Regulation of Cardiac Myocyte Differentiation

描述（由申请人提供）：拟议的研究具有广泛的重要性和影响，因为心脏发育缺陷可能导致胚胎发育问题，但也可能导致儿童甚至成年生活中出现的问题。控制心肌细胞的分裂、形状和囊泡运输对心脏形态发生至关重要。虽然由单个基因调控这些事件似乎是不可能的，但我们最近发表的研究表明，Lek 1在细胞分裂，形状和运输的调控中起着关键作用。对最初申请的主要批评是，我们缺乏支持心肌细胞中Lek 1功能的数据，并且Lek 1功能的抑制与任何发育缺陷或心脏疾病无关。我们现在证明，1）Lek 1调节心肌细胞中的这些关键功能，2）Lek 1基因的条件性破坏导致心壁发育异常，导致扩张型心肌病。这些新的数据使我们的核心假设，Lek 1在心脏形态发生中起着核心作用，该基因的突变导致分化的心脏功能受损。三个直接的目标将检验这一假设。第一个目标是确定心肌细胞分裂、生长和/或凋亡的变化是否是“小心脏”表型的原因。这将伴随着对出生后心脏的生理分析，以确定肌细胞的正常适应是否被破坏。第二个目标将使用GLUT 4贩运作为模型，量化发育和出生后心脏中囊泡运输中断的程度和时间。这一目标的第二部分更具推测性，但就其分析而言很简单。也就是说，我们将确定如何产生和维持的闰盘是改变Lek 1基因的突变。这将不仅作为囊泡运输和肌细胞形状的模型，而且提供了对心脏功能至关重要的椎间盘调节的关键见解。第三个目标是确定nuc和cyt-LEK 1在心脏发育中的具体作用。这将通过Lek 1-/-心脏的转基因“拯救”来实现。没有其他研究小组拥有阐明这种独特基因在心脏发育和疾病中的功能所需的遗传、分子、免疫化学或细胞生物学工具。

DESCRIPTION (provided by applicant): The proposed studies are of broad importance and impact because developmental defects in the heart can lead to problems in embryogenesis but also problems that are manifested in childhood or even adult life. Control of cardiac myocyte division, shape and vesicular transport is critical for cardiac morphogenesis. While regulation of these events by a single gene may seem improbable, our recently published studies demonstrate that Lek1 plays critical roles in regulation of cell division, shape and transport. The major criticism of the original application was that we lacked data supporting Lek1 function in cardiac myocytes and that inhibition of Lek1 function was not linked to any developmental defect or cardiac disease. We now demonstrate that 1) Lek1 regulates these critical functions in cardiac myocytes and that 2) conditional disruption of the Lek1 gene results in developmental abnormalities of the heart wall leading to dilated cardiomyopathy. These new data lead us to the central hypothesis that Lek1 plays a central role in cardiac morphogenesis and that mutation of this gene leads to impaired function in the differentiated heart. Three straightforward aims will test this hypothesis. The first aim will determine whether changes in cardiac myocyte division, growth and/or apoptosis account for the "small heart" phenotype. This will be accompanied by physiological analyses of the postnatal heart to determine whether the normal adaptation of the myocyte is disrupted. The second aim will quantify the degree and timing of vesicular transport disruption in the developing and postnatal heart using GLUT4 trafficking as a model. The second part of this aim is more speculative but simple in terms of its analysis. Namely, we will determine how generation and maintenance of the intercalated disc is altered with mutation of the Lek1 gene. This will not only serve as a model for vesicular transport and myocyte shape but provide critical insight into regulation of the disc which is essential for cardiac function. The third aim focuses on determining the specific roles of nuc- and cyt-LEK1 in heart development. This will be accomplished using transgenic "rescue" of the Lek1-/- heart. No other group has the genetic, molecular, immunochemical or cell biological tools needed to elucidate the function of this unique gene in cardiac development and disease.