James C. Garrison
Primary AppointmentProfessor, Pharmacology
<body>Role of the G protein &alpha and &beta&gamma Subunits in Cell Signaling </body>
Receptors which raise cyclic AMP levels inhibit the activation of hematopoietic cells. Thus, phosphorylation of important regulatory sites via the cyclic AMP depended protein kinase in hematopoietic cells is central to the inhibitory response. Both PI 3-kinase and P-Rex1 can be phosphorylated in vitro by the cyclic AMP depended protein kinase (PKA). This event inhibits the activity of both of these enzymes. Our work examines the ability of pure G protein α and βγ subunits to regulate PI 3-kinase and P-Rex1 in synthetic lipid vesicles containing PIP3 and PI 3-kinase or Rac to determine which G protein subunits modify the activity of these enzymes. In another project we are phosphorylating pure, recombinant PI 3-kinase and P-Rex1 with the cyclic AMP depended protein kinase and measuring the effect of phoshorylation on their activity in the presence of PIP3 and the G protein subunits.In a third project, we are examining how PI 3-kinase and P-Rex1 respond to activation of G protein coupled receptors in HEK-293 cells, macrophages and neutrophils. We are also using small, inhibitory RNA's delivered to these cells by transfection or stable infection with lentiviruses to determine which isoforms of G proteins regulate PI 3-kinase and P-Rex1 activity in a cellular context. Other effectors examined in these experiments include: adenylyl cyclase, PLC-β, PtdIns 3-kinase and production of certain cytokines such as TNF-α.