Laboratoire d'Immunologie Moléculaire, Département de Microbiologie, Infectiologie et Immunologie, Faculté de Médecine, Université de Montréal, Succ Centre-Ville, CP 6128, Montréal, QC, H3C 3J7, Canada.
BMC Immunol. 2021 Aug 12;22(1):56. doi: 10.1186/s12865-021-00444-6.
Invariant chain (CD74, Ii) is a multifunctional protein expressed in antigen presenting cells. It assists the ER exit of various cargos and serves as a receptor for the macrophage migration inhibitory factor. The newly translated Ii chains trimerize, a structural feature that is not readily understood in the context of its MHCII chaperoning function. Two segments of Ii, the luminal C-terminal region (TRIM) and the transmembrane domain (TM), have been shown to participate in the trimerization process but their relative importance and impact on the assembly with MHCII molecules remains debated. Here, we addressed the requirement of these domains in the trimerization of human Ii as well as in the oligomerization with MHCII molecules. We used site-directed mutagenesis to generate series of Ii and DR mutants. These were transiently transfected in HEK293T cells to test their cell surface expression and analyse their interactions by co-immunoprecipitations.
Our results showed that the TRIM domain is not essential for Ii trimerization nor for intracellular trafficking with MHCII molecules. We also gathered evidence that in the absence of TM, TRIM allows the formation of multi-subunit complexes with HLA-DR. Similarly, in the absence of TRIM, Ii can assemble into high-order structures with MHCII molecules.
Altogether, our data show that trimerization of Ii through either TM or TRIM sustains nonameric complex formation with MHCII molecules.
不变链（CD74，Ii）是一种在抗原呈递细胞中表达的多功能蛋白。它协助各种货物从内质网中输出，并作为巨噬细胞迁移抑制因子的受体。新翻译的 Ii 链三聚化，这一结构特征在 MHCII 伴侣功能的背景下不易理解。Ii 的两个片段，即腔内侧 C 末端区域（TRIM）和跨膜结构域（TM），已被证明参与三聚化过程，但它们的相对重要性及其对与 MHCII 分子组装的影响仍存在争议。在这里，我们研究了这些结构域在人 Ii 三聚化以及与 MHCII 分子寡聚化中的要求。我们使用定点突变技术生成了一系列 Ii 和 DR 突变体。将这些突变体瞬时转染到 HEK293T 细胞中，以测试它们的细胞表面表达，并通过共免疫沉淀分析它们的相互作用。
我们的结果表明，TRIM 结构域对于 Ii 三聚化以及与 MHCII 分子的细胞内运输不是必需的。我们还收集了证据表明，在没有 TM 的情况下，TRIM 允许与 HLA-DR 形成多亚基复合物。同样，在没有 TRIM 的情况下，Ii 可以与 MHCII 分子组装成高阶结构。
总的来说，我们的数据表明，通过 TM 或 TRIM 三聚化维持了与 MHCII 分子的九聚体复合物形成。
