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

β-抑制蛋白在β-肾上腺素能受体（β-AR）脱敏和生理环境中的再循环中的功能作用仍然很难理解。在这里，我们证明了小鼠中β-arrestin 2的缺乏（β-arrestin 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反应性降低与Gs或Gi蛋白表达或受体密度或其配体结合特性的改变无关，但伴随着两种bAR亚型磷酸化的显著增加。此外，b2 AR的磷酸化增加与受体与PP 2A的结合显著减少相关，表明b-抑制蛋白对于b2 AR和PP 2之间的物理相互作用是必需的，因此对于受体再敏化是必需的。事实上，通过BARK 1肽抑制剂BARK-ct的腺病毒基因转移抑制BARK 1对b2 AR磷酸化能够拯救b-arrestin 2敲除小鼠肌细胞中b1 AR和b2 AR的功能。这一出乎意料的发现与β-arrestin 2促进β-AR脱敏的既定范式形成鲜明对比，并反驳了β-arrestin 2减少或缺乏应增强β-AR信号传导效率的假设。重要的是，腺病毒基因转移的β-arrestin 2能够完全恢复β-AR介导的收缩反应，从β-arrestin 2基因敲除小鼠的心肌细胞，表明β-AR收缩反应的缺陷是由于缺乏β-arrestin 2，而不是基因敲除相关的非特异性适应性变化。这些结果还表明，β-arrestin 2在β-AR再敏化中起重要作用，但在受体脱敏中不起作用。在自发性高血压大鼠（SHR）的衰竭心脏中，心力衰竭发作前β-arrestin 2的丰度显著降低，并且β-arrestin 2的下调伴随着对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。 这些发现揭示了以前未被认识到的β-arrestin 2在儿茶酚胺介导的收缩支持中的关键作用，以及β-arresin 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新的心力衰竭的致病因素和潜在的治疗靶点。

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.