RHOA 突变存在于各种癌症类型的不同残基上，这意味着肿瘤发生的突变和细胞特异性机制。在这里，我们重点关注在成人 T 细胞白血病/淋巴瘤中发现的两种功能获得性 RHOA 突变 A161P 和 A161V 的潜在机制。我们发现 RHOAA161P 和 RHOAA161V 都是快速循环突变体，与 RHOAWT 相比，鸟嘌呤核苷酸解离/结合速率增加，并且 GTP 水解活性降低。晶体结构揭示了 RHOAA161P 中改变的核苷酸关联和 RHOAA161V 中的开放核苷酸口袋。如 31P NMR 和分子动力学模拟所示，这两种突变都会扰乱 RHOA 开关区域的动态特性，并改变对 RHOA 活性重要的构象景观。有趣的是，RHOAA161P 和 RHOAA161V 可以与 GDP 结合状态下的效应器相互作用。 1H-15N HSQC NMR 谱支持 RHOAA161V-GDP 中存在活跃群体。突变产生的独特相互作用机制可能有利于类似 RHOAWT 的“ON”构象，从而赋予 GDP 结合状态效应器结合活性。© 2024。作者。

RHOA mutations are found at diverse residues in various cancer types, implying mutation- and cell-specific mechanisms of tumorigenesis. Here, we focus on the underlying mechanisms of two gain-of-function RHOA mutations, A161P and A161V, identified in adult T-cell leukemia/lymphoma. We find that RHOAA161P and RHOAA161V are both fast-cycling mutants with increased guanine nucleotide dissociation/association rates compared with RHOAWT and show reduced GTP-hydrolysis activity. Crystal structures reveal an altered nucleotide association in RHOAA161P and an open nucleotide pocket in RHOAA161V. Both mutations perturb the dynamic properties of RHOA switch regions and shift the conformational landscape important for RHOA activity, as shown by 31P NMR and molecular dynamics simulations. Interestingly, RHOAA161P and RHOAA161V can interact with effectors in the GDP-bound state. 1H-15N HSQC NMR spectra support the existence of an active population in RHOAA161V-GDP. The distinct interaction mechanisms resulting from the mutations likely favor an RHOAWT-like "ON" conformation, endowing GDP-bound state effector binding activity.© 2024. The Author(s).