Postdoctoral fellow University of Michigan Ann Arbor, Michigan, United States
Hoa Phan (University of Michigan)| Joseph Loomis (University of Michigan)| Murat Bastepe (Harvard Medical School, Harvard Medical School)| Qing He (Harvard Medical School, Harvard Medical School)| Alan Smrcka (University of Michigan)
Phospholipase C (PLC) enzymes are essential for normal cellular function. Upon stimulation by GPCRs or RTKs, PLCs hydrolyze phosphatidylinositol 4,5-bisphosphate (PIP2) at cell membranes to generate diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP3). DAG and IP3 are important second messengers in signal transduction. DAG regulates activity of protein kinase C and IP3 mobilizes intracellular Ca2+, which initiate multiple signaling cascades and cellular processes. Dysregulation of PLC function gives rise to pathological processes. Among six mammalian PLC subfamilies, PLCβ isozymes are activated by Gαq, Gβγ, small G proteins and thus, play significant roles in various cellular activities.
Extra-large stimulatory Gα (XLαs) is a large variant of Gαs. Biochemically, XLαs activation by GPCR mediates cAMP generation similarly to Gαs, however, Gαs and XLαs have distinct cellular and physiological functions. He et al. suggests that XLαs is able to stimulate IP3 signaling in vivo. In this study, we aim to identify the PLC that mediates this phenomenon. By co-expressing XLαs with different PLC constructs in cells and measuring total IP production, we identify that XLαs stimulates PLCβ4 to generate IP3. This activity is specific to XLαs because wild-type and constitutively active Gαs do not activate PLCβ4. Using purified proteins, we show that XLαs directly and specifically activates PLCβ4. Interestingly, unlike Gαq, XLαs does not require GTP to activate its effector PLCβ4. We also identify that the helical domain preceding the nucleotide-binding domain in XLαs is required for its ability to stimulate PLCβ4. Further studies are underway to determine the mode of activation of PLCβ4 by XLαs and the role of XLαs – mediated stimulation of PLCβ4 in physiological systems.Overall, we identify XLαs as a novel direct activator of PLCβ4 and this novel signaling axis potentially play important roles physiologically.
Support or Funding Information
Upjohn postdoctoral fellowship to HP NIH GM R35 to AVS