Modulators of cardiomyocyte structure to promote functional recovery during cardiac regeneration and repair

项目摘要 该应用的支配前提是心脏损伤反应福尔斯沿着连续体下降，取决于 物种、发育阶段以及治疗/实验操作。低等生物，如 斑马鱼和新生小鼠在部分截肢或缺血后能够完全心脏再生， 诱导心肌细胞（CM）死亡。再肌化是由增殖的预先存在的 心肌细胞（CM）。另一方面，在对成年哺乳动物的心脏损伤或应激的反应中，CM不能 增殖并相反经历可导致心脏功能障碍的不良肥大性重塑。有 对识别引导CM进行增殖与增殖的分子和信号线索有很大的兴趣。 因此我们可以调节这些因子来促进成年哺乳动物的心脏再生 并在受伤后进行修复。在这个建议中，我们调查的假设，连接和 细胞结构蛋白是调节CM如何响应损伤的关键效应物。调节这些蛋白质 协调CM的增殖状态，而且还促进CM与邻近细胞的适当再整合 在心脏再生过程中。在初步研究中，我们启动了心脏发育筛查， 斑马鱼突变体的再生，破坏已知调节细胞连接和细胞骨架蛋白的因子。 所选择的蛋白质具有已知的与Hippo-Yap信号传导的相互作用，Hippo-Yap信号传导是一种对免疫调节具有重要意义的途径。 促进心脏再生从这个筛选中，我们确定Llgl 1对斑马鱼心脏至关重要 形态发生和发育，以及Shroom 3作为调控CM增殖的关键因子， 斑马鱼心脏再生随后，我们发现CM特异性缺失Llgl 1或Shroom 3， 损害闰盘的完整性，这与未损伤心脏中异常的CM细胞周期活性有关。 在这里，我们建议描绘Llgl 1和Shroom 3在哺乳动物CM中的细胞和生理作用， 发育和新生和成年小鼠心脏损伤后的反应，并进一步研究 这些蛋白质与Hippo-Yap通路组分相互作用的机制。目标1和2采用 分别用CM特异性缺失Llgl 1或Shroom 3的转基因小鼠，以研究这两个基因的作用。 蛋白质在心脏发育和损伤后的反应，重点是细胞形态和 连接复合体和闰盘的建立。我们采用斑马鱼模型来研究 Llgl 1和Shroom 3功能的潜在机制，以及这些因子是否受Hippo 激酶，Lats 1/2。总的来说，我们提出的研究将阐明心脏损伤的关键效应物的作用 反应，能够调节CM细胞结构和细胞-细胞连接，以促进功能性再生 恢复健康的心肌

PROJECT SUMMARY The governing premise of this application is that cardiac injury response falls along a continuum, depending on species, developmental stage, as well as therapeutic/experimental manipulation. Lower organisms such as zebrafish and neonatal mice are capable of complete heart regeneration following partial amputation or ischemic- induced cardiomyocyte (CM) death. Remuscularization is facilitated by proliferation of pre-existing cardiomyocytes (CMs). On the other hand, in response to cardiac injury or stress in adult mammals, CMs fail to proliferate and instead undergo adverse hypertrophic remodeling that can result in cardiac dysfunction. There is great interest in identifying molecules and signaling cues that guide CMs to undergo a proliferative versus hypertrophic response so that we can modulate these factors to promote adult mammalian cardiac regeneration and repair in response to injury. In this proposal we investigate the hypothesis that junctional and cytoarchitectural proteins are key effectors that regulate how CMs respond to injury. Modulation of these proteins coordinate the proliferative state of CMs, but also facilitate proper re-integration of CMs with neighboring cells during cardiac regeneration. In preliminary studies, we initiated a screen for cardiac development and regeneration in zebrafish mutants that disrupt factors known to regulate cell junctional and cytoskeletal proteins. The selected proteins have known interactions with Hippo-Yap signaling, a pathway of significant interest for promoting cardiac regeneration. From this screen, we identified Llgl1 as critical for zebrafish heart morphogenesis and development, and Shroom3 as a factor that is critical for regulation of CM proliferation during zebrafish heart regeneration. Subsequently, we found that CM specific deletion of either Llgl1 or Shroom3 impairs intercalated disc integrity which was associated with aberrant CM cell cycle activity in uninjured hearts. Here, we propose to delineate the cellular and physiological role of Llgl1 and Shroom3 in mammalian CMs during development and the post cardiac injury response in neonatal and adult mice, and to further investigate the mechanism by which these proteins interact with Hippo-Yap pathway components. Aims 1 and 2 employ transgenic mice with CM specific deletion of Llgl1 or Shroom3, respectively, to investigate the role of these two proteins in cardiac development and the post injury response with emphasis on cell morphology and establishment of junctional complexes and intercalated discs. We employ the zebrafish model to investigate the mechanisms underlying Llgl1 and Shroom3 functions, and whether these factors are co-regulated by the Hippo kinase, Lats1/2. Collectively, our proposed studies will elucidate the role of key effectors of the cardiac injury response, capable of modulating CM cytoarchitecture and cell-cell junctions to promote functional regeneration and restore a healthy myocardium.