Wang Ping, Kumar Puneet, Wang Chang, Defea Kathryn A
Division of Biomedical Sciences, Cell, Molecular and Developmental Biology Program and Biochemistry and Molecular Biology Program, University of California, Riverside, CA 92521, USA.
Biochem J. 2007 Dec 1;408(2):221-30. doi: 10.1042/BJ20070483.
PAR-2 (protease-activated receptor 2) is a GPCR (G-protein-coupled receptor) that can elicit both G-protein-dependent and -independent signals. We have shown previously that PAR-2 simultaneously promotes Galphaq/Ca2+-dependent activation and beta-arrestin-1-dependent inhibition of class IA PI3K (phosphoinositide 3-kinase), and we sought to characterize further the role of beta-arrestins in the regulation of PI3K activity. Whereas the ability of beta-arrestin-1 to inhibit p110alpha (PI3K catalytic subunit alpha) has been demonstrated, the role of beta-arrestin-2 in PI3K regulation and possible differences in the regulation of the two catalytic subunits (p110alpha and p110beta) associated with p85alpha (PI3K regulatory subunit) have not been examined. In the present study we have demonstrated that: (i) PAR-2 increases p110alpha- and p110beta-associated lipid kinase activities, and both p110alpha and p110beta are inhibited by over-expression of either beta-arrestin-1 or -2; (ii) both beta-arrestin-1 and -2 directly inhibit the p110alpha catalytic subunit in vitro, whereas only beta-arrestin-2 directly inhibited p110beta; (iii) examination of upstream pathways revealed that PAR-2-induced PI3K activity required the small GTPase Cdc (cell-division cycle)42, but not tyrosine phosphorylation of p85; and (iv) beta-arrestins inhibit PAR-2-induced Cdc42 activation. Taken together, these results indicated that beta-arrestins could inhibit PAR-2-stimulated PI3K activity, both directly and through interference with upstream pathways, and that the two beta-arrestins differ in their ability to inhibit the p110alpha and p110beta catalytic subunits. These results are particularly important in light of the growing interest in PAR-2 as a pharmacological target, as commonly used biochemical assays that monitor G-protein coupling would not screen for beta-arrestin-dependent signalling events.
蛋白酶激活受体2（PAR-2）是一种G蛋白偶联受体（GPCR），可引发G蛋白依赖性和非依赖性信号。我们之前已经表明，PAR-2同时促进Gαq/ Ca2+依赖性激活和β-抑制蛋白1依赖性对IA类磷脂酰肌醇3激酶（PI3K）的抑制，并且我们试图进一步阐明β-抑制蛋白在PI3K活性调节中的作用。虽然已经证实β-抑制蛋白1具有抑制p110α（PI3K催化亚基α）的能力，但尚未研究β-抑制蛋白2在PI3K调节中的作用以及与p85α（PI3K调节亚基）相关的两个催化亚基（p110α和p110β）在调节方面可能存在的差异。在本研究中，我们已经证明：（i）PAR-2增加与p110α和p110β相关的脂质激酶活性，并且p110α和p110β均受到β-抑制蛋白1或β-抑制蛋白2过表达的抑制；（ii）β-抑制蛋白1和β-抑制蛋白2在体外均直接抑制p110α催化亚基，而只有β-抑制蛋白2直接抑制p110β；（iii）对上游途径的研究表明，PAR-2诱导的PI3K活性需要小GTP酶细胞分裂周期蛋白（Cdc）42，但不需要p85的酪氨酸磷酸化；并且（iv）β-抑制蛋白抑制PAR-2诱导的Cdc42激活。综上所述，这些结果表明，β-抑制蛋白可以直接并通过干扰上游途径来抑制PAR-2刺激的PI3K活性，并且两种β-抑制蛋白在抑制p110α和p110β催化亚基的能力上有所不同。鉴于对PAR-2作为药理学靶点的兴趣日益增加，这些结果尤为重要，因为常用的监测G蛋白偶联的生化分析方法不会筛选β-抑制蛋白依赖性信号事件。
