited the differentiation of ESCs into pacemaker-like cells by inhibiting the expression of HCN4. Our results confirm the roles and mechanism of lncRNA RCPCD in the differentiation of ESCs into pacemaker-like cells, which could pave the path for the development of a cell-based biological pacemaker.
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影响因子：
7.5
作者：
Schweizer PA;Darche FF;Ullrich ND;Geschwill P;Greber B;Rivinius R;Seyler C;Müller-Decker K;Draguhn A;Utikal J;Koenen M;Katus HA;Thomas D
通讯作者：
Thomas D
影响因子：
7.5
作者：
Liu, Zheying;Liu, Liya;Han, Liping
通讯作者：
Han, Liping
HCN2 和 TBX3 将人类诱导的多能干细胞来源的心肌细胞重新编程为起搏器样细胞
影响因子：
3.1
作者：
Zhao, Hongyi;Wang, Fengyuan;Huang, Congxin
通讯作者：
Huang, Congxin
影响因子：
20.1
作者：
D'Souza A;Pearman CM;Wang Y;Nakao S;Logantha SJRJ;Cox C;Bennett H;Zhang Y;Johnsen AB;Linscheid N;Poulsen PC;Elliott J;Coulson J;McPhee J;Robertson A;da Costa Martins PA;Kitmitto A;Wisløff U;Cartwright EJ;Monfredi O;Lundby A;Dobrzynski H;Oceandy D;Morris GM;Boyett MR
通讯作者：
Boyett MR
影响因子：
5.9
作者：
Ionta, Vittoria;Liang, Wenbin;Kim, Elizabeth H.;Rafie, Reza;Giacomello, Alessandro;Marban, Eduardo;Cho, Hee Cheol
通讯作者：
Cho, Hee Cheol