B-Arrestin2 Is Required for BAR Resensitization But Not its Desensitization

在生理环境中，β-阻滞素在β-肾上腺素能受体(β-AR)脱敏和再循环中的作用在很大程度上仍不清楚。在这里，我们证明了小鼠缺乏β-拦阻蛋白2(β-拦阻蛋白2 KO)会导致对两种β-AR亚型刺激的心脏收缩反应的缺陷。β-AR反应性的降低与Gs或Gi蛋白的表达、受体密度或其配体结合特性的变化无关，但伴随着这两种BAR亚型磷酸化的显著增加。此外，B2AR的磷酸化增加与受体与PP2A的结合显著减少相关，这表明b-arrestins对于B2AR和PP2之间的物理相互作用是必要的，从而对受体的再增敏是必要的。事实上，通过腺病毒转移BARK1多肽抑制剂Bark-ct来抑制BARK1对B2AR磷酸化的抑制，能够从b-arrestin2基因敲除小鼠的心肌细胞中挽救b1AR和B2AR的功能。这一出人意料的发现与已建立的研究范式形成鲜明对比，即β-arrestin2促进β-AR脱敏，并反对β-arrestin2的减少或缺失应该提高β-AR信号转导效率的假设。重要的是，腺病毒转导β-Arrstin2能够完全恢复β-AR介导的心肌细胞收缩反应，提示β-AR收缩反应的缺陷是由于β-arrestin2的缺陷，而不是基因敲除引起的非特异性适应性改变。这些结果还表明，β-arrestin2在β-AR再增敏中起重要作用，但在受体脱敏中不起作用。在自发性高血压大鼠(SHR)衰竭的心脏中，β-arrestin2的丰度在心力衰竭发作前显著降低，并且β-arrestin2的下调伴随着对任何一种β-AR亚型刺激的明显减弱的心脏收缩反应，这一事实证实了这一结论。这些发现揭示了β-抑制素2在儿茶酚胺介导的收缩支持中先前未被认识到的关键作用，以及β-精氨酸2下调与心力衰竭发展之间的潜在因果关系。因此，我们的研究可能揭示心力衰竭的新的病因和潜在的治疗靶点。

The functional role of beta-arrestins in beta-adrenergic receptor (beta-AR) desensitization and recycling in physiological contexts remains largely elusive. Here, we demonstrate that deficiency of beta-arrestin2 in mice (beta-arrestin2 KO) causes defects in cardiac contractile response to stimulation of both beta-AR subtypes. The reduced beta-AR responsiveness is not associated with alterations in the expression of Gs or Gi proteins or the receptor density or its ligand binding properties, but is accompanied by a marked increase in phosphorylation of both bAR subtypes. Furthermore, the increased phosphorylation of b2AR is associated with markedly reduced binding of the receptor to PP2A, indicating that b-arrestins is necessary for the physical interaction between b2AR and PP2, thus for the receptor resensitization. Indeed, inhbition of b2AR phosphorylation by BARK1 via adenoviral gene transfer of a BARK1 peptide inhibitor, BARK-ct, is able to rescue the function of both b1AR and b2AR in myocytes from b-arrestin2 knockout mice. This unexpected finding is in sharp contrast to the established paradigm that beta-arrestin2 promotes beta-AR desensitization, and argues against the hypothesis that reduction or deficiency of beta-arrestin2 should enhance beta-AR signaling efficiency. Importantly, adenoviral gene transfer of beta-arrstin2 is able to fully restore beta-AR mediated contractile response in cardiomyocytes from beta-arrestin2 KO mice, indicating that the defect of beta-AR contractile response is attributable to the deficiency of beta-arrestin2 rather that the gene knockout associated nonspecific adaptive changes. These results also suggest that beta-arrestin2 plays an essential role in beta-AR resensitiztion, but not in the receptor desensitization. This conclusion is corroborated by the fact that in the failing hearts from spontaneous hypertensive rats (SHR), the abundance of beta-arrestin2 is markedly decreased before the onset of heart failure, and that the downregulation of beta-arrestin2 is accompanied by overtly attenuated cardiac contractile response to either beta-AR subtype stimulation. These findings have revealed a previously unrecognized crucial role of beta-arrestin2 in catecholamine-mediated contractile support and a potential causal relation between downregulation of beta-arresin2 and the development of heart failure. Thus, our study might reveals novel causal factors and potential therapeutic targets of heart failure.