Regulation of intracellular cyclic AMP concentrations is principally controlled at the level of its synthesis, through the hormonal regulation of adenylylcyclase, the enzyme responsible for the conversion of ATP into cyclic AMP. The adenylylcyclase system is comprised of three components: heptahelical, G protein-coupled receptors for a variety of hormones, neurotransmitters, and autocoids; heterotrimeric G proteins; and the catalytic entity itself. The G proteins regulate the activity of the enzyme in response to the interaction of ligands with an appropriate receptor. This molecular architecture is common to all G protein regulated effector systems identified to date and include hormone regulated phospholipases and ion channels, as well as the light activated cyclic GMP phosphodiesterases. Central to the regulation of adenylylcyclase, or any of the other effector molecules, is the specificity of the G proteins to couple an appropriate receptor to the correct effector. An additional level of regulation is achieved by the cross-talk of different G protein-coupled effector systems, often mediated by the action of downstream protein kinases. The recent cloning of multiple isoforms of adenylylcyclases has permitted the biochemical demonstration that these cyclases are regulated by G proteins and protein kinases in an isoform specific fashion. The two aims of this project are: 1) to determine the precise molecular signals underlying the specificity of recognition of adenylylcyclase by G proteins, and 2) to characterize the role of phosphorylation in the regulation of adenylylcyclase isoforms. Two genetic system (based on the expression of mammalian adenylylcyclases and G protein subunits in yeast) will be used to select for regulatory mutants of both adenylylcyclases and G protein subunits. Isolation of mutant defective in coupling G protein subunits to adenylylcyclases will allow to probe the molecular basis of G protein recognition and regulation of adenylylcyclase. In addition, in vitro and in vivo approaches will be used to elucidate the regulation of specific adenylylcyclase isoforms by phosphorylation. These studies will have a significant impact on the understanding of the mechanisms underlying the regulation of effector systems by G proteins and the intricate cross-talk among different hormone regulated signaling pathways.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM053645-04
Application #
2872704
Study Section
Biochemistry Study Section (BIO)
Project Start
1996-02-01
Project End
2001-01-14
Budget Start
1999-02-01
Budget End
2001-01-14
Support Year
4
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Biochemistry
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
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Clapp, Peter; Capper, Austin B; Taussig, Ronald (2002) Genetic selection of regulatory mutants of mammalian adenylyl cyclases. Methods Enzymol 345:241-51
Scott, J K; Huang, S F; Gangadhar, B P et al. (2001) Evidence that a protein-protein interaction 'hot spot' on heterotrimeric G protein betagamma subunits is used for recognition of a subclass of effectors. EMBO J 20:767-76
Watts, V J; Taussig, R; Neve, R L et al. (2001) Dopamine D2 receptor-induced heterologous sensitization of adenylyl cyclase requires Galphas: characterization of Galphas-insensitive mutants of adenylyl cyclase V. Mol Pharmacol 60:1168-72
Hou, Y; Chang, V; Capper, A B et al. (2001) G Protein beta subunit types differentially interact with a muscarinic receptor but not adenylyl cyclase type II or phospholipase C-beta 2/3. J Biol Chem 276:19982-8
Shoshani, I; Taussig, R; Iyengar, R et al. (2000) Synthesis and use of 3'-(azidoiodosalicyl) derivatives of 2', 5'-dideoxyadenosine as photoaffinity ligands for adenylyl cyclase. Arch Biochem Biophys 376:221-8
Zimmermann, G; Zhou, D; Taussig, R (1999) Activating mutation of adenylyl cyclase reverses its inhibition by G proteins. Mol Pharmacol 56:895-901
Lan, K L; Sarvazyan, N A; Taussig, R et al. (1998) A point mutation in Galphao and Galphai1 blocks interaction with regulator of G protein signaling proteins. J Biol Chem 273:12794-7
Taussig, R; Zimmermann, G (1998) Type-specific regulation of mammalian adenylyl cyclases by G protein pathways. Adv Second Messenger Phosphoprotein Res 32:81-98
Zimmermann, G; Zhou, D; Taussig, R (1998) Genetic selection of mammalian adenylyl cyclases insensitive to stimulation by Gsalpha. J Biol Chem 273:6968-75

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