CD4+T细胞活化是由五模块受体复合物驱动的。T细胞受体(TCR)是结合嵌入MHCII分子(pMHCII)内的肽抗原的复合表面的受体模块。它与三个信号模块(CD3γε,CD3δε,和CD3ζ)形成TCR-CD3复合物。CD4是共受体模块。它与CD4+T细胞外部的TCR-CD3-pMHCII组装体和Src激酶相互关联,LCK,在里面。以前,我们报道了在eutherian(胎盘哺乳动物)CD4中发现的CD4跨膜GGXXG和细胞质近膜(C/F)CVC基序具有在纯化选择下进化的组成残基(Lee等人。,2022年)。在T细胞杂交瘤中一起表达这些基序的突变体增加了CD4-LCK关联,但降低了CD3ζ,ZAP70和PLCγ1磷酸化水平,以及IL-2的生产,响应于激动剂pMHCII。因为这些突变体优先将CD4-LCK对定位到非移植物膜部分,我们的结果的一个解释是,他们通过隔离LCK远离TCR-CD3而损害了近端信号传导.另一种假设是突变直接影响信号传导,因为基序通常在信号传导中起LCK-非依赖性作用。这项研究的目的是区分这些可能性。利用T细胞杂交瘤,我们的结果表明:pMHCII特异性信号启动不需要细胞内CD4-LCK相互作用;GGXXG和(C/F)CV+C基序是CD4介导的pMHCII特异性信号扩增的关键决定因素;GGXXG和(C/F)CV+C基序独立于直接CD4-LCK关联而发挥其功能.这些数据为为什么这些基序中的残基在下颌脊椎动物中处于纯化选择中提供了机制解释。结果对于合成受体的仿生工程也很重要。
CD4+ T cell activation is driven by five-module receptor complexes. The T cell receptor (TCR) is the receptor module that binds composite surfaces of peptide antigens embedded within MHCII molecules (pMHCII). It associates with three signaling modules (CD3γε, CD3δε, and CD3ζζ) to form TCR-CD3 complexes. CD4 is the coreceptor module. It reciprocally associates with TCR-CD3-pMHCII assemblies on the outside of a CD4+ T cells and with the Src kinase,
LCK, on the inside. Previously, we reported that the CD4 transmembrane GGXXG and cytoplasmic juxtamembrane (C/F)CV+C motifs found in eutherian (placental mammal) CD4 have constituent residues that evolved under purifying selection (Lee et al., 2022). Expressing mutants of these motifs together in T cell hybridomas increased CD4-
LCK association but reduced CD3ζ, ZAP70, and PLCγ1 phosphorylation levels, as well as IL-2 production, in response to agonist pMHCII. Because these mutants preferentially localized CD4-
LCK pairs to non-raft membrane fractions, one explanation for our results was that they impaired proximal signaling by sequestering
LCK away from TCR-CD3. An alternative hypothesis is that the mutations directly impacted signaling because the motifs normally play an LCK-independent role in signaling. The goal of this study was to discriminate between these possibilities. Using T cell hybridomas, our results indicate that: intracellular CD4-
LCK interactions are not necessary for pMHCII-specific signal initiation; the GGXXG and (C/F)CV+C motifs are key determinants of CD4-mediated pMHCII-specific signal amplification; the GGXXG and (C/F)CV+C motifs exert their functions independently of direct CD4-LCK association. These data provide a mechanistic explanation for why residues within these motifs are under purifying selection in jawed vertebrates. The results are also important to consider for biomimetic engineering of synthetic receptors.