G protein coupled receptors and their Signaling Mechanism

G Protein Coupled Receptors and their Signaling Mechanisms

1. G Protein Coupled Receptors Faraza Javed Mphil Pharmacology

2. G Protein-Coupled R eceptors G protein-coupled receptors (GPCRs), also known as seven- transmembrane domain receptors, 7TM receptors,  serpentine receptor, and G protein-linked receptors (GPLR), constitute a large protein family of receptors that sense molecules  outside the cell and activate inside signal transduction pathways and ultimately, cellular responses. They are called seven- transmembrane receptors because they pass through the cell membrane seven times.

3. The  ligands  that bind and activate these receptors include: Light sensitive compounds H ormones and   Neurotransmitters That vary in size from small molecules to peptides to large proteins.

4. Families of GPCR 3 F amilies: A – Rhodopsin family B - Secretin /Glucagon receptor family e g . Peptide hormones. C - Metabotropic G lutamate family e g . GABA B , Glutamate.

5. Rhodopsin Receptor Family RLR  are a family of  proteins comprise of  G protein-coupled receptors and are extremely sensitive to light. It  activates the G protein  transducin  ( G t ) to activate the visual phototransduction  pathway . Mutation of the rhodopsin gene is a major contributor to various retinopathies.

6. R emaining receptors are  liganded  by known  E ndogenous   compounds. E xamples include receptor (FXR) farnesoid X receptor, which is activated by bile acid,  liver X receptor (LXR), and  peroxisome proliferator -activated receptor (PPAR ).

7. Secretin R eceptor F amily The secretin -receptor family of GPCRs include  V asoactive intestinal peptide receptors and receptors for  secretin ,  calcitonin   and parathyroid hormone/parathyroid hormone-related peptides. These receptors activate  adenylyl cyclase  and the  phosphatidyl - inositol -calcium pathway.

8. Metabotropic Glutamate F amily The metabotropic glutamate receptors ( mGluRs ) are family C GPCR that participate in the modulation of synaptic transmission and neuronal excitability throughout the central nervous system . They have been subdivided into three groups, based on intracellular signalling mechanisms. Group I mGlu receptors (coupled to PLC and intracellular calcium signalling ).

9. G roup II Group III receptors are negatively coupled to adenylyl cyclase . These receptors are generally widely distributed throughout the mammalian brain with high levels in the cerebellum and thalamus.  

10. Structure of G Protein G proteins , also known as  guanine nucleotide-binding proteins , involved in transmitting signals and function as molecular switches. Their activity is regulated by factors that control their ability to bind to and hydrolyze  guanosine triphosphate  (GTP) to  guanosine diphosphate  (GDP). When they bind GTP, they are 'on', and, when they bind GDP, they are 'off ' .

11. G protein complexes are Made up of  alpha  (α),  beta  (β) and gamma  (γ) subunits . Beta and gamma subunits can form a stable dimeric complex referred to as the  beta-gamma complex.

12. G proteins located within the cell are activated by   GPCRs that span the cell membrane. Inside the cell, on the plasma membrane, G Protein binds GDP when inactive and GTP when active. When the GPCRs binds to a signal molecule, the receptor is activated and changes shape, thereby allowing it to bind to an inactive G Protein. When this occurs, GTP displaces GDP which activates the G Protein .

13. The newly activated G Protein then migrates along the cell membrane until it binds to adenylyl cyclase which convert ATP to cAMP that leads to the next step in the pathway and generates a cellular response . After transduction, G Protein functions as a GTPase and hydrolyzes the bound GTP which causes a phosphate group to fall off. This regenerates GDP and inactivates the G Protein and the cycle repeats.

14. G Protein Mediated Pathways Secondary messenger Systems Involved In Signal Transduction: Adenylate cyclase cAMP mediated pathway P hospholipase mediated pathway

15. cAMP Mediated P athway T he   cAMP -dependent pathway , also known as the  adenylyl cyclase  pathway , is a G protein-coupled receptor triggered  signaling cascade used in cell communication . When a GPCR is activated by its extracellular ligand , a conformational change is induced in the receptor that is transmitted to an attached intracellular  heterotrimeric G protein complex.

16. The  G s  alpha subunit  of the stimulated G protein complex exchanges   GDP  for GTP and is released from the complex . In a cAMP -dependent pathway, the activated G s  alpha subunit binds to and activates an enzyme called  adenylyl cyclase , which, in turn, catalyzes the conversion of ATP  into ( cAMP ). G s cAMP Dependent Pathway

17. Increases in concentration of the second messenger  cAMP may lead to the activation of an enzyme called protein kinase A (PKA ). The PKA enzyme is also known as cAMP -dependent enzyme because it gets activated only if cAMP is present. Many different cell responses are mediated by cAMP . These include increase in heart rate, cortisol secretion, and breakdown of glycogen and fat.

18. GTP GDP  GDP GTP  ATP cAMP Cell response AT Protein kinase ADP P Inactive protein Active protein hormone Adenylate cyclase Signaling System AC R S Inhibitor R i  

19. This pathway can:   A ctivate enzymes and  R egulate gene expression If cAMP -dependent pathway is not controlled, it can ultimately lead to hyper-proliferation, which may contribute to the development and/or progression of cancer .

20. Alterations in number, structure or function of receptors will lead to disorder in cellular signal transduction. Up-regulation/hypersensitivity Down-regulation/desensitization Receptor Gene Mutation

21. Hyperthyroidism Hyperthyroidism , often called  overactive thyroid , is a condition in which the thyroid gland produces  and secretes excessive amounts of the  thyroid hormones T3 and/or T4 .   Grave disease is the most common cause of hyperthyroidism.

22. Mechanism: The  thyrotropin receptor   ( TSH receptor ) responds to thyroid-stimulating hormone and stimulates the production of  thyroxine  (T4) and  triiodothyronine  (T3). The TSH receptor is a member of the G protein-coupled receptor  and is coupled to the G s  protein . Mutation in TSHR gene (chromosome 14q31) lead to the hyperactivation of cAMP pathway results in hyperactivation of gland and make progress towards the development of tumor.

23. Treatment: Antithyroid Medicine including Propylthiouracil , Methimazole and Carbimazole . Radioactive Iodine

24. Cholera Toxin Cholera  is an infection of the small intestine caused by the bacterium  Vibrio cholerae . Mechanism:    When cholera toxin is released from the bacteria in the infected intestine, it binds to the intestinal cells known as enterocytes . T oxin enters, where it activates the G protein G s  through an ADP- ribosylation reaction that acts to lock the G protein in its GTP-bound form, thereby continually stimulating adenylate cyclase to produce cAMP .

25. Increased G s  activation leads to increased  adenylate cyclase activity , which increases the intracellular concentration of cAMP to more than 100-fold over normal and over-activates cytosolic   PKA. These active PKA then phosphorylate the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel proteins, which leads to ATP-mediated efflux of chloride ions and leads to secretion of H 2 O,  Na + ,K + , and HCO 3 -  into the intestinal lumen.

26.   In addition, the entry of Na +  and consequently the entry of water into enterocytes are diminished. The combined effects result in rapid fluid loss from the intestine, leading to severe  dehydration.

27. G-protein modification — cholera lumen of intestine Gs CT AC cAMP ↑ ↑ ↑ Cl - H 2 O Na + CT--Cholera toxin Gs  ribosylation

28. Treatment: Rehydration. The goal is to replace lost fluids and electrolytes using a simple rehydration solution, oral rehydration salts (ORS). Intravenous fluids . Antibiotics . Zinc supplements.

29. G i cAMP Dependent Pathway G i  mainly inhibits the  cAMP dependent pathway by inhibiting adenylate cyclase activity, decreasing the production of  cAMP   from ATP , which, in turn, results in decreased activity of  cAMP -dependent protein kinase . Therefore, the ultimate effect of G i  is the opposite of  cAMP -dependent protein kinase .

30. When G i receptors get activated, they  release activated G-protein  βγ- subunits from inactive  heterotrimeric G protein   complexes. G βγ   dimeric protein interacts with GIRK channels to open them so that they become permeable to potassium ions, resulting in hyperpolarization of the cell. These receptors are primarily found on heart as well as in brain.

31. Atrial fibrillation  (abnormal heart rhythm) is associated with shorter action potential duration and believed to be affected by the G protein-gated K +  channel, I K,Ach . The I K,ACh channel , when activated by G proteins, allows the flow of K +  across the plasma membrane and out of the cell. This current hyperpolarizes the cell, thus terminating the action potential.

32. In chronic atrial fibrillation there is an increase in this inwardly rectifying current because of constantly activated I K,ACh  channels. Increase in the current results in shorter action potential duration experienced in chronic atrial fibrillation and leads to the subsequent fibrillating of the cardiac muscle. Blocking I K,ACh  channel activity could be a therapeutic target in atrial fibrillation and is an area under study.

33. Opioids are prescribed to treat chronic pain in different diseases, GIRK channels are activated by certain GPC opioid receptors, which leads to the inhibition of nociceptive transmission, thus functioning in pain relief.  Studies have shown that G proteins directly activate GIRKs which were found to participate in propagation of morphine-induced analgesia in inflamed spines of mice. Research pertaining to chronic pain management continues to be performed in this field.

34. GPC Receptors G Protein Receptors Signaling Pathway G S Beta adrenergic receptors, glucagon, histamine, serotonin Increase Adenylyl cyclase CAMP Excitatory effects G i Alpha 2 adrenergic receptors, mAchR , opioid , serotonin Decrease Adenylyl cyclase CAMP Cardiac K + channel open- decrease heart rate G q mAchR , serotonin 5HT 1C PLC- IP 3 , DAG Increase Cytoplasmic Ca

35. G t Rhodopsin and colour opsins in retinal rod and cone cells Increase cGMP phosphodiesterase . Decrease cGMP

36. Gq Protein C oupled R eceptor G q protein is a heterotrimeric protein subunit that activates phospholipase C (PLC). PLC in turn hydrolyzes Phosphatidylinositol 4,5-bisphosphate (PIP 2 ) to diacyl glycerol (DAG) and inositol trisphosphate (IP 3 ) signal transduction pathway. DAG acts as a second messenger that activates Protein Kinase C (PKC) and IP 3 acts on calcium channels to release calcium from stores and phosphorylation of some proteins.

37. Cell Signaling Pathway: Activation of PKC through G protein coupled receptor

38. Receptors that are Gq protein coupled include : 5-HT 2 serotonergic receptors Alpha-1 adrenergic receptor Vasopressin type 1 receptors: 1A and 1B Angiotensin II receptor type 1 Histamine H1 receptor Metabotropic glutamate receptor, Group I M 1 , M 3 , and M 5 muscarinic receptors

39. Clinical Significance Ligands targeting the mAChR that are currently approved for clinical use include non-selective antagonists for the treatment of Parkinson's disease, atropine (to dilate the pupil), Scopolamine (used to prevent motion sickness), and ipratropium (used in the treatment of COPD ).

40. Pilocarpine can be given in glaucoma because it reduces intraocular pressure by contraction of the ciliary muscle, opening the trabecular meshwork and allowing increased outflow of the aqueous humour

41. Gt Protein C oupled R eceptors Gt protein coupled receptors are found in photoreceptos (rods and cons) of the eye. Photoreceptors are light sensitive and responsible for visual phototransduction process. These encode a light stimulus as a chemical output .

42. Photoreceptor Cells Two types of photoreceptors: rods and cones Rods are very sensitive cells specialized for night vision. In bright light conditions the response of the rods is saturated and cones, faster but less sensitive photoreceptors , mediate day vision.

43. Phototransduction Light activates the opsin molecules in the photoreceptors ( rhodopsin ). Upon activation becomes metarhodopsin II. Metarhodopsin II activates transducin , a Gt protein. GDP-bound inactive transducin will exchange GDP for GTP. GTP-bound active transducin will increase the activity of cGMP phosphodiesterase . The result is decreased levels of cGMP in the cytoplasm.

44. Decreased levels of cGMP cause the closing of cGMP -gated ion channels which will lead to membrane hyperpolarization .

45. Disorders of Phototransduction Bradyopsia (or ‘slow vision’) is a condition that results from mutations in genes encoding the transducin -inactivating protein RGS9 or the RGS9 anchor protein (R9AP). This protein inactivates transducin during light termination process. Patients with bradyopsia have trouble adjusting to changing light conditions, experiencing a temporary blindness when first exposed to bright light.

46. Congenital Stationary N ight B lindness is an inherited disorder that affects rod photoreceptors and impairs vision under low-light conditions. This disorder may result from missense mutations in the rhodopsin gene that cause the mutated rhodopsin protein to constitutively activate transducin . Persistent activation of the phototransduction cascade limits the fidelity of the light response by rod photoreceptors.

47. Retinitis Pigmentosa is an inherited disorder characterized by degeneration of photoreceptor cells and accumulation of retinal pigments. This disorder, which often leads to blindness, can result from mutations in a variety of genes expressed in photoreceptors.

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