Abstract：
Next to its classical role in MHC II-mediated antigen presentation, CD74 was identified as a high-affinity receptor for macrophage migration inhibitory factor (MIF), a pleiotropic cytokine and major determinant of various acute and chronic inflammatory conditions, cardiovascular diseases and cancer. Recent evidence suggests that CD74 is expressed in T cells, but the functional relevance of this observation is poorly understood. Here, we characterized the regulation of CD74 expression and that of the MIF chemokine receptors during activation of human CD4+ T cells and studied links to MIF-induced T-cell migration, function, and COVID-19 disease stage. MIF receptor profiling of resting primary human CD4+ T cells via flow cytometry revealed high surface expression of CXCR4, while CD74, CXCR2 and ACKR3/CXCR7 were not measurably expressed. However, CD4+ T cells constitutively expressed CD74 intracellularly, which upon T-cell activation was significantly upregulated, post-translationally modified by chondroitin sulfate and could be detected on the cell surface, as determined by flow cytometry, Western blot, immunohistochemistry, and re-analysis of available RNA-sequencing and proteomic data sets. Applying 3D-matrix-based live cell-imaging and receptor pathway-specific inhibitors, we determined a causal involvement of CD74 and CXCR4 in MIF-induced CD4+ T-cell migration. Mechanistically, proximity ligation assay visualized CD74/CXCR4 heterocomplexes on activated CD4+ T cells, which were significantly diminished after MIF treatment, pointing towards a MIF-mediated internalization process. Lastly, in a cohort of 30 COVID-19 patients, CD74 surface expression was found to be significantly upregulated on CD4+ and CD8+ T cells in patients with severe compared to patients with only mild disease course. Together, our study characterizes the MIF receptor network in the course of T-cell activation and reveals CD74 as a novel functional MIF receptor and MHC II-independent activation marker of primary human CD4+ T cells.
摘要：
除了其在MHCII介导的抗原呈递中的经典作用，CD74被鉴定为巨噬细胞迁移抑制因子（MIF）的高亲和力受体，多效细胞因子和各种急性和慢性炎症的主要决定因素，心血管疾病和癌症。最近的证据表明CD74在T细胞中表达，但对这种观察的功能相关性了解甚少。这里,我们表征了CD74表达和MIF趋化因子受体在人CD4+T细胞激活过程中的调控，并研究了与MIF诱导的T细胞迁移的联系，函数,和COVID-19疾病阶段。通过流式细胞术对静息的原代人CD4T细胞的MIF受体谱分析显示CXCR4的高表面表达，而CD74，CXCR2和ACKR3/CXCR7没有可测量的表达。然而,CD4+T细胞在细胞内组成型表达CD74，在T细胞激活后，它被显著上调，硫酸软骨素翻译后修饰，可以在细胞表面检测到，通过流式细胞术确定，蛋白质印迹,免疫组织化学，并重新分析可用的RNA测序和蛋白质组数据集。应用基于3D基质的活细胞成像和受体途径特异性抑制剂，我们确定CD74和CXCR4参与MIF诱导的CD4+T细胞迁移。机械上,邻近连接分析显示活化的CD4+T细胞上的CD74/CXCR4杂复，MIF治疗后显著减少，指向MIF介导的内化过程。最后,在30名COVID-19患者的队列中，与仅有轻度病程的患者相比，重度患者的CD4和CD8T细胞上的CD74表面表达被发现显着上调。一起,我们的研究描述了T细胞活化过程中MIF受体网络的特征,并揭示了CD74作为一种新型功能性MIF受体和非MHCII依赖性原代人CD4+T细胞活化标志物.
