神经纤维蛋白SECPH结构域的非典型机制提供了对Sumoylation Sertage选择的新见解
An Atypical Mechanism of SUMOylation of Neurofibromin SecPH Domain Provides New Insights into SUMOylation Site Selection
影响因子:4.50000
分区:生物学2区 / 生化与分子生物学3区
发表日期:2024 Nov 15
作者:
Mohammed Bergoug, Christine Mosrin, Amandine Serrano, Fabienne Godin, Michel Doudeau, Iva Dundović, Stephane Goffinont, Thierry Normand, Marcin J Suskiewicz, Béatrice Vallée, Hélène Bénédetti
摘要
神经纤维蛋白（NF1）是由肿瘤抑制剂基因NF1编码的巨型多域蛋白。 NF1在常见的遗传疾病，神经纤维瘤病I型（NF1）和各种癌症中被突变。该蛋白具有RAS-GAP（GTPase激活蛋白）活性，但也通过其SECPH结构域与不同的信号通路连接，该途径与脂质和不同的蛋白质伴侣相互作用。我们先前表明，NF1与PML核体中的寄生虫细胞性白血病（PML）蛋白部分共定位，这是Sumoylation的热点，从而表明NF1的潜在Sumoylation。在这里，我们证明了NF1的全长同工型2和SECPH片段是SUMO途径的底物，并识别Secph的明确定义的SECPH的semoylation曲线，具有两个主要的修饰赖氨酸。这些位点之一K1731是高度保守的，表面暴露。尽管存在围绕K1731的倒置共识基序，并且在SECPH中存在潜在的SUMO相互作用基序（SIM），但我们表明，对于K1731 Sumoylation而言，这两个元素都不是必不可少的，该元素也与K14上的UBC9 Sumoylation无关。以K1731为中心的SECPH和UBC9之间相互作用的3D模型，结合了定向诱变，确定了K1731 Sumoylation所需的SECPH的特定结构元素，其中一些在报告的NF1病原变体中受到影响。这项工作提供了取决于第三纪，而不是围绕修饰位点的初级蛋白质结构的新示例，从而扩展了我们对Sumoylation位点选择的机制的了解。
Abstract
Neurofibromin (Nf1) is a giant multidomain protein encoded by the tumour-suppressor gene NF1. NF1 is mutated in a common genetic disease, neurofibromatosis type I (NF1), and in various cancers. The protein has a Ras-GAP (GTPase activating protein) activity but is also connected to diverse signalling pathways through its SecPH domain, which interacts with lipids and different protein partners. We previously showed that Nf1 partially colocalized with the ProMyelocytic Leukemia (PML) protein in PML nuclear bodies, hotspots of SUMOylation, thereby suggesting the potential SUMOylation of Nf1. Here, we demonstrate that the full-length isoform 2 and a SecPH fragment of Nf1 are substrates of the SUMO pathway and identify a well-defined SUMOylation profile of SecPH with two main modified lysines. One of these sites, K1731, is highly conserved and surface-exposed. Despite the presence of an inverted SUMO consensus motif surrounding K1731, and a potential SUMO-interacting motif (SIM) within SecPH, we show that neither of these elements is necessary for K1731 SUMOylation, which is also independent of Ubc9 SUMOylation on K14. A 3D model of an interaction between SecPH and Ubc9 centred on K1731, combined with site-directed mutagenesis, identifies specific structural elements of SecPH required for K1731 SUMOylation, some of which are affected in reported NF1 pathogenic variants. This work provides a new example of SUMOylation dependent on the tertiary rather than primary protein structure surrounding the modified site, expanding our knowledge of mechanisms governing SUMOylation site selection.