Research ArticleGPCR SIGNALING

Phosphatidylinositol 4-phosphate is a major source of GPCR-stimulated phosphoinositide production

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Science Signaling  11 Sep 2018:
Vol. 11, Issue 547, eaan1210
DOI: 10.1126/scisignal.aan1210

Sourcing phosphoinositides

Agonist binding to certain G protein–coupled receptors (GPCRs) stimulates members of the phospholipase C (PLC) family of enzymes to hydrolyze the plasma membrane lipid phosphatidylinositol 4,5-bisphosphate (PI4,5P2), which results in the generation of the intracellular second messengers DAG and IP3. Noting that certain PLCs also hydrolyze the lipid phosphatidylinositol 4-phosphate (PI4P) at the Golgi, de Rubio et al. performed fluorescence-based imaging to monitor the spatiotemporal regulation of lipid hydrolysis in various cell types stimulated through different GPCRs, as well as the resulting activation of the DAG-dependent kinase PKD. Their findings suggest that PI4P, but not PI4,5P2, at the plasma membrane is the main source of GPCR-stimulated second messenger production.


Phospholipase C (PLC) enzymes hydrolyze the plasma membrane (PM) lipid phosphatidylinositol 4,5-bisphosphate (PI4,5P2) to generate the second messengers inositol trisphosphate (IP3) and diacylglycerol (DAG) in response to receptor activation in almost all mammalian cells. We previously found that stimulation of G protein–coupled receptors (GPCRs) in cardiac cells leads to the PLC-dependent hydrolysis of phosphatidylinositol 4-phosphate (PI4P) at the Golgi, a process required for the activation of nuclear protein kinase D (PKD) during cardiac hypertrophy. We hypothesized that GPCR-stimulated PLC activation leading to direct PI4P hydrolysis may be a general mechanism for DAG production. We measured GPCR activation–dependent changes in PM and Golgi PI4P pools in various cells using GFP-based detection of PI4P. Stimulation with various agonists caused a time-dependent reduction in PI4P-associated, but not PI4,5P2-associated, fluorescence at the Golgi and PM. Targeted depletion of PI4,5P2 from the PM before GPCR stimulation had no effect on the depletion of PM or Golgi PI4P, total inositol phosphate (IP) production, or PKD activation. In contrast, acute depletion of PI4P specifically at the PM completely blocked the GPCR-dependent production of IPs and activation of PKD but did not change the abundance of PI4,5P2. Acute depletion of Golgi PI4P had no effect on these processes. These data suggest that most of the PM PI4,5P2 pool is not involved in GPCR-stimulated phosphoinositide hydrolysis and that PI4P at the PM is responsible for the bulk of receptor-stimulated phosphoinositide hydrolysis and DAG production.

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