Effects of lipidomic diversity on GPCR

整合膜蛋白(IMP)的功能依赖于膜环境，两者都通过活性 特定的脂类作为变构调节剂，通过体膜性质，如粘度、刚性和有序性。近期 脂组分析已经确定了膜复杂性的组成细节，并提供了明确的 证据表明，不同的细胞类型--甚至不同个体中相同的细胞类型--呈现出独特的、 截然不同的膜环境。我们的初步观察表明，这些不同的膜 环境影响G蛋白偶联受体的构象和活性，并可能 调节他们对靶向治疗的反应。此外，我们最近的观察显示，膜 表型对外源干扰非常敏感，这表明饮食或其他外在因素 脂质合成抑制剂可与GPCR靶向治疗药物协同作用。因此，定义 脂质双层对蛋白质的调节机制的光谱，特别是对此有了解 对高价值目标产生影响，如GPCRs。 我们的长期目标是利用膜环境中特定组织的差异来预测和调整 针对GPCRs的治疗的有效性和特异性。这项提议的目的是利用 脂质组学和GPCR重组在连接活细胞膜复杂性和 由模型系统提供的受控环境。通过结合遗传学、细胞生物学、脂质组学、GPCR 重组成蛋白脂质体，和分子模拟，我们的目标是确定通过 膜成分决定腺苷A2a受体的配体结合和信号特征 (A2AR)。我们的中心假设是配体脱落率和G蛋白偶联受局部因素的影响 膜的有序性，这是胆固醇浓度和总脂肪组成的函数。 在整个模型系统中，我们的初步观察表明，胆固醇浓度是一个关键 GPCR活性的调节剂，对配体结合热力学和信号动力学都有影响。我们会 破译胆固醇的影响是由于与受体的特定相互作用还是由于对 通过在模拟、模型中独立地改变和测量膜的物理特性 膜和活细胞。A2AR相关靶细胞类型的脂组分析-神经元，白细胞， 心肌细胞和上皮细胞-将为复杂混合物的分子模拟和重建提供信息 从这些相同的细胞获得的A2AR进入细胞膜将决定信号变化的程度 单元类型。最后，我们将测试脂体扰动的影响--包括胆固醇消耗/ 负荷、脂肪酸摄入以及调节脂肪酸和胆固醇合成的药物--重述A2AR 在模型系统中观察到的功能依赖。这一最终目标将测试基于脂肪组学的原型疗法。 微扰，以局部和具体地加强A2AR治疗。

Integral membrane protein (IMP) function depends on the membrane environment, both through the activity of specific lipids as allosteric modulators and via bulk membrane properties like viscosity, rigidity, and order. Recent lipidomic analyses have established the compositional details of membrane complexity, and provided clear evidence that disparate cell types — and even the same cell type in different individuals — present unique, grossly different membrane environments. Our preliminary observations suggest that these distinct membrane environments influence the conformation and activity of G protein-coupled receptors (GPCRs), and likely modulate their response to targeted therapeutics. Moreover, our recent observations reveal that membrane phenotypes are remarkably susceptible to exogenous perturbations, suggesting that extrinsic factors like diet or lipid synthesis inhibitors could synergize with GPCR-targeted therapeutics. It is therefore critical to define the spectrum of regulatory mechanisms imparted on proteins by lipid bilayers, with this knowledge especially impactful for high value targets, such as GPCRs. Our long-term goal is to exploit tissue-specific differences in membrane environments to predict and tune the efficacy and specificity of therapeutics targeting GPCRs. The objective of this proposal is to leverage advances in lipidomics and GPCR reconstitution to bridge the membrane complexity of live cells with the controlled environments provided by model systems. By combining genetics, cell biology, lipidomics, GPCR reconstitution into proteoliposomes, and molecular simulations we aim to determine the mechanisms by which membrane composition determines ligand binding and signaling characteristics for the adenosine A2A receptor (A2AR). Our central hypothesis is that ligand off-rates and G protein coupling are affected by local membrane order, which is a function of both cholesterol concentration and overall lipid composition. Across model systems, our preliminary observations suggest that cholesterol concentration is a key modulator of GPCR activity, with effects on both ligand binding thermodynamics and signaling kinetics. We will decipher whether the effects of cholesterol are due to specific interactions with the receptor or effects on membrane physical properties by independently varying and measuring these properties in simulations, model membranes, and live cells. Lipidomic analysis of A2AR-relevant target cell types — neurons, leukocytes, cardiomyocytes, and epithelial cells — will inform molecular simulations of complex mixtures, and reconstitution of A2AR into membranes obtained from these same cells will determine the extent of signaling variation across cell types. Finally, we will test whether the effects of lipidomic perturbations — including cholesterol depletion / loading, fatty acid feeding, and drugs that modulate fatty acid and cholesterol synthesis — recapitulate A2AR functional dependence observed in model systems. This final goal will test a prototype therapy based on lipidomic perturbation, to locally and specifically enhance A2AR therapeutics..