Divergent Folding-Mediated Epistasis Among Unstable Membrane Protein Variants.
Divergent Folding-Mediated Epistasis Among Unstable Membrane Protein Variants.
复制标题
不稳定膜蛋白变体中不同折叠介导的上位性。
DOI：
10.1101/2023.08.25.554866
复制
发表时间：
2024
期刊：
影响因子：
--
通讯作者：
Schlebach,JonathanP
中科院分区：
文献类型：
--
作者：
Chamness,LauraM;Kuntz,CharlesP;McKee,AndrewG;Penn,WesleyD;Hemmerich,ChristopherM;Rusch,DouglasB;Woods,Hope;Dyotima;Meiler,Jens;Schlebach,JonathanP
许多膜蛋白易于错误折叠，这损害了它们在质膜上的功能表达。哺乳动物促性腺激素释放激素受体（GnRHR）尤其如此。我们最近证明，进化GnRHR修改似乎已经符合适应性变化的共翻译折叠效率。虽然蛋白质的稳定性是已知的形状进化，目前还不清楚如何共翻译折叠的限制调节突变之间的协同作用，上位相互作用。因此，我们比较了由破坏GnRHR的膜拓扑结构（V276T）或三级结构（W107A）的突变形成的成对相互作用。使用深度突变扫描，我们评估了这些变体的质膜表达如何被数百个次级突变修饰。在三个遗传背景的251个突变体的分析表明，V276T和W107A形成不同的上位相互作用，这取决于严重程度和机制的不稳定。V276T主要与可溶性环中的不稳定突变形成负上位相互作用。相比之下，W107 A与环和跨膜结构域中的突变形成正相互作用，这反映了已经不稳定的变体中不稳定突变的影响逐渐减小。这些发现揭示了上位性是如何通过膜蛋白和更普遍的不稳定蛋白的构象缺陷来重塑的。
Many membrane proteins are prone to misfolding, which compromises their functional expression at the plasma membrane. This is particularly true for the mammalian gonadotropin-releasing hormone receptor GPCRs (GnRHR). We recently demonstrated that evolutionary GnRHR modifications appear to have coincided with adaptive changes in cotranslational folding efficiency. Though protein stability is known to shape evolution, it is unclear how cotranslational folding constraints modulate the synergistic, epistatic interactions between mutations. We therefore compared the pairwise interactions formed by mutations that disrupt the membrane topology (V276T) or tertiary structure (W107A) of GnRHR. Using deep mutational scanning, we evaluated how the plasma membrane expression of these variants is modified by hundreds of secondary mutations. An analysis of 251 mutants in three genetic backgrounds reveals that V276T and W107A form distinct epistatic interactions that depend on both the severity and the mechanism of destabilization. V276T forms predominantly negative epistatic interactions with destabilizing mutations in soluble loops. In contrast, W107A forms positive interactions with mutations in both loops and transmembrane domains that reflect the diminishing impacts of the destabilizing mutations in variants that are already unstable. These findings reveal how epistasis is remodeled by conformational defects in membrane proteins and in unstable proteins more generally.