Differential regulation of NPR-B/GC-B by protein kinase c and calcium
Differential regulation of NPR-B/GC-B by protein kinase c and calcium
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DOI：
10.1016/j.bcp.2005.04.034
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发表时间：
2005-09-01
影响因子：
5.8
通讯作者：
Potter, LR
中科院分区：
文献类型：
--
作者：
Abbey-Hosch, SE;Smirnov, D;Potter, LR
c型利钠肽（CNP）活化的胍基环化酶连接的利钠肽受体- b （NPR-B）刺激血管松弛和骨骼生长。激素和磷酯（PMA）分别以钙依赖性和蛋白激酶c依赖性的方式抑制NPR-B。在这里，我们描述了暴露于PMA、钙离子载体、离子霉素或鞘氨醇-1-磷酸（S1P）的细胞膜上的NPR-B的动力学性质。PMA和离子霉素主要提高了NPR-B的K-m，降低了V-max；，而SIP对这两个参数的影响都不大。PMA和SIP处理使CNP激活的EC50分别提高了8倍和3倍，而离子霉素则无效。所有三种药物都引起了NPR-B的去磷酸化，但磷酸盐损失的基础在不同的处理之间有所不同。体外膜上的NPR-B磷酸化被先前全细胞PMA或S1P暴露明显减少，而离子霉素预处理没有影响。已知磷酸化残基在每个过程中的参与是用含有谷氨酸替代这些位点的突变受体来测试的。虽然PMA对该受体的作用丧失，但SIP和离子霉素的作用仅被部分阻断。我们的数据表明，PMA和钙依赖抑制NPR-B的分子基础是独特的。前者是由于一个已知位点的磷酸化减少，主要影响NPR-B对CNP和GTP的亲和力。后者与最大速度的降低有关，其机制不涉及抑制NPR-B磷酸化，除了已知位点的去磷酸化外，还需要一个过程。(c) 2005爱思唯尔公司版权所有。
C-type natriuretic peptide (CNP) activation of the guanylyl cyclase-linked natriuretic peptide receptor-B (NPR-B) stimulates vasorelaxation and bone growth. Hormones and phorbol esters (PMA) inhibit NPR-B in calcium and protein kinase c-dependent manners, respectively. Here, we characterize the kinetic properties of NPR-B in membranes from cells exposed to PMA, the calcium ionophore, ionomycin, or sphingosine-1-phosphate (S1P). PMA and ionomycin primarily increased the K-m and decreased the V-max of NPR-B; for GTP, respectively, whereas SIP caused modest changes in both parameters. PMA and SIP treatment increased the EC50 for CNP activation by eight- and three-fold, whereas ionomycin was ineffective. All three agents caused NPR-B dephosphorylation, but the basis for the loss of phosphate differed between treatments. In vitro phosphorylation of NPR-B in membranes was markedly diminished by prior whole cell PMA or S1P exposure, whereas ionomycin pretreatment had no effect. The involvement of the known phosphorylated residues in each process was tested with a mutant receptor containing glutamates substituted for these sites. While the effect of PMA was lost on this receptor, the effects of SIP and ionomycin were only partially blocked. Our data suggest that the molecular bases for PMA- and calcium-dependent inhibition of NPR-B are unique. The former results from reduced phosphorylation of a known site and primarily affects the affinity of NPR-B for CNP and GTP. The latter is associated with reductions in maximal velocities by a mechanism that does not involve inhibition of NPR-B phosphorylation and requires a process in addition to the dephosphorylation of the known sites. (c) 2005 Elsevier Inc. All rights reserved.