Dupré Denis J, Robitaille Mélanie, Richer Maxime, Ethier Nathalie, Mamarbachi Aida M, Hébert Terence E
Department of Pharmacology and Therapeutics, McGill University, Montréal, Québec, H3G 1Y6, Canada.
J Biol Chem. 2007 May 4;282(18):13703-15. doi: 10.1074/jbc.M608846200. Epub 2007 Mar 15.
Heterotrimeric G proteins play a central role in intracellular communication mediated by extracellular signals, and both Galpha and Gbetagamma subunits regulate effectors downstream of activated receptors. The particular constituents of the G protein heterotrimer affect both specificity and efficiency of signal transduction. However, little is known about mechanistic aspects of G protein assembly in the cell that would certainly contribute to formation of heterotrimers of specific composition. It was recently shown that phosducin-like protein (PhLP) modulated both Gbetagamma expression and subsequent signaling by chaperoning nascent Gbeta and facilitating heterodimer formation with Ggamma subunits (Lukov, G. L., Hu, T., McLaughlin, J. N., Hamm, H. E., and Willardson, B. M. (2005) EMBO J. 24, 1965-1975; Humrich, J., Bermel, C., Bunemann, M., Harmark, L., Frost, R., Quitterer, U., and Lohse, M. J. (2005) J. Biol. Chem. 280, 20042-20050). Here we demonstrate using a variety of techniques that DRiP78, an endoplasmic reticulum resident protein known to regulate the trafficking of several seven transmembrane receptors, interacts specifically with the Ggamma subunit but not Gbeta or Galpha subunits. Furthermore, we demonstrate that DRiP78 and the Gbeta subunit can compete for the Ggamma subunit. DRiP78 also protects Ggamma from degradation until a stable partner such as Gbeta is provided. Furthermore, DRiP78 interaction may represent a mechanism for assembly of specific Gbetagamma heterodimers, as selectivity was observed among Ggamma isoforms for interaction with DRiP78 depending on the presence of particular Gbeta subunits. Interestingly, we could detect an interaction between DRiP78 and PhLP, suggesting a role of DRiP78 in the assembly of Gbetagamma by linking Ggamma to PhLP.Gbeta complexes. Our results, therefore, suggest a role of DRiP78 as a chaperone in the assembly of Gbetagamma subunits of the G protein.
异源三聚体G蛋白在由细胞外信号介导的细胞内通讯中起核心作用，Gα和Gβγ亚基均调节活化受体下游的效应器。G蛋白异源三聚体的特定组成成分会影响信号转导的特异性和效率。然而，对于细胞中G蛋白组装的机制方面了解甚少，而这肯定有助于形成特定组成的异源三聚体。最近的研究表明，视紫红质样蛋白（PhLP）通过陪伴新生的Gβ并促进其与Gγ亚基形成异二聚体，调节Gβγ的表达及随后的信号传导（Lukov，G.L.，Hu，T.，McLaughlin，J.N.，Hamm，H.E.和Willardson，B.M.（2005年）《欧洲分子生物学组织杂志》24，1965 - 1975；Humrich，J.，Bermel，C.，Bunemann，M.，Harmark，L.，Frost，R.，Quitterer，U.和Lohse，M.J.（2005年）《生物化学杂志》280，20042 - 20050）。在此，我们使用多种技术证明，DRiP78是一种内质网驻留蛋白，已知其调节几种七跨膜受体的运输，它与Gγ亚基特异性相互作用，但不与Gβ或Gα亚基相互作用。此外，我们证明DRiP78和Gβ亚基可以竞争Gγ亚基。DRiP78还能保护Gγ不被降解，直到提供稳定的伴侣如Gβ。此外，DRiP78的相互作用可能代表了一种组装特定Gβγ异二聚体的机制，因为根据特定Gβ亚基的存在情况，在Gγ同工型与DRiP78的相互作用中观察到了选择性。有趣的是，我们能够检测到DRiP78与PhLP之间的相互作用，这表明DRiP78通过将Gγ连接到PhLP.Gβ复合物，在Gβγ的组装中发挥作用。因此，我们的结果表明DRiP78作为伴侣蛋白在G蛋白的Gβγ亚基组装中发挥作用。
