Department of Surgery, Burn and Shock Trauma Research Institute, Loyola University Chicago Stritch School of Medicine, Maywood, Illinois, United States of America.
Department of Surgery, Morsani College of Medicine, University of South Florida, Tampa, Florida, United States of America.
PLoS One. 2021 Jun 24;16(6):e0253821. doi: 10.1371/journal.pone.0253821. eCollection 2021.
Recently, we reported that the chemokine (C-X-C motif) receptor 4 (CXCR4) and atypical chemokine receptor 3 (ACKR3) heteromerize with α1A/B/D-adrenoceptors (ARs) and arginine vasopressin receptor 1A (AVPR1A) in recombinant systems and in rodent and human vascular smooth muscle cells (hVSMCs). In these studies, we observed that heteromerization between two receptor partners may depend on the presence and the expression levels of other partnering receptors. To test this hypothesis and to gain initial insight into the formation of these receptor heteromers in native cells, we utilized proximity ligation assays in hVSMCs to visualize receptor-receptor proximity and systematically studied how manipulation of the expression levels of individual protomers affect heteromerization patterns among other interacting receptor partners. We confirmed subtype-specific heteromerization between endogenously expressed α1A/B/D-ARs and detected that AVPR1A also heteromerizes with α1A/B/D-ARs. siRNA knockdown of CXCR4 and of ACKR3 resulted in a significant re-arrangement of the heteromerization patterns among α1-AR subtypes. Similarly, siRNA knockdown of AVPR1A significantly increased heteromerization signals for seven of the ten receptor pairs between CXCR4, ACKR3, and α1A/B/D-ARs. Our findings suggest plasticity of seven transmembrane helix (7TM) receptor heteromerization in native cells and could be explained by a supramolecular organization of these receptors within dynamic clusters in the plasma membrane. Because we previously observed that recombinant CXCR4, ACKR3, α1a-AR and AVPR1A form hetero-oligomeric complexes composed of 2-4 different protomers, which show signaling properties distinct from individual protomers, re-arrangements of receptor heteromerization patterns in native cells may contribute to the phenomenon of context-dependent GPCR signaling. Furthermore, these findings advise caution in the interpretation of functional consequences after 7TM receptor knockdown in experimental models. Alterations of the heteromerization patterns among other receptor partners may alter physiological and pathological responses, in particular in more complex systems, such as studies on the function of isolated organs or in in vivo experiments.
最近，我们报道趋化因子（C-X-C 基序）受体 4（CXCR4）和非典型趋化因子受体 3（ACKR3）在重组系统中和在啮齿动物和人血管平滑肌细胞（hVSMCs）中与α1A/B/D-肾上腺素能受体（ARs）和精氨酸加压素受体 1A（AVPR1A）形成异源二聚体。在这些研究中，我们观察到两个受体伴侣之间的异源二聚体形成可能取决于其他伴侣受体的存在和表达水平。为了检验这一假设，并初步了解这些受体在天然细胞中的异源二聚体形成，我们利用 hVSMCs 中的邻近连接分析来可视化受体-受体接近度，并系统地研究了单个原聚体表达水平的操纵如何影响其他相互作用的受体伴侣之间的异源二聚体形成模式。我们证实了内源性表达的α1A/B/D-AR 之间的亚型特异性异源二聚体，并检测到 AVPR1A 也与α1A/B/D-AR 异源二聚体。CXCR4 和 ACKR3 的 siRNA 敲低导致α1-AR 亚型之间的异源二聚体形成模式发生显著重排。同样，AVPR1A 的 siRNA 敲低显著增加了 CXCR4、ACKR3 和α1A/B/D-AR 之间十个受体对中的七个的异源二聚体信号。我们的发现表明，天然细胞中七跨膜螺旋（7TM）受体异源二聚体具有可塑性，这可以通过这些受体在质膜中动态簇内的超分子组织来解释。因为我们之前观察到重组 CXCR4、ACKR3、α1a-AR 和 AVPR1A 形成由 2-4 种不同原聚体组成的异源寡聚复合物，其表现出与单个原聚体不同的信号特性，因此天然细胞中受体异源二聚体形成模式的重排可能导致 GPCR 信号的上下文依赖性现象。此外，这些发现告诫我们在实验模型中进行 7TM 受体敲低后，在解释功能后果时要谨慎。其他受体伴侣之间异源二聚体形成模式的改变可能会改变生理和病理反应，特别是在更复杂的系统中，例如对分离器官功能的研究或在体内实验中。
