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. 1996 Apr 2;93(7):2964–2968. doi: 10.1073/pnas.93.7.2964

Identification of a phospholipase C beta2 region that interacts with Gbeta-gamma.

Y Kuang 1, Y Wu 1, A Smrcka 1, H Jiang 1, D Wu 1
PMCID: PMC39743  PMID: 8610151

Abstract

To delineate the phospholipase C (PLC; EC 3.1.4.3) beta2 sequences involved in interactions with the beta-gamma subunits of G proteins, we prepared a number of mammalian expression plasmids encoding a series of PLC beta2 segments that span the region from the beginning of the X box to the end of the Y box. We found the sequence extending from residue Glu-435 to residue Val-641 inhibited Gbeta-gamma-mediated activation of PLC beta2 in transfected COS-7 cells. This PLC beta2 sequence also inhibited ligand-induced activation of PLC in COS-7 cells cotransfected with cDNAs encoding the complement component C5a receptor and PLC beta2 but not in cells transfected with the alpha1B-adrenergic receptor, suggesting that the PLC beta2 residues (Glu-435 to Val-641) inhibit the Gbeta-gamma-mediated but not the Galpha-mediated effect. The inhibitory effect on Gbeta-gamma-mediated activation of PLC beta2 may be the result of the interaction between Gbeta-gamma and the PLC beta2 fragment. This idea was confirmed by the observation that a fusion protein comprising these residues (Glu-435 to Val-641) of PLC beta2 and glutathione S-transferase (GST) bound to Gbeta-gamma in an in vitro binding assay. The Gbeta-gamma-binding region was further narrowed down to 62 amino acids (residues Leu-580 to Val-641) by testing fusion proteins comprising various PLC beta2 sequences and GST in the in vitro binding assay.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Blank J. L., Brattain K. A., Exton J. H. Activation of cytosolic phosphoinositide phospholipase C by G-protein beta gamma subunits. J Biol Chem. 1992 Nov 15;267(32):23069–23075. [PubMed] [Google Scholar]
  2. Boyer J. L., Waldo G. L., Harden T. K. Beta gamma-subunit activation of G-protein-regulated phospholipase C. J Biol Chem. 1992 Dec 15;267(35):25451–25456. [PubMed] [Google Scholar]
  3. Camps M., Carozzi A., Schnabel P., Scheer A., Parker P. J., Gierschik P. Isozyme-selective stimulation of phospholipase C-beta 2 by G protein beta gamma-subunits. Nature. 1992 Dec 17;360(6405):684–686. doi: 10.1038/360684a0. [DOI] [PubMed] [Google Scholar]
  4. Chen J., DeVivo M., Dingus J., Harry A., Li J., Sui J., Carty D. J., Blank J. L., Exton J. H., Stoffel R. H. A region of adenylyl cyclase 2 critical for regulation by G protein beta gamma subunits. Science. 1995 May 26;268(5214):1166–1169. doi: 10.1126/science.7761832. [DOI] [PubMed] [Google Scholar]
  5. Crespo P., Xu N., Simonds W. F., Gutkind J. S. Ras-dependent activation of MAP kinase pathway mediated by G-protein beta gamma subunits. Nature. 1994 Jun 2;369(6479):418–420. doi: 10.1038/369418a0. [DOI] [PubMed] [Google Scholar]
  6. Hawes B. E., Touhara K., Kurose H., Lefkowitz R. J., Inglese J. Determination of the G beta gamma-binding domain of phosducin. A regulatable modulator of G beta gamma signaling. J Biol Chem. 1994 Nov 25;269(47):29825–29830. [PubMed] [Google Scholar]
  7. Inglese J., Koch W. J., Caron M. G., Lefkowitz R. J. Isoprenylation in regulation of signal transduction by G-protein-coupled receptor kinases. Nature. 1992 Sep 10;359(6391):147–150. doi: 10.1038/359147a0. [DOI] [PubMed] [Google Scholar]
  8. Katz A., Wu D., Simon M. I. Subunits beta gamma of heterotrimeric G protein activate beta 2 isoform of phospholipase C. Nature. 1992 Dec 17;360(6405):686–689. doi: 10.1038/360686a0. [DOI] [PubMed] [Google Scholar]
  9. Kim H. K., Kim J. W., Zilberstein A., Margolis B., Kim J. G., Schlessinger J., Rhee S. G. PDGF stimulation of inositol phospholipid hydrolysis requires PLC-gamma 1 phosphorylation on tyrosine residues 783 and 1254. Cell. 1991 May 3;65(3):435–441. doi: 10.1016/0092-8674(91)90461-7. [DOI] [PubMed] [Google Scholar]
  10. Lee C. H., Park D., Wu D., Rhee S. G., Simon M. I. Members of the Gq alpha subunit gene family activate phospholipase C beta isozymes. J Biol Chem. 1992 Aug 15;267(23):16044–16047. [PubMed] [Google Scholar]
  11. Lee C. W., Park D. J., Lee K. H., Kim C. G., Rhee S. G. Purification, molecular cloning, and sequencing of phospholipase C-beta 4. J Biol Chem. 1993 Oct 5;268(28):21318–21327. [PubMed] [Google Scholar]
  12. Logothetis D. E., Kurachi Y., Galper J., Neer E. J., Clapham D. E. The beta gamma subunits of GTP-binding proteins activate the muscarinic K+ channel in heart. Nature. 1987 Jan 22;325(6102):321–326. doi: 10.1038/325321a0. [DOI] [PubMed] [Google Scholar]
  13. Park D., Jhon D. Y., Lee C. W., Lee K. H., Rhee S. G. Activation of phospholipase C isozymes by G protein beta gamma subunits. J Biol Chem. 1993 Mar 5;268(7):4573–4576. [PubMed] [Google Scholar]
  14. Reuveny E., Slesinger P. A., Inglese J., Morales J. M., Iñiguez-Lluhi J. A., Lefkowitz R. J., Bourne H. R., Jan Y. N., Jan L. Y. Activation of the cloned muscarinic potassium channel by G protein beta gamma subunits. Nature. 1994 Jul 14;370(6485):143–146. doi: 10.1038/370143a0. [DOI] [PubMed] [Google Scholar]
  15. Rhee S. G., Choi K. D. Regulation of inositol phospholipid-specific phospholipase C isozymes. J Biol Chem. 1992 Jun 25;267(18):12393–12396. [PubMed] [Google Scholar]
  16. Smrcka A. V., Hepler J. R., Brown K. O., Sternweis P. C. Regulation of polyphosphoinositide-specific phospholipase C activity by purified Gq. Science. 1991 Feb 15;251(4995):804–807. doi: 10.1126/science.1846707. [DOI] [PubMed] [Google Scholar]
  17. Smrcka A. V., Sternweis P. C. Regulation of purified subtypes of phosphatidylinositol-specific phospholipase C beta by G protein alpha and beta gamma subunits. J Biol Chem. 1993 May 5;268(13):9667–9674. [PubMed] [Google Scholar]
  18. Stephens L., Smrcka A., Cooke F. T., Jackson T. R., Sternweis P. C., Hawkins P. T. A novel phosphoinositide 3 kinase activity in myeloid-derived cells is activated by G protein beta gamma subunits. Cell. 1994 Apr 8;77(1):83–93. doi: 10.1016/0092-8674(94)90237-2. [DOI] [PubMed] [Google Scholar]
  19. Tang W. J., Gilman A. G. Type-specific regulation of adenylyl cyclase by G protein beta gamma subunits. Science. 1991 Dec 6;254(5037):1500–1503. doi: 10.1126/science.1962211. [DOI] [PubMed] [Google Scholar]
  20. Taylor S. J., Chae H. Z., Rhee S. G., Exton J. H. Activation of the beta 1 isozyme of phospholipase C by alpha subunits of the Gq class of G proteins. Nature. 1991 Apr 11;350(6318):516–518. doi: 10.1038/350516a0. [DOI] [PubMed] [Google Scholar]
  21. Wickman K. D., Iñiguez-Lluhl J. A., Davenport P. A., Taussig R., Krapivinsky G. B., Linder M. E., Gilman A. G., Clapham D. E. Recombinant G-protein beta gamma-subunits activate the muscarinic-gated atrial potassium channel. Nature. 1994 Mar 17;368(6468):255–257. doi: 10.1038/368255a0. [DOI] [PubMed] [Google Scholar]
  22. Wu D. Q., Lee C. H., Rhee S. G., Simon M. I. Activation of phospholipase C by the alpha subunits of the Gq and G11 proteins in transfected Cos-7 cells. J Biol Chem. 1992 Jan 25;267(3):1811–1817. [PubMed] [Google Scholar]
  23. Wu D., Jiang H., Katz A., Simon M. I. Identification of critical regions on phospholipase C-beta 1 required for activation by G-proteins. J Biol Chem. 1993 Feb 15;268(5):3704–3709. [PubMed] [Google Scholar]
  24. Wu D., Katz A., Lee C. H., Simon M. I. Activation of phospholipase C by alpha 1-adrenergic receptors is mediated by the alpha subunits of Gq family. J Biol Chem. 1992 Dec 25;267(36):25798–25802. [PubMed] [Google Scholar]
  25. Wu D., Katz A., Simon M. I. Activation of phospholipase C beta 2 by the alpha and beta gamma subunits of trimeric GTP-binding protein. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5297–5301. doi: 10.1073/pnas.90.11.5297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wu D., LaRosa G. J., Simon M. I. G protein-coupled signal transduction pathways for interleukin-8. Science. 1993 Jul 2;261(5117):101–103. doi: 10.1126/science.8316840. [DOI] [PubMed] [Google Scholar]
  27. Xu J., Wu D., Slepak V. Z., Simon M. I. The N terminus of phosducin is involved in binding of beta gamma subunits of G protein. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2086–2090. doi: 10.1073/pnas.92.6.2086. [DOI] [PMC free article] [PubMed] [Google Scholar]

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