Role of FKBP12.6 in cADPR-induced activation of reconstituted ryanodine receptors from arterial smooth muscle
Role of FKBP12.6 in cADPR-induced activation of reconstituted ryanodine receptors from arterial smooth muscle
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DOI：
10.1152/ajpheart.00843.2001
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发表时间：
2002-04-01
影响因子：
4.8
通讯作者：
Li, PL
中科院分区：
文献类型：
--
作者：
Tang, WX;Chen, YF;Li, PL
CADP核糖(CADPR)作为第二信使，激活肌浆网(SR)上的兰尼定受体(RyRs)，动员血管平滑肌细胞内的钙离子。然而，cADPR的作用机制尚不清楚。本研究旨在确定RyRs的辅助蛋白FK-506结合蛋白12.6(FKBP12.6)是否在cADPR诱导的RyRs激活中发挥作用。用抗FKBP12的抗体免疫印迹牛冠状动脉平滑肌细胞(BCASM)和培养的CASM细胞，检测到一个12.6 kDa的蛋白，该抗体也与FKBP12.6反应。FK-506(0.01-10微米)可显著增加重组RyR/Ca(2+)释放通道的开放概率(NPO)。抗FKBP12抗体可阻断FK-506对RyR/Ca~(2+)释放通道的激活作用。RyRs激动剂cADPR(0.1-0um)显著增加RyR/Ca~(2+)释放通道活性。在FK-506存在下，cADPR不进一步增加RyR/Ca~(2+)释放通道的NPO。加入抗FKBP12抗体也可完全阻断cADPR诱导的这些通道的激活，用FK-506预育和随后的梯度离心法去除FKBP12.6可取消cADPR诱导的RyR/Ca~(2+)释放通道NPO的增加。我们认为，FKBP12.6在介导cADPR通过BCASM的SR激活RyR/Ca~(2+)释放通道中起重要作用。
cADP ribose (cADPR) serves as second messenger to activate the ryanodine receptors (RyRs) of the sarcoplasmic reticulum (SR) and mobilize intracellular Ca2+ in vascular smooth muscle cells. However, the mechanisms mediating the effect of cADPR remain unknown. The present study was designed to determine whether FK-506 binding protein 12.6 (FKBP12.6), an accessory protein of the RyRs, plays a role in cADPR-induced activation of the RyRs. A 12.6-kDa protein was detected in bovine coronary arterial smooth muscle (BCASM) and cultured CASM cells by being immunoblotted with an antibody against FKBP12, which also reacted with FKBP12.6. With the use of planar lipid bilayer clamping techniques, FK-506 (0.01-10 muM) significantly increased the open probability (NPO) of reconstituted RyR/Ca(2+)release channels from the SR of CASM. This FK-506-induced activation of RyR/Ca2+ release channels was abolished by pretreatment with anti-FKBP12 antibody. The RyRs activator cADPR (0.1-0 muM) markedly increased the activity of RyR/Ca2+ release channels. In the presence of FK-506, cADPR did not further increase the NPO of RyR/Ca2+ release channels. Addition of anti-FKBP12 antibody also completely blocked cADPR- induced activation of these channels, and removal of FKBP12.6 by preincubation with FK-506 and subsequent gradient centrifugation abolished cADPR- induced increase in the NPO of RyR/Ca2+ release channels. We conclude that FKBP12.6 plays a critical role in mediating cADPR- induced activation of RyR/Ca2+ release channels from the SR of BCASM.