Research Article: G Protein Subunit Dissociation and Translocation Regulate Cellular Response to Receptor Stimulation

Date Published: November 11, 2009

Publisher: Public Library of Science

Author(s): Mariangela Chisari, Deepak Kumar Saini, Joon-Ho Cho, Vani Kalyanaraman, N. Gautam, Richard Steinhardt.

Abstract: We examined the role of G proteins in modulating the response of living cells to receptor activation. The response of an effector, phospholipase C-β to M3 muscarinic receptor activation was measured using sensors that detect the generation of inositol triphosphate or diacylglycerol. The recently discovered translocation of Gβγ from plasma membrane to endomembranes on receptor activation attenuated this response. A FRET based G protein sensor suggested that in contrast to translocating Gβγ, non-translocating Gβγ subunits do not dissociate from the αq subunit on receptor activation leading to prolonged retention of the heterotrimer state and an accentuated response. M3 receptors with tethered αq induced differential responses to receptor activation in cells with or without an endogenous translocation capable γ subunit. G protein heterotrimer dissociation and βγ translocation are thus unanticipated modulators of the intensity of a cell’s response to an extracellular signal.

Partial Text: G proteins are the major modulators of cellular responses to external signals in mammalian cells [1], [2]. There is limited information on the role that G proteins play in directly regulating the sensitivity of a cell’s response to an external stimulus in living cells. Studies in intact yeast cells [3] and in a mammalian cell line [4] have attempted to address the quantitative relationship between G protein activation and activity downstream. The kinetics of the rod photoreceptor G protein, Gt, mediated phototransduction activity has also been examined in highly specialized rod photoreceptor cells which are amenable to such studies [5]. Overall however, little is known about such processes with regard to the large families of G protein subunits that are expressed widely in all mammalian cell types. Here we have used imaging methods to examine whether mechanisms at the level of the G protein subunits control the intensity of the response to receptor activation in intact living cells.

We have used live cell imaging of single mammalian cells to identify mechanisms at the basis of the sensitivity of cellular responses to G protein activation by an extracellular signal. We show here that the ability of G protein γ subunit types to translocate away from the plasma membrane on receptor activation controls sensitivity of response. Cells that express a translocating γ subunit demonstrate attenuated PIP2 breakdown in response to M3 receptor stimulation compared to cells that express a non translocating γ subunit. The lowered sensitivity is not due to a specific property of a γ subunit type but due to the translocation of the βγ complex because cells expressing a translocation capable γ subunit mutated to abolish translocation showed heightened sensitivity similar to non translocating γ subunits.



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