G protein βγ subunits directly interact with and activate phospholipase Cϵ

J Biol Chem. 2018 Apr 27;293(17):6387-6397. doi: 10.1074/jbc.RA118.002354. Epub 2018 Mar 13.


Phospholipase C (PLC) enzymes hydrolyze membrane phosphatidylinositol 4,5 bisphosphate (PIP2) and regulate Ca2+ and protein kinase signaling in virtually all mammalian cell types. Chronic activation of the PLCϵ isoform downstream of G protein-coupled receptors (GPCRs) contributes to the development of cardiac hypertrophy. We have previously shown that PLCϵ-catalyzed hydrolysis of Golgi-associated phosphatidylinositol 4-phosphate (PI4P) in cardiac myocytes depends on G protein βγ subunits released upon stimulation with endothelin-1. PLCϵ binds and is directly activated by Ras family small GTPases, but whether they directly interact with Gβγ has not been demonstrated. To identify PLCϵ domains that interact with Gβγ, here we designed various single substitutions and truncations of WT PLCϵ and tested them for activation by Gβγ in transfected COS-7 cells. Deletion of only a single domain in PLCϵ was not sufficient to completely block its activation by Gβγ, but blocked activation by Ras. Simultaneous deletion of the C-terminal RA2 domain and the N-terminal CDC25 and cysteine-rich domains completely abrogated PLCϵ activation by Gβγ, but activation by the GTPase Rho was retained. In vitro reconstitution experiments further revealed that purified Gβγ directly interacts with a purified fragment of PLCϵ (PLCϵ-PH-RA2) and increases PIP2 hydrolysis. Deletion of the RA2 domain decreased Gβγ binding and eliminated Gβγ stimulation of PIP2 hydrolysis. These results provide first evidence that Gβγ directly interacts with PLCϵ and yield insights into the mechanism by which βγ subunits activate PLCϵ.

Keywords: G protein; Ras homolog gene family; Ras protein; member A (RhoA); phosphatidylinositol signaling; phospholipase C.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Endothelin-1 / genetics
  • Endothelin-1 / metabolism
  • GTP-Binding Protein beta Subunits / genetics
  • GTP-Binding Protein beta Subunits / metabolism*
  • GTP-Binding Protein gamma Subunits / genetics
  • GTP-Binding Protein gamma Subunits / metabolism*
  • Golgi Apparatus / enzymology*
  • Golgi Apparatus / genetics
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / enzymology*
  • Phosphatidylinositol Phosphates / genetics
  • Phosphatidylinositol Phosphates / metabolism
  • Phosphoinositide Phospholipase C / genetics
  • Phosphoinositide Phospholipase C / metabolism*
  • Protein Domains
  • Rats
  • cdc25 Phosphatases / genetics
  • cdc25 Phosphatases / metabolism*
  • rho GTP-Binding Proteins / genetics
  • rho GTP-Binding Proteins / metabolism


  • Endothelin-1
  • GTP-Binding Protein beta Subunits
  • GTP-Binding Protein gamma Subunits
  • Phosphatidylinositol Phosphates
  • phosphatidylinositol 4-phosphate
  • cdc25 Phosphatases
  • Phosphoinositide Phospholipase C
  • phospholipase C epsilon
  • rho GTP-Binding Proteins