Regulation of F-actin and endoplasmic reticulum organization by the trimeric G-protein G(i2) in rat hepatocytes. Implication for the activation of store-operated Ca2+ inflow

Ying Jie Wang, Roland B. Gregory, Greg J. Barritt

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    Abstract

    The roles of the heterotrimeric G-protein, G(i2), in regulating the actin cytoskeleton and the activation of store-operated Ca2+ channels in rat hepatocytes were investigated. Gα(i2) was principally associated with the plasma membrane and microsomes. Both F-actin and Gα(i2) were detected by Western blot analysis in a purified plasma membrane preparation, the supernatant and pellet obtained by treating the plasma membrane with Triton X-100, and after depolymerization and repolymerization of F-actin in the Triton X-100-insoluble pellet. Actin in the Triton X-100-soluble supernatant coprecipitated with Gα(i2) using either anti-Gα(i2) or antiactin antibodies. The principally cortical location of F-actin in hepatocytes cultured for 0.5 h changed to a pericanalicular distribution over a further 3.5 h. Some Gα(i2) co-localized with F-actin at the plasma membrane. Pretreatment with pertussis toxin ADP-ribosylated 70-80% of Gα(i2) in the plasma membrane and microsomes, prevented the redistribution of F-actin, caused redistribution and fragmentation of the endoplasmic reticulum, and inhibited vasopressin-stimulated Ca2+ inflow. It is concluded that (i) a significant portion of hepatocyte Gα(i2) associates with, and regulates the arrangement of, cortical F-actin and the endoplasmic reticulum and (ii) either or both of these regulatory roles are likely to be required for normal vasopressin activation of Ca2+ inflow.

    Original languageEnglish
    Pages (from-to)22229-22237
    Number of pages9
    JournalJournal of Biological Chemistry
    Volume275
    Issue number29
    DOIs
    Publication statusPublished - 21 Jul 2000

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