G Protein Signaling

 G Protein Signaling Background

The heterotrimeric guanine nucleotide-binding proteins (G proteins) are membrane-associated signal transducers that transmit signals from cell surface receptors to intracellular effectors. In humans, there are 21 Gα subunits, 6 Gβ subunits, and 12 Gγ subunits. Without stimulation, G proteins exist as a heterotrimer, i.e., αβγ, associated with seven transmembrane G-protein-coupled receptors (GPCRs). On agonist binding to a GPCR, a conformational change in the receptor catalyses the G protein α subunit to release GDP, bind GTP, and dissociate from the G protein βγ subunits. Depending on the system, either Gα or Gβγ or both transmits the signal to activate the downstream effectors, such as adenylyl cyclase, ion channels, phospholipases, metabolic enzymes, transcriptional regulators or protein kinase cascade. Signaling ends when GTP is hydrolyzed to GDP and Gα reassociates with Gβγ, completing the cycle of activation. Gα itself is a GTPase, which catalyzes hydrolysis of GTP to GDP. The GTPase activity of Gα is promoted by RGS proteins, the regulator of G protein signaling, which act as GTPase accelerating proteins (GAPs).

G protein signaling pathways are involved in a wide variety of cellular responses to an enormous array of stimuli, including hormones, neurotransmitters, odors, light cytokines, antigens and stress, as well as to changes in extracellular matrix and cell-cell contacts. A large number of cellular processes including transcription, proliferation, differentiation, apoptosis, motility, contractility and secretion depend on normal activity of G proteins. In a variety of organisms, cell polarity is determined by G protein signaling. For example, G proteins regulate appropriate orientation of cell division axes in early Caenorhabditis elegans embryos, cellular response to chemotactic cytokines in leukocytes, and polarization of precursor cells as well as asymmetric cell divisions in Drosophila melanogaster.

Defects in G protein signaling pathways and the aberrant activation of G proteins can lead to human diseases and debilitating conditions, such as cardiac hypertrophy, cardiac arrest, changes of glucose metabolism in liver and muscle, pain, inflammation and cancer. For instance, cholera, a condition associated with severe diarrhea, is caused by Vibrio cholera. This pathogenic exotoxin promotes ADP-ribosylation of Gαs in the intestinal epithelium. Whooping cough is caused by Bordetella pertussis, whose toxin ADP-ribosylates Gαi. Endocrine hyperfunction and hyperplasia in gonad, adrenal cortex, thyroid, or pituitary somatrotrophs results from mutations that block the GTPase activity of Gαs and Gαi.

Given the importance of G protein signaling to human pathogenesis, G proteins could serve as potential drug targets, particularly in a situation where the receptor loses its control due to a genetic defect and becomes unresponsive, constitutively active, or chronically desensitized. Given that a large number of receptor subtypes act on a much smaller number of G proteins, drugs targeting at a specific G protein should have a greater impact on cell signaling. Thus, a more thorough understanding of how G protein signaling is regulated would be useful in revealing novel disease mechanisms and could dramatically extend our ability to better define the targets for therapeutic intervention.


G protein signaling in yeast

In Baker’s yeast Saccharomyces cerevisia, only two GPCR systems have been identified. One is responsible for glucose-sensing, and another is responsible for pheromone-detection. The former mediates glucose activation of adenylate cyclase during the switch from respirative/gluconeogenic metabolism to fermentation. The latter mediates pheromone induced mating response, when naïve yeast cells are exposed to the mating pheromone. Compared to the yeast mating signaling pathway, less is known about the glucose-sensing signaling pathway. It is known that the glucose receptor Gpr1 is a putative G-protein coupled receptor and Gpa2 is the Gα. However, the identity of Gβ and Gγ remains to be revealed. For this pathway, binding of glucose to the receptor Gpr1 activates the Gα protein Gpa2. Gpa2 is then thought to activate adenylate cyclase (Cdc35/Cyr1), which produces cAMP to activate protein kinase A, leading to stimulation of invasive growth and pseudohyphal differentiation, loss of stress resistance, mobilization of trehalose and glycogen, and reduced life-span.

As for the GPCR system of the mating pathway, the pheromone receptor Ste2 or Ste3 (depending on the cell type), is the GPCR coupled to the heterotrimeric G-protein consisting of the Gα Gap1, Gβ Ste4 and Gγ Ste18 in naïve cells not exposed to pheromone. When cells are stimulated with pheromone, binding of pheromone to its receptor leads to a conformational change and activation of the receptor. The activated receptor then catalyzes the exchange of guanosine diphosphate (GDP) for guanosine triphosphate (GTP) on Gpa1. Guanine-nucleotide exchange on Gα Gpa1 leads to its dissociation from the Gβγ composed of Ste4 and Ste18. The Gβγ dimer can then propagate the mating signal through activation of effector proteins, including a protein kinase (Ste20), a kinase scaffolding protein (Ste5), the GDP-GTP exchange factor of a small GTPase Cdc42 (Cdc24), and a scaffolding protein (Far1) that is critical for cells growth arrest and polarized cell growth. Activation of Ste20 and recruitment of Ste5 to the Ste4 at the plasma membrane result in the activation of a mitogen-activated protein kinase (MAPK) cascade comprised of a MAPK kinase kinase (Ste11), which phosphorylates and activates a dual specificity MAPK kinase (Ste7), which in turn phosphorylates and activates two related MAPKs (Fus3 and Kss1), leading to new gene transcription, changes in nuclear architecture, polarized cell growth, cell division arrest, and ultimately cell fusion between two different cell types (mating). Signaling ends when GTP is hydrolyzed and G protein subunits reassociate. GTP hydrolysis by GTPase Gpa1 is accelerated by an RGS protein (Sst2).

The pheromone signaling in Saccharomyces cerevisia,has served as an important system for the study of GPCR signaling. Many important mechanisms that regulate the G protein signaling were initially discovered through studying this pathway. Examples include the definitive demonstration of a positive signaling role for G protein β subunits, the first discovery of three-tiered structure of the MAP kinase module, development of the concept of a kinase-scaffold protein, the first demonstration that RGS proteins desensitize the G protein, and the first identification of mono ubiquitination as a signal for receptor endocytosis.