Beilstein Frauke, Obiang Linda, Raux Hélène, Gaudin Yves
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris-Sud, Gif-sur-Yvette, France
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris-Sud, Gif-sur-Yvette, France.
J Virol. 2015 Nov;89(21):11019-29. doi: 10.1128/JVI.01753-15. Epub 2015 Aug 26.
The matrix protein (M) of vesicular stomatitis virus (VSV) is involved in virus assembly, budding, gene regulation, and cellular pathogenesis. Using a yeast two-hybrid system, the M globular domain was shown to interact with LMP2, a catalytic subunit of the immunoproteasome (which replaces the standard proteasome catalytic subunit PSMB6). The interaction was validated by coimmunoprecipitation of M and LMP2 in VSV-infected cells. The sites of interaction were characterized. A single mutation of M (I96A) which significantly impairs the interaction between M and LMP2 was identified. We also show that M preferentially binds to the inactive precursor of LMP2 (bearing an N-terminal propeptide which is cleaved upon LMP2 maturation). Furthermore, taking advantage of a sequence alignment between LMP2 and its proteasome homolog, PSMB6 (which does not bind to M), we identified a mutation (L45R) in the S1 pocket where the protein substrate binds prior to cleavage and a second one (D17A) of a conserved residue essential for the catalytic activity, resulting in a reduction of the level of binding to M. The combination of both mutations abolishes the interaction. Taken together, our data indicate that M binds to LMP2 before its incorporation into the immunoproteasome. As the immunoproteasome promotes the generation of major histocompatibility complex (MHC) class I-compatible peptides, a feature which favors the recognition and the elimination of infected cells by CD8 T cells, we suggest that M, by interfering with the immunoproteasome assembly, has evolved a mechanism that allows infected cells to escape detection and elimination by the immune system.
The immunoproteasome promotes the generation of MHC class I-compatible peptides, a feature which favors the recognition and the elimination of infected cells by CD8 T cells. Here, we report on the association of vesicular stomatitis virus (VSV) matrix protein (M) with LMP2, one of the immunoproteasome-specific catalytic subunits. M preferentially binds to the LMP2 inactive precursor. The M-binding site on LMP2 is facing inwards in the immunoproteasome and is therefore not accessible to M after its assembly. Hence, M binds to LMP2 before its incorporation into the immunoproteasome. We suggest that VSV M, by interfering with the immunoproteasome assembly, has evolved a mechanism that allows infected cells to escape detection and elimination by the immune system. Modulating this M-induced immunoproteasome impairment might be relevant in order to optimize VSV for oncolytic virotherapy.
水泡性口炎病毒（VSV）的基质蛋白（M）参与病毒组装、出芽、基因调控和细胞发病机制。利用酵母双杂交系统，发现M球状结构域与免疫蛋白酶体的催化亚基LMP2相互作用（LMP2取代了标准蛋白酶体催化亚基PSMB6）。通过在VSV感染细胞中共免疫沉淀M和LMP2验证了这种相互作用。对相互作用位点进行了表征。鉴定出M的一个单突变（I96A），该突变显著损害M与LMP2之间的相互作用。我们还表明，M优先结合LMP2的无活性前体（带有在LMP2成熟时被切割的N端前肽）。此外，利用LMP2与其蛋白酶体同源物PSMB6（不与M结合）之间的序列比对，我们在蛋白质底物切割前结合的S1口袋中鉴定出一个突变（L45R）和一个对催化活性至关重要的保守残基的第二个突变（D17A），导致与M的结合水平降低。这两个突变的组合消除了相互作用。综上所述，我们的数据表明M在LMP2整合到免疫蛋白酶体之前就与其结合。由于免疫蛋白酶体促进主要组织相容性复合体（MHC）I类兼容肽的产生，这一特征有利于CD8 T细胞识别和清除感染细胞，我们认为M通过干扰免疫蛋白酶体组装，进化出了一种机制，使感染细胞能够逃避免疫系统的检测和清除。
免疫蛋白酶体促进MHC I类兼容肽的产生，这一特征有利于CD8 T细胞识别和清除感染细胞。在此，我们报道了水泡性口炎病毒（VSV）基质蛋白（M）与免疫蛋白酶体特异性催化亚基之一LMP2的关联。M优先结合LMP2无活性前体。LMP2上的M结合位点在免疫蛋白酶体中面向内部，因此在其组装后M无法接近。因此，M在LMP2整合到免疫蛋白酶体之前就与其结合。我们认为VSV M通过干扰免疫蛋白酶体组装，进化出了一种机制，使感染细胞能够逃避免疫系统的检测和清除。调节这种由M诱导的免疫蛋白酶体损伤可能与优化VSV用于溶瘤病毒疗法相关。
