Wang Xinhua, Zeng Weizhong, Kim Min Seuk, Allen Patrick B, Greengard Paul, Muallem Shmuel
Department of Physiology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390-9040, USA.
EMBO J. 2007 Jun 6;26(11):2768-76. doi: 10.1038/sj.emboj.7601701. Epub 2007 Apr 26.
Spinophilin (SPL) and neurabin (NRB) are structurally similar scaffolding proteins with several protein binding modules, including actin and PP1 binding motifs and PDZ and coiled-coil domains. SPL also binds regulators of G protein signaling (RGS) proteins and the third intracellular loop (3iL) of G protein-coupled receptors (GPCRs) to reduce the intensity of Ca(2+) signaling by GPCRs. The role of NRB in Ca(2+) signaling is not known. In the present work, we used biochemical and functional assays in model systems and in SPL(-/-) and NRB(-/-) mice to show that SPL and NRB reciprocally regulate Ca(2+) signaling by GPCRs. Thus, SPL and NRB bind all members of the R4 subfamily of RGS proteins tested (RGS1, RGS2, RGS4, RGS16) and GAIP. By contract, SPL, but not NRB, binds the 3iL of the GPCRs alpha(1B)-adrenergic (alpha(1B)AR), dopamine, CCKA, CCKB and the muscarinic M3 receptors. Coexpression of SPL or NRB with the alpha(1B)AR in Xenopus oocytes revealed that SPL reduces, whereas NRB increases, the intensity of Ca(2+) signaling by alpha(1B)AR. Accordingly, deletion of SPL in mice enhanced binding of RGS2 to NRB and Ca(2+) signaling by alphaAR, whereas deletion of NRB enhanced binding of RGS2 to SPL and reduced Ca(2+) signaling by alphaAR. This was due to reciprocal modulation by SPL and NRB of the potency of RGS2 to inhibit Ca(2+) signaling by alphaAR. These findings suggest a novel mechanism of regulation of GPCR-mediated Ca(2+) signaling in which SPL/NRB forms a functional pair of opposing regulators that modulates Ca(2+) signaling intensity by GPCRs by determining the extent of inhibition by the R4 family of RGS proteins.
亲嗜素（SPL）和神经肌动蛋白（NRB）是结构相似的支架蛋白，具有多个蛋白质结合模块，包括肌动蛋白和蛋白磷酸酶1（PP1）结合基序以及PDZ和卷曲螺旋结构域。SPL还与G蛋白信号调节（RGS）蛋白以及G蛋白偶联受体（GPCR）的第三个细胞内环（3iL）结合，以降低GPCR介导的Ca（2+）信号强度。NRB在Ca（2+）信号传导中的作用尚不清楚。在本研究中，我们在模型系统以及SPL（-/-）和NRB（-/-）小鼠中使用生化和功能分析，以表明SPL和NRB相互调节GPCR介导的Ca（2+）信号传导。因此，SPL和NRB与所测试的RGS蛋白R4亚家族的所有成员（RGS1、RGS2、RGS4、RGS16）和GAIP结合。相比之下，SPL而非NRB与GPCRα（1B）-肾上腺素能受体（α（1B）AR）、多巴胺、胆囊收缩素A（CCKA）、胆囊收缩素B（CCKB）和毒蕈碱M3受体的3iL结合。在非洲爪蟾卵母细胞中，SPL或NRB与α（1B）AR共表达表明，SPL降低而NRB增加α（1B）AR介导的Ca（2+）信号强度。相应地，小鼠中SPL的缺失增强了RGS2与NRB的结合以及αAR介导的Ca（2+）信号传导，而NRB的缺失增强了RGS2与SPL的结合并降低了αAR介导的Ca（2+）信号传导。这是由于SPL和NRB对RGS2抑制αAR介导的Ca（信号传导能力的相互调节。这些发现提示了一种GPCR介导的Ca（2+）信号传导调节的新机制，其中SPL/NRB形成一对功能相反的调节因子，通过确定R4家族RGS蛋白的抑制程度来调节GPCR介导的Ca（2+）信号强度。
