The small G protein Rac1 activates phospholipase Cdelta1 through phospholipase Cbeta2.
Rac1, which is associated with cytoskeletal pathways, can activate phospholipase Cbeta2 (PLCbeta2) to increase intracellular Ca(2+) levels. This increased Ca(2+) can in turn activate the very robust PLCdelta1 to synergize Ca(2+) signals. We have previously found that PLCbeta2 will bind to and inhibit PLCdelta1 in solution by an unknown mechanism and that PLCbeta2.PLCdelta1 complexes can be disrupted by Gbetagamma subunits. However, because the major populations of PLCbeta2 and PLCdelta1 are cytosolic, their regulation by Gbetagamma subunits is not clear. Here, we have found that the pleckstrin homology (PH) domains of PLCbeta2 and PLCbeta3 are the regions that result in PLCdelta1 binding and inhibition. In cells, PLCbeta2.PLCdelta1 form complexes as seen by Förster resonance energy transfer and co-immunoprecipitation, and microinjection of PHbeta2 dissociates the complex. Using PHbeta2 as a tool to assess the contribution of PLCbeta inhibition of PLCdelta1 to Ca(2+) release, we found that, although PHbeta2 only results in a 25% inhibition of PLCdelta1 in solution, in cells the presence of PHbeta2 appears to eliminates Ca(2+) release suggesting a large threshold effect. We found that the small plasma membrane population of PLCbeta2.PLCdelta1 is disrupted by activation of heterotrimeric G proteins, and that the major cytosolic population of the complexes are disrupted by Rac1 activation. Thus, the activity of PLCdelta1 is controlled by the amount of bound PLCbeta2 that changes with displacement of the enzyme by heterotrimeric or small G proteins. Through PLCbeta2, PLCdelta1 activation is linked to surface receptors as well as signals that mediate cytoskeletal pathways.
Department of Physiology and Biophysics, Stony Brook University, Stony Brook, New York 11794-8661, USA.
Yuanjian
Y
Guo
Urszula
U
Golebiewska
Stephen
S
D'Amico
Suzanne
S
Scarlata
D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D019204:GTP-Binding Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D008954:Models, Biological; D010716:Phosphatidylinositols / Q000737:chemistry; D054799:Phospholipase C beta / Q000378:metabolism*; D054803:Phospholipase C delta / Q000378:metabolism*; D011485:Protein Binding; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D010716:Phosphatidylinositols; C107953:RAC1 protein, human; D049452:Green Fluorescent Proteins; D054799:Phospholipase C beta; D054803:Phospholipase C delta; D019204:GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; D002118:Calcium