Nuclear envelope protein LEMD2 in heart

项目总结 编码核膜(NE)成分的基因突变会导致一系列疾病，称为 核包膜病，通常表现为心肌病。去甲肾上腺素将核质从 细胞质，由外核膜和内核膜组成。核层(NL)是一种广泛的 嵌入内核膜(INM)的层蛋白聚合物和相关蛋白质的网络。 在功能上，NL和INM蛋白不仅为细胞核提供机械稳定性，而且还作为 染色质在核周的锚固点，在染色质的组织和 通过与表观遗传机制组件的相互作用和调控来调节基因表达。这个 LAP2-MERIN-MAN1结构域(LEM-D)家族在细胞与细胞之间的相互作用中发挥重要作用。 NL和染色质。LEMD2(LEM Domaining Protein 2，LEM结构域包含蛋白2)是一种位于INM的跨膜蛋白， 参与核完整性、核周系留和异染色质转录沉默。 最近有报道，LEMD2中亮氨酸13的单一氨基酸取代为精氨酸(L13R 导致常染色体隐性遗传性人类心肌病。然而，尽管它与临床相关，但很少有人知道 LEMD2在心肌细胞(CMS)中的特殊作用，以及L13R突变导致 心肌病。为了解决这一知识差距，我们产生了三个小鼠模型：结构性CM- 特异性Lemd2基因敲除(CKO)、可诱导的CM特异性Lemd2基因敲除(IcKO)和LEMD2 L13R敲除 老鼠。我们还将利用LEMD2 L13R的人诱导多能干细胞(IPSC)来源的CM模型 LEMD2中L13R突变对人类CMS的影响。初步研究显示， 胚胎或成年CMS中LEMD2缺失是有害的，LEMD2 L13R突变会影响心脏功能。 小鼠模型，表明LEMD2在维持正常的CM结构和功能中起关键作用 发育中的和成人的心脏。因此，我们假设LEMD2介导的NE完整性和/或 异染色质连接和沉默的调节对CM的结构和功能以及LEMD2是必不可少的 L13R突变损害LEMD2功能的特定方面，导致心肌病。因此，我们的 具体目标是：1.通过分析LEMD2在发育心肌和成年心肌中的作用 结构型(CKO)和诱导型(ICKO)Lemd2 CM特异性基因敲除小鼠心脏形态发生、结构 和功能，以及心肌病的进展；以及2.阐明潜在的分子机制 LEMD2基因L13R敲入详细分析L13R突变所致心肌病 小鼠和人类诱导的多能干细胞(IPSC)衍生的LEMD2 L13R突变CMS。

PROJECT SUMMARY Mutations in genes encoding nuclear envelope (NE) components cause an array of diseases referred to as nuclear envelopathies that often manifest as cardiomyopathies. The NE separates the nucleoplasm from cytoplasm and is composed of outer and inner nuclear membranes. The nuclear lamina (NL) is an extensive network of lamin polymers and associated proteins that are embedded in the inner nuclear membrane (INM). Functionally, NL and INM proteins not only provide mechanical stability to the nucleus but also serve as the anchoring point for chromatin at the nuclear periphery, playing a critical role in chromatin organization and regulation of gene expression via interaction with and modulation of epigenetic machinery components. The LAP2-Emerin-MAN1-domain (LEM-D) family of proteins play an important role in the association between the NL and chromatin. LEMD2 (LEM domain-containing protein 2), is a transmembrane protein located in the INM, involved in nuclear integrity and perinuclear tethering and transcriptional silencing of heterochromatin. Recently, it has been reported that a single amino acid substitution of leucine 13 to arginine (L13R) in LEMD2 leads to autosomal recessive human cardiomyopathy. However, despite its clinical relevance, little is known as to the specific role of LEMD2 in cardiomyocytes (CMs), and mechanisms by which the L13R mutation leads to cardiomyopathy. To address this gap in knowledge, we have generated three mouse models: constitutive CM- specific Lemd2 knockout (cKO), inducible CM-specific Lemd2 knockout (icKO), and LEMD2 L13R knock-in mice. We will also utilize a human induced pluripotent stem cell (iPSC)-derived CM model of the LEMD2 L13R mutation to address the impact of L13R mutation in LEMD2 on human CMs. Preliminary studies revealed that loss of LEMD2 in embryonic or adult CMs is detrimental, and LEMD2 L13R mutation affects cardiac function in a murine model, indicating that LEMD2 plays a critical role in maintaining normal CM structure and function in developing and adult hearts. Thus, we hypothesize that LEMD2-mediated maintenance of NE integrity and/or regulation of heterochromatin tethering and silencing is essential for CM structure and function, and LEMD2 L13R mutation impairs specific aspects of LEMD2 function leading to cardiomyopathy. Accordingly, Our Specific Aims are: 1. To determine the role of LEMD2 in the developing and adult myocardium by analyzing constitutive (cKO) and inducible (icKO) Lemd2 CM-specific knockout mice for heart morphogenesis, structure and function, and the progression of cardiomyopathy; and 2. To elucidate molecular mechanisms underlying cardiomyopathy consequent to the L13R mutation in LEMD2 by detailed analyses of LEMD2 L13R knock-in mice and human induced pluripotent stem cell (iPSC)-derived LEMD2 L13R mutant CMs.