BICD 150 Sp 14 Fortes 2 4 8 14 1 of 9 Lecture 3 GPCR signaling cont 3 main families of subunits G s Increase Adenylyl cyclase turns ATP to cAMP Stimulates phosphorylates Protein Kinase A PKA at a Ser or Thr Phosphorylates other cytosolic proteins and catalytic subunits Can also enter the nucleus and affect gene transcription by phosphorylating CREB G s is seen in all of the adrenergic receptors o PKA is a tetramer w 4 subunits two catalytic and 2 regulatory cAMP binding changes regulatory subunits conformation causing the m to dissociate exposing catalytic subunits o Reaction performed by kinases ATP Protein Protein Phosphate ADP G i inhibitory opposite effects of Gas Decrease AC Decrease cAMP Decrease PKA Seen in 2 receptors G q Activates Phospholipase C Cleaves PIP2 into IP3 and DAG Seen in 1 receptors o IP3 binds to gated Ca channel receptors in the ER to increase Ca Signals smooth muscle contraction and exocytosis of neurotransmitters Ca binds to Calmodulin component of troponin Activation of CaM Kinase phosphorylates proteins and enters nucleus to phosphorylate CREB and increase transcription o DAG activates Protein Kinase C PKC which phosphorylates proteins PKC also needs Ca to be activated Summary adrenergic receptors Gs 2 adrenergic receptors Gi 1 adrenergic receptors Gq How to terminate signaling Dissociating the hormone from the receptor Page 9 of 1 GTPase hydrolyzes GTP bound to the subunit to GDP turning it off after it rebinds subunits o Bacterium Cholera bacteria has an enzyme toxin it secretes which is taken up into cells Catalyzes the reaction G ADPribose G ADPribose inhibiting the GTPase activity causing continual stimulation of the cAMP PKA pathway A gated Cl channel gated by cAMP in the intestinal cells is continuously open releasing Cl Cl attracts Na ions which in turn attract H20 Diarrhea High mortality from dehydration o Pertussis bacteria also has a toxin with similar effects Second messengers can also be dephosphorylated by phosphodiesterases to stop signal pathway cAMP AMP Protein phosphatases dephosphorylate proteins to stop signaling Synthesis and expression of receptors can increase or decrease Receptors can also be removed from the membrane by endocytosis down regulation When there is overstimulation of specifically adrenergic receptor over a period of time PKA will phosphorylate and activate ARK B ARK Phosphorylates the receptor phosphorylated receptor binds the soluble protein Arrestin which stops the receptor from functioning Later a phosphatase will remove the phosphate removes Arrestin reactivates receptor Growth Factor Receptors Insulin and IGFs bind these receptors Single transmembrane segment how does the info of the hormone binding on the outside reach the inside These receptors frequently dimerize after binding their ligand or already exist as dimers On the intracellular portion of these receptors there is at least one Tyrosine residue The receptors have intrinsic tyrosine kinase activity on their intracellular domain After binding the ligand the dimerized receptors will phosphorylate each other The phosphorylated tyrosine will bind proteins w SH2 domains which will attract proteins w SH3 domains that also contain tyrosines This results in the recruitment of other proteins that get phosphorylated that result in cell division and growth Remove ligand tyrosine phosphatases turn off signaling Cytokine receptors Growth hormone EPO Prolactin bind these receptors Used GH receptor specifically as an example Receptors are dimers in the membrane and bind a single GH molecule at different spots Receptors have intracellular tyrosines No intrinsic enzymatic activity Binding GH will cause a change that allows a soluble kinase called JAK to bind and phosphorylate the tyrosines and itself Phosphotyrosines on the receptor can bind another protein called STAT JAK phosphorylates STAT Page 9 of 2 STAT enters the nucleus and stimulates transcription Remove hormone phosphatases end signaling TGF receptors not covered in this class Guanylyl cyclase receptors Natriuretic peptides bind these receptors inhibit Na reabsorption in kidneys regulate pressure in CNS to decrease secretion of ADH Decrease Na and BP They have Guanylyl cyclase activity Synthesizes cGMP Soluble guanylyl cyclase situations Smooth muscle Receptor for cytokines Receptor stimulates NOS to make NO NO will increase synthesis of cGMP cGMP will activate PKG PKG causes Vasodilation by inhibiting contraction Endothelium Receptor for Acetylcholine remember parasympathetic system is responsible for arousal in the genitals or Bradykinin which will increase Ca by binding to specific Gaq receptors Ca stimulates NOS to make NO NO diffuses out of endothelium into the smooth muscle to increase cGMP End signaling Remove hormones NO has a very short half life Phosphodiesterases cleave cGMP Phosphatases remove phosphates from PKG NO derivatives are used as vasodilators to treat angina pectoris chest pain from heart disease Nitroglycerin pills release NO Years ago a drug company made a compound as a dilator to treat angina pectoris It didn t work that well but it was found that patients were frequently getting erections The drug Viagra is a phosphodiesterase inhibitor Increase cGMP Increase PKG Increase vasodilation Erection The signaling pathway upstream of cGMP must still be working for the drug to be effective Photoreceptors in the retina function by activating a Guanylyl cyclase ED drugs can cause visual changes Don t take ED drugs with nitrates Can cause such vasodilation that the BP drops tremendously and fatally Page 9 of 3 Intracellular receptors Hormones must be able to enter the cell steroid hormones Eicosanoids and vitamin D Cytoplasmic receptors Exist in an inactive state with hydrophobic binding sites They would tend to clump together to hide these sites and precipitate Instead the sites are covered by what are known as Heat Shock Proteins HSPs In the rough ER proteins that are destined to be secreted like milk digestive enzymes protein hormones and antibodies are synthesized HSPs prevent mis folding of hydrophobic sites in these proteins Steroid hormone enters the cell binds to receptor cause a conformational change that dissociates the HSP The receptors are then translocated across the nuclear membrane into the nucleus They bind to hormone response elements to alter transcription Most steroid hormones get their changes via changes in protein synthesis Glucocorticoids mineralocorticoids androgens