Heterotrimeric G proteins mediate signaling across the plasma membranes of cells for many hormones and neurotransmitters, as well as for a wide variety of other regulatory agents. They are known or likely sites for disease processes such as cholera and pertussis, and their role in cellular regulation makes them likely contributors to the pathology, if not etiology, of complex diseases such as diabetes, essential hypertension and cancer. Understanding how these proteins function in normal signaling events is essential to describing their role in disease states and to providing information about the future possible development of these proteins as targets for pharmacologic intervention. One of the two main hypotheses upon which this research is based is that the widespread involvement of these proteins in cell signaling depends upon the immense structural and functional diversity of the G protein subunits. A primary objective of the proposed studies is to define the role of subunit diversity in signaling by heterotrimeric G proteins. Thus, the first two Specific Aims are: (1) To determine the origin and nature of the structural diversity of the beta and gamma subunits of G proteins. These studies take advantage of protein chemistry and mass spectrometry expertise associated with this project. (2) To determine the functional differences between Gbetagamma dimers containing (a) different beta or gamma isoforms or (b) differently modified gamma subunit isoforms. These studies use primarily molecular biology approaches to apply the information generated under Specific Aim 1 to the biochemical characterization of the G protein subunits. An important functional step in the action of G proteins is the GTP-dependent dissociation of their a subunit from their associated betagamma dimer. This reaction has diverse implications for the cellular signaling processes mediated by G proteins, in large part due to their structural and functional diversity. We hypothesize that G protein subunit dissociation is a key step in signal transduction that can determine to what signals cells respond, as well as the nature of that response. To test this hypothesis the third Specific Aim is: (3) To determine the physiological role of subunit dissociation in intact membranes and intact cells. These studies will specifically test whether the G protein subunit diversity characterized under Specific Aims 1 and 2 is utilized by cells to generate changing patterns of G protein heterotrimers with varying signaling properties. The results of this project will define the potential range of functional diversity associated with these proteins, and in so doing define their diversity as potential drug targets.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK037219-15
Application #
6380543
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Abraham, Kristin M
Project Start
1991-07-01
Project End
2004-06-30
Budget Start
2001-07-01
Budget End
2002-06-30
Support Year
15
Fiscal Year
2001
Total Cost
$294,580
Indirect Cost
Name
Medical University of South Carolina
Department
Pharmacology
Type
Schools of Medicine
DUNS #
183710748
City
Charleston
State
SC
Country
United States
Zip Code
29425
Kilpatrick, Eric L; Hildebrandt, John D (2007) Sequence dependence and differential expression of Ggamma5 subunit isoforms of the heterotrimeric G proteins variably processed after prenylation in mammalian cells. J Biol Chem 282:14038-47
Yang, Wanling; Hildebrandt, John D (2006) Genomic analysis of G protein gamma subunits in human and mouse - the relationship between conserved gene structure and G protein betagamma dimer formation. Cell Signal 18:194-201
Cook, Lana A; Schey, Kevin L; Wilcox, Michael D et al. (2006) Proteomic analysis of bovine brain G protein gamma subunit processing heterogeneity. Mol Cell Proteomics 5:671-85
Wells, Christopher A; Dingus, Jane; Hildebrandt, John D (2006) Role of the chaperonin CCT/TRiC complex in G protein betagamma-dimer assembly. J Biol Chem 281:20221-32
Dingus, Jane; Wells, Christopher A; Campbell, Lia et al. (2005) G Protein betagamma dimer formation: Gbeta and Ggamma differentially determine efficiency of in vitro dimer formation. Biochemistry 44:11882-90
Yang, Wanling; White, Brook; Spicer, Eleanor K et al. (2004) Complex haplotype structure of the human GNAS gene identifies a recombination hotspot centred on a single nucleotide polymorphism widely used in association studies. Pharmacogenetics 14:741-7
Cleator, John H; Ravenell, Roneka; Kurtz, David T et al. (2004) A dominant negative Galphas mutant that prevents thyroid-stimulating hormone receptor activation of cAMP production and inositol 1,4,5-trisphosphate turnover: competition by different G proteins for activation by a common receptor. J Biol Chem 279:36601-7
Ribas, Catalina; Takesono, Aya; Sato, Motohiko et al. (2002) Pertussis toxin-insensitive activation of the heterotrimeric G-proteins Gi/Go by the NG108-15 G-protein activator. J Biol Chem 277:50223-5
Cook, Lana A; Wilcox, Michael D; Dingus, Jane et al. (2002) Separation and analysis of G protein gamma subunits. Methods Enzymol 344:209-33
Ribas, Catalina; Sato, Motohiko; Hildebrandt, John D et al. (2002) Analysis of signal transfer from receptor to Go/Gi in different membrane environments and receptor-independent activators of brain G protein. Methods Enzymol 344:140-52

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