Characterization of phosphoinositide-specific phospholipase C in rat colonocyte membranes.
Characterization of phosphoinositide-specific phospholipase C in rat colonocyte membranes.
复制标题
大鼠结肠细胞膜中磷酸肌醇特异性磷脂酶 C 的表征。
DOI：
10.1042/bj2920271
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Sitrin,MD
中科院分区：
文献类型：
--
作者：
Bolt,MJ;Bissonnette,BM;Wali,RK;Hartmann,SC;Brasitus,TA;Sitrin,MD
The phosphoinositide signal transduction pathway mediates important processes in intestinal physiology, yet the key enzyme, phosphoinositide-specific phospholipase C (PI-PLC), is not well-characterized in the colon. PI-PLC activity was examined in rat colonic membranes using exogenous [3H]phosphatidylinositol 4,5-bisphosphate (PIP2) as substrate, and beta-glycerophosphate to suppress degradation of substrate or product. The activity of membrane PI-PLC increased 6-fold with the addition of alamethicin, and a further 2-3-fold enhancement was observed with 10 microM guanosine 5′-[gamma-thio]triphosphate (GTP[S]), suggesting the involvement of G-protein(s). The effect of GTP[S] appeared to be specific, as up to 100 microM adenosine 5′-[gamma-thio]-triphosphate failed to stimulate PI-PLC activity, and guanosine 5′-[beta-thio]diphosphate inhibited activity. The response of membrane PI-PLC to Ca2+ was biphasic, while > 0.5 mM Mg2+ was inhibitory with or without GTP[S]. Comparable total PI-PLC activities and responses to GTP[S] and Ca2+ were observed in purified brush-border and basolateral membranes. Western immunoblots probed with monoclonal antibodies to PLC isoenzymes PLC-beta 1, -gamma 1 and -delta 1 demonstrated that these antipodal plasma membranes contain predominantly the PLC-delta 1 isoform, with small amounts of PLC-gamma 1 present but no detectable PLC-beta 1. PLC-gamma 1 was the major isoform detected in cytosol.
