| Many transmembrane signaling systems consist of specific G protein-coupled receptors (GPCRs) that transduce the binding of extracellular ligands into intracellular signaling events. GPCRs modulate the activity of numerous effector molecules including various adenylyl cyclases, PI 3-kinases, non-receptor tyrosine kinases, small G proteins, phospholipases, phosphodiesterases and ion channels. GPCRs regulate multiple biological functions including neurotransmission, sensory perception, chemotaxis, embryogenesis, development, HIV infection, cell growth, differentiation, and apoptosis, and oncogenesis. In particular, a number of oncogenic GPCRs have been identified including the mas oncogene, several thyroid-stimulating hormone and luteinizing hormone receptor mutants, a GPCR encoded by the Kaposi's sarcoma-associated herpesvirus, and receptors for the Wnt signaling pathway. Moreover, paracrine and autocrine activation of various neuropeptide and prostanoid GPCRs has been implicated in small cell lung carcinoma, colon adenomas and carcinomas, and gastric cancer.|
One of the principal mechanisms for regulating GPCR responsiveness involves a rapid hormone-dependent phosphorylation of the receptor that functions to turn-off signaling. This phosphorylation is primarily mediated by specific G protein-coupled receptor kinases (GRKs) that have the unique ability to recognize and phosphorylate only the activated conformation of the receptor. The subsequent loss of receptor signaling is then mediated by arrestin proteins that specifically bind to the phosphorylated GPCR and uncouple receptor and G protein. Given the tremendous functional and cellular diversity of GPCR-mediated signaling, GRKs and arrestins likely serve a critical role in controlling cellular activation and growth.
While GRKs mediate agonist-dependent phosphorylation and desensitization of multiple GPCRs, recent studies suggest that GRKs may have additional cellular functions (e.g., in development). In an effort to further elucidate such functions, we have attempted to identify novel GRK-interacting proteins utilizing affinity-chromatography and yeast two-hybrid strategies. Interestingly, these studies reveal that GRKs are capable of interacting with a number of additional proteins including the cytoskeleton proteins tubulin and actin, G protein α and βγ subunits, various calcium binding proteins, and the protein phosphatase PP2A. In some cases these interactions appear to regulate GRK activities in either a positive (G
Recent insight into the role of arrestins in GPCR regulation has demonstrated that while arrestin/GPCR interaction plays an important role in quenching signaling, some arrestins are also capable of binding to clathrin and the adaptor protein AP2 and mediating GPCR endocytosis. Further mechanistic and structural insight into arrestin/clathrin interaction is being performed in collaboration with Drs. Keen and Brenner and should help to delineate the role of arrestins in GPCR endocytosis, recycling, and downregulation. Interestingly, recent studies suggest that arrestins also play a role in the formation of signalling complexes that enable activated GPCRs to couple to MAP kinase stimulation. In this regard, non-visual arrestins contain a proline-rich domain that mediates arrestin interaction with SH3 domain containing proteins such as src. Thus, arrestins may have a dual role in turning-off some signaling pathways while turning-on MAP kinase cascades. Current efforts are focused on further defining the role of GRKs and arrestins in regulating GPCR signaling and trafficking.
Another area of investigation in my laboratory involves the role of GPCRs and GPCR/growth factor receptor cross-talk in cellular activation and growth. In this regard, multiple GPCRs including the receptors for thrombin, bombesin, neurokinins, endothelin, serotonin, acetylcholine, and LPA elicit a mitogenic response in various cell types. While the pathways responsible for GPCR activation of MAP kinases are not completely defined, several key players have been identified including G
Keywords: receptors, protein kinases, signal transduction, endocytosis
|Selected Publications Carman, C. V., Barak, L. S., Chen, C., Liu-Chen, L.-Y., Onorato, J. J., Kennedy, S. P., Caron, M. G., and Benovic, J. L. Mutational analysis of G|
Mundell, S. J. and Benovic, J. L. Selective regulation of endogenous G protein-coupled receptors by arrestins in HEK293 cells. J. Biol. Chem. 275: 12900-12908, 2000.
Tiruppathi, C., Yan, W., Sandoval, R., Naqvi, T., Pronin, A. N., Benovic, J. L., and Malik, A. B. G protein-coupled receptor kinase-5 regulates thrombin-activated signalling in endothelial cells. Proc. Natl. Acad. Sci. USA 97: 7440-7445, 2000.
Pronin, A.N., Morris, A.J., Surguchov, A., and Benovic, J.L. Synucleins are a novel class of substrates for G protein-coupled receptor kinases. J. Biol. Chem. 275: 26515-26522, 2000.
Kallal, L. and Benovic, J. L. Using green fluorescent proteins to study G-protein-coupled receptor localization and trafficking. Trends Pharm. Sci. 21: 175-180, 2000.
Mundell, S.J., Matharu, A.-L., Kelly, E., and Benovic, J.L. Arrestin isoforms dictate differential kinetics of A2B adenosine receptor trafficking. Biochemistry 39: 12828-12836, 2000.
Penn, R.B., Pronin, A.N., and Benovic, J.L. Regulation of G protein-coupled receptor kinases. Trends Cardiovasc. Med. 10: 81-88, 2000.
Parent, J.-L., Labrecque, P., and Benovic, J. L. Role of the differentially spliced carboxyl-terminus in thromboxane A2 receptor trafficking: Identification of a distinct motif for tonic internalization. J. Biol. Chem. 276: 7079-7085, 2001.
Penn, R. B., Pascual, R. M., Kim, Y.-M., Mundell, S. J., Krymskaya, V.P. Jr. Panettieri, R. and Benovic, J. L. Arrestin specificity for G protein-coupled receptors in human airway smooth muscle. J. Biol. Chem. 276: 32648-32656, 2001.
Marchese, A. and Benovic, J. L. Agonist-promoted ubiquitination of the G-protein-coupled receptor CXCR4 mediates lysosomal sorting. J Biol Chem. 276: 45509-45512, 2001.
Milano, S.K., Pace, H.C., Kim, Y.-M., Brenner, C., and Benovic, J.L. Scaffolding functions of arrestin-2 revealed by crystal structure and mutagenesis. Biochemistry 41: 3321-3328, 2002.
Kim, Y.M., Barak, L.S., Caron, M.G., and Benovic, J.L. Regulation of arrestin-3 phosphorylation by casein kinase II. J. Biol. Chem. 277: 16837-16846, 2002.
Kim, Y.M. and Benovic, J. L. Differential roles of arrestin-2 interaction with clathrin and AP2 in G protein-coupled receptor trafficking. J. Biol. Chem. 277: 30760-30768, 2002.
Bhattacharya, M., Anborgh, P. H., Babwah, A. V., Dale, L. B., Dobransky, T., Benovic, J. L., Feldman, R.D., Verdi, J. M., Rylett, R.J., and Ferguson, S.S.G. β-Arrestins regulate a Ral-GDS-Ral effector pathway that mediates cytoskeletal reorganization. Nat. Cell Biol. 4: 547-555, 2002.
DeGraff, J.L., Gurevich, V.V., and Benovic, J.L. The third intracellular loop of α2-adrenergic receptors determines subtype specificity of arrestin interaction. J. Biol. Chem. 277: 43247-43252, 2002.
Pao, C. and Benovic, J. Phosphorylation-independent desensitization of G protein-coupled receptors? Science's STKE (2002), http://stke.sciencemag.org/cgi/content/full/sigtrans;2002/ 153/pe42
Sterne-Marr, R., Tesmer, J.J., Day, P.W., Stracquatanio, R.P., Cilente, J.A., O'Connor, K.E., Pronin, A.N., Benovic, J.L., and Wedegaertner, P.B. GRK2:Gαq/11 interaction: A novel surface on an RGS homology domain for binding G subunits. J. Biol. Chem., in press.
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