Structural and Molecular Mechanisms for Dysregulation of Protein Kinase C Gamma in Cerebellar Ataxia

摘要/摘要 我们提议的研究的总体愿景是理解结构、分子和细胞 蛋白激酶C-γ(PKCG)胚系突变驱动神经细胞凋亡的机制 退行性脊髓小脑性共济失调14(SCA14)。PKCG是一种钙/甘油调节的激酶 仅在神经元中表达，包括浦肯野细胞，其退化是近50个 SCA亚型。我们已经组建了一支拥有广泛和互补的结构专业知识的团队 了解这些突变是如何改变结构和蛋白激酶C机制的 PKCG在疾病表型中的作用。这一提议背后的假设是 突变集中在PKCG的特定区域，这些区域打破自抑制接触以增强其 通过一种新的机制，避免了正常的质量控制降级。这种逃避 降解可能是共济失调突变的一个独特特征，因为与癌症相关的突变 自身抑制接触会破坏PKC的稳定性，并将其分流到降解。这种对正常品质的回避 控制允许异常活跃的PKCG增强其信号输出，这在浦肯野细胞中 小脑会导致变性。此外，我们假设PKCG增强的信号转导可能 通常，SCA的病理基础是很大一部分SCA是由蛋白质突变引起的 控制钙离子动态平衡或信号的物质。我们的目标是将计算、结构、生化、 活细胞成像和磷蛋白组学方法来了解分子细节如何 PKCG影响功能的疾病相关突变，未来的长期目标是利用这一点 治疗这种毁灭性疾病的知识。

Summary/Abstract The overall vision of our proposed research is to understand the structural, molecular, and cellular mechanisms by which germline mutations in protein kinase C gamma (PKCg) drive the neuro- degenerative disease Spinocerebellar Ataxia 14 (SCA14). PKCg is a Ca2+/diacylglycerol-regulated kinase expressed only in neurons, including Purkinje cells whose degeneration is a hallmark of the almost 50 subtypes of SCA. We have assembled a team with extensive and complementary expertise in structural biology of kinases and in PKC mechanisms to understand how these mutations alter the structure and function of PKCg to contribute to the disease phenotype. The hypothesis driving this proposal is that mutations are concentrated at specific regions of PKCg that break autoinhibitory contacts to enhance its activity by a novel mechanism that evades normal quality control degradation. This evasion of degradation may be a unique feature of the Ataxia mutations as cancer-associated mutations that break autoinhibitory contacts destabilize PKC and shunt it to degradation. Such evasion of normal quality control allows aberrantly active PKCg to enhance its signaling output, which in Purkinje cells in the cerebellum leads to degeneration. Furthermore, we hypothesize that enhanced signaling by PKCg may underlie the pathology of SCA, in general, as a large fraction of SCAs are caused by mutations in proteins that control Ca2+ homeostasis or signaling. We aim to combine computational, structural, biochemical, live-cell imaging, and phosphoproteomics approaches to understand the molecular details of how disease-associated mutations in PKCg impact function, with the long-term future goal of using this knowledge to treat this devastating disease.