Poldip2: structural and functional implications for vascular disease

项目总结 NADPH氧化酶产生的活性氧物种和炎症与大多数 血管疾病，也通过对正常生理和血管愈合的顺畅调节发挥作用 肌细胞(SMC)增殖和基质分泌。我们实验室最近发现了一种新的p22Phox 结合蛋白，聚合酶三角洲相互作用蛋白-2(Poldip2)，调节NOX4，聚合酶-δ和细胞 循环蛋白，在生长、迁移和胶原蛋白分泌中起重要作用。在这个提案中，我们测试 一种假设，即Poldip2在血管细胞中的多种作用是其结合伙伴的功能，并且 这些功能可以使用小干扰肽进行特定的靶向。在目标1中，我们计划描述 Poldip2和p22Phox之间的物理相互作用，目的是定义可用于 特别是在治疗上破坏这种互动。截断突变体和下拉分析将用于 定义相互作用区域并设计用于细胞和动物递送的多肽。Aim 2致力于使用这些 确定Poldip2调节细胞外胶原水平的机制。我们将测试 假设Poldip2的缺失减少了自噬，从而促进了胶原的分泌。在《目标3》中，我们将 确定平滑肌Poldip2介导的基质调节对动脉僵硬的具体影响 用Poldipflx/Flox动物与平滑肌特异性Cre小鼠杂交在两种情况下形成动脉瘤 慢性和急性环境，以及水凝胶或微粒介导的多肽诱饵输送。 总之，这项提案中描述的实验不仅将扩大我们对血管的理解 Poldip2的生物学，但这将使我们迈出将我们的发现转化为治疗的第一步 可能对治疗血管疾病有用的模式。

PROJECT SUMMARY Reactive oxygen species derived from NADPH oxidases and inflammation have been implicated in most vascular diseases, but also play a role in normal physiology and vascular healing via regulation of smooth muscle cell (SMC) proliferation and matrix secretion. Our laboratory has recently discovered a novel p22phox binding protein, polymerase delta interacting protein-2 (Poldip2), which regulates Nox4, polymerase-δ and cell cycle proteins and has important roles in growth, migration and collagen secretion. In this proposal, we test the hypothesis that the multiple roles of Poldip2 in vascular cells are a function of its binding partners, and that these functions can be specifically targeted using small interfering peptides. In Aim 1, we plan to characterize the physical interaction between Poldip2 and p22phox with the goal of defining peptides that can be used to specifically disrupt this interaction therapeutically. Truncation mutants and pulldown assays will be used to define interacting regions and to design peptides for cell and animal delivery. Aim 2 is devoted to using these peptides to determine the mechanism by which Poldip2 regulates extracellular collagen levels. We will test the hypothesis that depletion of Poldip2 reduces autophagy, thus enhancing collagen secretion. In Aim 3, we will determine the specific consequence of smooth muscle Poldip2-mediated matrix regulation on arterial stiffness and aneurysm formation using Poldipflox/flox animals crossed with smooth muscle-specific Cre mice in both chronic and acute settings, as well as hydrogel- or microparticle-mediated delivery of peptide decoys. Together, the experiments described in this proposal will not only extend our understanding of the vascular biology of Poldip2, but will allow us to take the first step towards translating our findings into a therapeutic modality potentially useful for treating vascular disease.