Folding, Misfolding, and Function of PMP22

 描述(申请人提供)：PMP22的折叠、错误折叠和功能外周髓鞘蛋白22(PMP22)是一种四聚糖完整的膜蛋白，在周围神经系统(PNS)的髓鞘中含量丰富。编码PMP22氨基酸序列变化的基因突变是常见的人类周围神经病Charcot-Marie-Tooth病(CMTD)的原因之一。CMTD突变促进PMP22在分泌途径早期的错误折叠、保留和可能的降解，导致功能丧失。这会导致PNS轴突的髓鞘障碍和CMTD的发病。尽管PMP22显然很重要，但它在髓鞘和雪旺细胞生理学中的作用还不是很清楚。我们最近对一系列CMTD突变形式的PMP22的热力学折叠稳定性进行了量化，并发现PMP22的热力学稳定性与细胞表面转运效率和患者神经传导速度大致呈线性相关。在目标1中，我们将比较和对比不同CMTD突变体PMP22的折叠动力学和其他性质，以检验引发PMP22不稳定和细胞内末端错误折叠的特定缺陷是否在突变体之间是相同的，或者除了稳定性的差异之外，不同的CMTD突变体是否具有明显不同的缺陷。在目标2中，我们将确定PMP22的CMTD突变形式的不稳定性是如何通过内质网质量控制来感知的。来自这一目标的细胞数据将与目标1的生物物理实验结果相结合，以提供关于PMP22中的缺陷如何被ER质量控制识别的统一的生物物理和细胞生物学理解。在目标3中，我们将确定纯化的PMP22在重组为脂泡后如何促进髓磷样集合体(MLA)的形成。这将探索这样的假设，即PMP22在MLA形成中的作用与其至少一个固有功能密切相关，并且至少一些CMTD突变可能以一种反映突变如何促进PNS组织中的髓鞘形成的方式来干扰MLA的形成。虽然CMTD是一种相对常见的疾病(1：3000)，但据我们所知，这是 世界，包括PMP22的生物物理特征。上述目标的完成将极大地促进我们对这种髓膜蛋白在健康和疾病条件下的折叠、错误折叠和功能的理解。这一结果将直接揭示CMTD的分子机制，并可能为未来抗CMTD疗法的发展铺平道路。

 DESCRIPTION (provided by applicant): Folding, Misfolding, and Function of PMP22 Peripheral myelin protein 22 (PMP22) is a tetraspan integral membrane protein that is abundant in myelin of the peripheral nervous system (PNS). Genetic mutations that encode changes in the amino acid sequence of PMP22 are one cause of the common human peripheral neuropathy Charcot-Marie-Tooth Disease (CMTD). CMTD mutations promote misfolding, retention, and possible degradation of PMP22 early in the secretory pathway, leading to loss of function. This results in dysmyelination of PNS axons and the onset of CMTD. While clearly important, the function of PMP22 in myelin and Schwann cell physiology is not well understood. We recently quantitated the thermodynamic folding stabilities of a series of CMTD mutant forms of PMP22 and established that there is a roughly linear correlation between the thermodynamic stability of PMP22 and both cell surface trafficking efficiencies and patient nerve conduction velocities. In Aim 1 we will compare and contrast the folding kinetics and other properties of different CMTD mutant forms of PMP22 in order to examine whether the specific defects that trigger PMP22 instability and terminal misfolding in the cell are the same from mutant to mutant or whether different CMTD mutants have distinctly different defects beyond differences in stability. In Aim 2 we will determine how the instability of CMTD mutant forms of PMP22 is sensed by endoplasmic reticulum quality control. The cellular data from this aim will be integrated with the results of the biophysical experiments of Aim 1 to provide a unified biophysical and cell biological understanding of how defects in PMP22 are recognized by ER quality control. In Aim 3 we will determine how purified PMP22 promotes formation of myelin-like assemblies (MLAs) following reconstitution into lipid vesicles. This will explore the hypotheses that the role of PMP22 in MLA formation is closely related to at least one of its native functions and also that at least some CMTD mutants are likely to disrupt MLA formation in a manner that reflects how mutations promote dysmyelination in PNS tissue. While CMTD is a relatively common disorder (1:3000), to the best of our knowledge this is the only project in the world that includes the biophysical characterization of PMP22. Completion of the above aims will significantly advance our understanding of the folding, misfolding, and function of this myeli membrane protein under conditions of both health and disease. Results will provide direct insight into the molecular mechanisms underlying CMTD and may pave the way for future development of anti-CMTD therapeutics.