Cellular Regulation of Dictyostelium Myosin Heavy Chain Kinases

描述（由申请人提供）：本区域更新中提出的研究的总体目标是深入了解控制非肌肉细胞中局部肌球蛋白II细丝组装和收缩的复杂且仍知之甚少的过程。肌球蛋白II双极丝的组装是由肌球蛋白II单体的重链“尾”之间的相互作用引起的。与单体不同，这些纤维可以以导致细胞形状局部变化的方式收缩肌动蛋白纤维。我们建议进一步研究肌球蛋白II依赖的细胞活动（特别是胞质分裂和细胞迁移）可以调节到新确定的候选肌球蛋白II重链激酶（MHCKs）在Dictyosteopathy，并通过扩展，这些过程如何可以在癌细胞表现出不受控制的细胞分裂和转移出错。这里提出的研究也有可能影响我们对其他背景下驱动细胞迁移的潜在过程的理解，如伤口愈合，趋化性和后生动物发育。我们建议研究MHCK-D和AK 1的细胞和酶特性，目的不仅是提供与其他Dictyosteoblastoma α-激酶进行比较的基础，而且还为高等真核系统中的类似研究提供框架。这些研究将由广泛的假设驱动，即由于迄今为止研究的所有网骨藻α-激酶在调节肌球蛋白II中发挥一定作用，并且MHCK-D和AK 1是α-激酶，那么MHCK-D和AK 1也应该调节肌球蛋白II功能。因此，通过我们的研究，我们将解决以下问题：1）MHCK-D是否在细胞内肌球蛋白II丝更新中起作用？2)MHCK-D的定位特性是什么？3)MHCK-D的酶和调节特性是什么？α激酶-1（AK 1）是否在调节肌球蛋白II功能中起作用？总的来说，拟议的研究具有重要意义，因为它们有可能有助于我们了解人肌球蛋白II双极丝转换缺陷如何导致异常血小板形成、肾小球肾炎以及与MYH 9相关疾病相关的其他病理。在更广泛的背景下，我们的MHCK-D和AK 1的研究可能揭示了哺乳动物α-激酶家族成员，瞬时受体电位melastatin 6和7，这两者都涉及在调节肌球蛋白II双极丝周转通过MHC磷酸化在哺乳动物系统中的底物靶向和亚细胞定位的机制。 公共卫生相关性：正常的细胞功能依赖于细胞以高度特异性和受调控的方式改变其形状的能力。我们正在使用一种模式生物，盘基网柄藻，来研究肌球蛋白II的调节，肌球蛋白II是一种通过驱动细胞收缩来促进细胞形状改变的蛋白质。我们研究的首要目标是了解肌球蛋白II调节缺陷如何损害细胞功能，并可能导致疾病状态（如癌症）的发展，或导致伤口愈合缺陷，细胞免疫反应和动脉粥样硬化的发展。

DESCRIPTION (provided by applicant): The overarching goal of the studies proposed in this AREA renewal is to gain insight into the complex, and still poorly understood, processes controlling localized myosin II filament assembly and contraction in a nonmuscle cell context. The assembly of myosin II bipolar filaments results from interactions between the heavy chain "tails" of myosin II monomers. These filaments, unlike monomers, can contract actin filaments in a manner leading to localized changes in cell shape. We propose to examine further how myosin II-dependent cellular activities (specifically, cytokinesis and cellular migration) can be regulated to newly indentified candidate myosin II heavy chain kinases (MHCKs) in Dictyostelium, and by extension, how these processes can go awry in cancer cells exhibiting uncontrolled cell division and metastasis. The studies proposed here also have the potential to impact our understanding of the underlying processes driving cellular migration in other contexts such as wound healing, chemotaxis, and metazoan development. We propose to examine the cellular and enzymatic characteristics of MHCK-D and AK1, with the goal of providing not only a basis for comparison with the other Dictyostelium a- kinases, but also a framework for similar studies in higher eukaryotic systems. These studies will be driven by the broad hypothesis that since all of the Dictyostelium a-kinases studied thus far play some role in regulating myosin II, and MHCK-D and AK1 are a-kinases, then MHCK-D and AK1 should also regulate myosin II function. Thus, with our studies, we will address the following questions: 1) Does MHCK-D function in myosin II filament turnover in the cell? 2) What are the localization properties of MHCK-D? 3) What are the enzymatic and regulatory properties of MHCK-D? and 4) Does alpha kinase- 1 (AK1) play a role in regulating myosin II function? Collectively, the proposed studies are of significance since they have the potential to contribute to our understanding of how defects in human myosin II bipolar filament turnover can lead to abnormal platelet formation, glomerulonephritis, among other pathologies associated with MYH9-related disorders. In a broader context, our studies of MHCK-D and AK1 may shed light on the mechanisms for substrate targeting and subcellular localization for the mammalian a-kinase family members, transient receptor potential melastatins 6 and 7, both of which have been implicated in the regulation of myosin II bipolar filament turnover via MHC phosphorylation in mammalian systems. PUBLIC HEALTH RELEVANCE: Normal cell function relies on the ability of the cell to change its shape in highly specific and regulated ways. We are using a model organism, Dictyostelium discoideum, to study the regulation of myosin II, a protein that facilitates cell shape change by driving contraction of the cell. The overarching goal of our studies is understand how defects in myosin II regulation can compromise cellular function and perhaps contribute to the development of disease states such as cancer, or lead to defects in wound healing, cellular immune responses, and the development of atherosclerosis.