05-07 Signalling.pdf

Intracellular Signaling pathways Dr. Hader I. Sakr Associate Professor, Medical Physiology  ‫ﺴﻢ ﷲ اﻟﺮﲪﻦ اﻟﺮﺣﲓ‬ ‫﴿ َواﺗ ُﻘﻮا ا َ ۖ َوﯾ ُ َﻌ ُ ُ َ ُ‬ ‫ا‬‫و‬ ‫ۗ‬ ‫ا‬ ‫ُ‬ ‫ﲂ‬ ‫ُ‬ ‫ﻤ‬‫ّ‬ ‫ِ‬ ‫ﻠ‬ ‫ﳾ ٍء َ ِﻠ ٌﲓ﴾‬ ‫ِ ُ ِّ‬ ‫ﲁ َْ‬ Learning ob...

Intracellular Signaling pathways Dr. Hader I. Sakr Associate Professor, Medical Physiology  ‫ﺴﻢ ﷲ اﻟﺮﲪﻦ اﻟﺮﺣﲓ‬ ‫﴿ َواﺗ ُﻘﻮا ا َ ۖ َوﯾ ُ َﻌ ُ ُ َ ُ‬ ‫ا‬‫و‬ ‫ۗ‬ ‫ا‬ ‫ُ‬ ‫ﲂ‬ ‫ُ‬ ‫ﻤ‬‫ّ‬ ‫ِ‬ ‫ﻠ‬ ‫ﳾ ٍء َ ِﻠ ٌﲓ﴾‬ ‫ِ ُ ِّ‬ ‫ﲁ َْ‬ Learning objectives: Regulatory systems. Endocrine glands. Hormones characteristics and nature. Hormones receptors structure, distribution and ligands. Hormones receptors intracellular signaling pathways. Regulatory systems Regulatory systems Regulatory systems The endocrine system: The nervous system Structural unit Ductless (endocrine glands) Neurons and neuroglia Functional unit chemical messengers (hormones) Reflex arc Onset Rapid, moderate or slow Rapid Duration Short or Prolonged Short Example Metabolic functions Muscular contractions Endocrine glands.  Endocrine glands  The endocrine system: - Endocrine glands constitute a slow control system of the body. - They produce their actions by secreting hormones. Co-ordination of Body Functions by Chemical Messengers: 1. Endocrine hormones. 2. Paracrine. 3. Autocrine. 4. Neurotransmitters: (rapid mechanism). 5. Neuroendocrine hormones: (intermediate mechanism). 6. Gap Junctions. Endocrine glands  Endocrine glands  Pituitary gland which is the master gland.  The pituitary is under the control of the Hypothalamus (neuro-endocrine).  Three endocrine glands under the control of pituitary gland: 1. Thyroid gland. 2. Suprarenal cortex. 3. Gonads (testis & ovaries)*.  Three endocrine glands not under the control of pituitary gland: 1. Parathyroid glands. 2. Suprarenal medulla. 3. Pancreas*.  Endocrine glands  In addition, there are organs which possess endocrine function together with their non- endocrine function: - The heart secretes atrial natriuretic peptide (ANP). - The kidney secretes erythropoietin, 1,25(OH) 2 cholecalciferol, renin among others. - The liver secretes somatomedines, 25 hydroxychole- calciferol & hepcidin. - The skin secretes calciferol (vitamin D). - The gastrointestinal tract secretes GIT hormones as gastrin, CCK, secretin, VIP….. etc. - The pineal gland secretes melatonin. Hormones characteristics and nature.  Hormones characteristics and nature  Physiological Properties of Hormones 1. Produced in small amounts (μg to ng). 2. The rate of secretion is determined by the body needs. 3. Super-added rhythms: - Circadian rhythm (i.e., daily rhythm), e.g. ACTH. - Monthly rhythm, e.g. female gonadotropic hormone. 4. They may affect various tissues (e.g. insulin) or specific target organs (e.g. sex hormones). 5. Some hormones antagonize the effect of others (e.g. insulin and glucagon). 6. They may trigger substances initiating biochemical reactions that persist after the disappearance of the hormone.  Hormones characteristics and nature Inter-relation between endocrine and nervous systems  Hormones characteristics and nature  The nervous system can affect the endocrine system: 1. Hypothalamo-hypophyseal portal circulation: Between the hypothalamus (nervous system) and the anterior pituitary gland. Releasing or inhibiting hormones are released from the hypothalamus to alter the secretion of the different anterior pituitary hormones. 2. Hypothalamo-hypophyseal tract: between the hypothalamus and the posterior pituitary. The ADH and oxytocin hormones are formed in the hypothalamus. They are stored and secreted by their nerve endings present in the posterior pituitary. 3. Direct connection between hypothalamus and suprarenal medulla: Affects the secretion of epinephrine & norepinephrine.  Hormones characteristics and nature  On the other hand, the endocrine system can also affect the nervous system: - Parathyroid glands control Ca+2 level in blood that affects the nerves excitability. - Thyroid hormones affect the excitability of the nervous system.  Hormones characteristics and nature  Chemical Nature of Hormones:  Hormones characteristics and nature  Chemical Nature of Hormones: a) Steroid hormones: secreted by the adrenal cortex (cortisol and aldosterone), the ovaries (estrogen & progesterone), the testes (testosterone) and the placenta (estrogen & progesterone). b) Derivatives of the amino acid: tyrosine, secreted by the thyroid (thyroxine and tri- iodothyronine) and the adrenal medullae (dopamine, epinephrine and norepinephrine). c) Protein and polypeptide hormones: including hormones secreted by the pituitary gland, the pancreas (insulin, glucagon & somatostatin), the parathyroid gland (parathyroid hormone), and many others. Hormones receptors structure, distribution and ligands Hormones receptors structure, distribution & ligands  Mechanism of Action of Hormones - The first step of a hormone's action is to bind to specific receptors at the target cell. Cells that lack receptors for the hormones do not respond. - The locations for the different types of hormone receptors are generally the following: 1. In or on the surface of the cell membrane: Specific mostly for the protein, peptide, and catecholamine hormones. Hormones receptors structure, distribution & ligands 2. In the cell cytoplasm: The primary receptors for the different steroid hormones. 3. In the cell nucleus: Receptors for the thyroid hormones are found in the nucleus and are located in direct association with one or more of the chromosomes. Hormones are called "primary messengers", while the chemical substance that results inside the cell from the combination of hormone and receptor is known as the "second messengers". Hormones receptors structure, distribution & ligands  Characters of cell receptors: - The number of receptors in a target cell does not remain constant. - Increased hormone concentration and increased binding with its target cell receptors cause the number of receptors to decrease. - This is known as down regulation that decreases the target tissue's responsiveness to the hormone. Hormones receptors structure, distribution & ligands - Down-regulation can occur through: 1. Decreased production of the receptors. 2. Temporary sequestration of the receptor to the inside of the cell, away from the site of action of hormones. 3. Destruction of the receptors by lysosomes after they are internalized. 4. Inactivation of some receptor molecules. 5. Inactivation of some intracellular protein signaling molecules. Hormones receptors structure, distribution & ligands  Up-regulation of receptors: - It is increased cell response 2ry to prolonged exposure to decreased amounts of hormone. - The target tissue becomes progressively more sensitive to the stimulating effects of the hormone - This can happen either by increase in receptor expression or decrease in rate of internalization of receptors. Hormones receptors intracellular signaling pathways Hormones receptors intracellular signaling pathways Trans- Cytoplasmic membrane Channels G-Protein Altering Altering Transcription Opening activation enzyme enzyme modulation Hormones receptors intracellular signaling pathways  The mechanisms by which chemical messengers exert their IC effects may be through: A. Opening of certain ion channels (trans-membrane). B. G-Protein activation (trans-membrane). C. Altering enzyme activity within the cell (trans-membrane or cytoplasmic). D. Direct activation of transcription. The resulting cell-signaling pathways provide: 1. Amplification (same) and distribution (different) of the effect. 2. Regulation and feedback control. Hormones receptors intracellular signaling pathways 1-3. Trans-membrane receptors  For lipid insoluble hormones (protein & peptide, catecholamines, serotonin, melatonin).  In or on the surface of the cell membrane.  Combination of the hormone with the cell membrane receptors will regulate the activity of target proteins (enzymes or ion channels): o Directly o Through coupling with G proteins. Hormones receptors intracellular signaling pathways A. Opening of certain ion channels. a, In tetrameric (voltage-gated) K+ channels, a single pore is formed by 4 identical or structurally similar proteins (subunits). b, In pentameric 'ligand-gated' cation (K+) or anion (Cl-) channels (acetylcholine or GABA), the single pore is formed by 5 identical or structurally similar subunits. c, Cl- channels from the CLC family are dimers, in which each subunit has its own pore. d, Aquaporin water channels are tetramers, again with one pore per subunit. Hormones receptors intracellular signaling pathways B. G- Protein activation (G-protein coupled receptors (GPCR):  Termed so because of their ability to bind guanosine nucleotide.  When the activating signal reaches the GPCR, they exchange GDP for GTP.  The GTP-protein complex brought about the effect.  Inherent GTPase activity of the protein converts GTP to GDP restoring the inactive resting state.  Occur in two large families: 1. Small G proteins. 2. Heterotrimeric G proteins (1st identified). Hormones receptors intracellular signaling pathways  Heterotrimeric G proteins - Couple surface receptor (GPCR) to ion channel or intracellular catalytic unit forming 2nd messenger. - Over 1000 receptors are coupled; each has 7 trans-membrane loops. - Activation of a single GPCR can result in 1, 10 or more active G proteins (amplification). - Made up of 3 subunits: α, β & γ. - Subunit α subunits are divided into three subfamilies: s, I and q, and a single GPCR can couple to either one or more families Gαs Gαi Gαq ↑ Adenyl cyclase ↓ Phospholipase Cβ Hormones receptors intracellular signaling pathways  Each G protein activates several downstream effectors: - Gαs stimulates adenylyl cyclase and increases levels of cAMP. - Gαi inhibits adenylyl cyclase and lowers cAMP levels. - Gαq family bind to and activate phospholipase C (PLC).  The Gβγ subunits dimer activates many signaling molecules, including phospholipases, ion channels and lipid kinases. Hormones receptors intracellular signaling pathways C. Enzyme-Linked Hormone Receptors: I II  When activated, they function directly as enzymes (I, part of the transmembrane receptor) or are closely associated with enzymes (II, the H-R complex activates cytoplasmic enzyme).  They have their hormone-binding site on the outside of the cell membrane and their catalytic (inherited) or enzyme-binding (cytoplasmic) site on the inside. Hormones receptors intracellular signaling pathways 2. Cytoplasmic receptors (Steroids, Lipid-soluble) Hormones receptors intracellular signaling pathways  They can cross the cell membrane (lipid-soluble) and interact with cytoplasmic (or nuclear) receptors rather than in the cell membrane.  The activated H-R complex then binds with the hormone response element (HRE) within a specific gene regulatory sequence (promoter), altering its transcription & mRNA formation.  There are 3 distinct pathways for 1ry messenger to alter transcription: 1. Activation of cytoplasmic latent transcription factor. 2. Activation of cytoplasmic protein kinase cascade  migrate to the nucleus  phosphorylate a nuclear latent transcription factor. 3. Binding to cytoplasmic or nuclear receptor. Different 2nd messenger systems Hormones receptors intracellular signaling pathways a) Intracellular Ca+2 as 2nd messenger:  Cytoplasmic Ca+2 can increase by: o Entrance through membrane channels (external source): 1. Voltage-gated. 2- Ligand-gated. 3- Stretch-gated. 4. SOCCs (store operated Ca+2 channels) where Ca+2 from depleted stores triggers their opening. o Release from internal stores (SR): 1. IP3 receptors (ligand-gated channels) on SR. 2. CICR (Ca+2-induced Ca+2 release) through foot processes opening of SR terminal cisternae (Ligand-gated DHPRs). 3. Voltage-gated DHPRs. Hormones receptors intracellular signaling pathways  Increased cytoplasmic Ca+2 binds to and activates calcium-binding proteins: 1. Troponin C  Muscle contraction. 2. Calmodulin  activates different kinases. 3. Calbindins  GIT. Hormones receptors intracellular signaling pathways b) Phospholipase C (PLC)  IP3 & DAG as 2nd messengers:  PLCβ is activated through Gαq.  PLCγ is tyrosine kinase-activated.  This enzyme catalyzes the breakdown of phosphatidyl inositol-di-phosphate (PIP2) into: 1. Inositol-tri-phosphate (IP3). 2.Di-acyl glycerol (DAG).  The IP3 mobilizes Ca+2 from mitochondria and ER  elevating cytoplasmic [Ca+2].  DAG stays in the cell membrane activating PKC  phosphorylates a large number of proteins  cell response. Hormones receptors intracellular signaling pathways  In addition, the lipid portion of DAG is the arachidonic acid, the precursor for PGs and other local hormones causing multiple effects in tissues throughout the body.  Hormones acting through PLC system: 1. Angiotensin-II (vascular smooth muscle). 2. Catecholamines (α1 receptors). 3. Vasopressin (V1 receptor) 4. Oxytocin. 5. GHRH. 6. TRH. 7. GnRH. 8. Muscarinic 1, 3 and 5. Hormones receptors intracellular signaling pathways c) cAMP as 2nd messenger: Adenylyl cyclase  Produced from ATP 3,5’cAMP Phosphodiestrase 5’AMP  Phosphodiestrase is inhibited by methylxanthines (caffeine and theophylline).  cAMP activates PKA  protein phosphorylation.  The active catalytic subunit of PKA moves to the nucleus  phosphorylating CREB (cAMP-responsive element- binding protein)  altering different genes transcription. Hormones receptors intracellular signaling pathways  Adenylyl cyclase activity can be altered through: 1. Gαs or Gαi subunits. 2. Cholera toxin (contaminated drinking water): inhibits Gαs GTPase activity prolonging its stimulatory effect  ↑ adenyl cyclase activity  ↑ intestinal cAMP  ↑ intestinal motility  watery diarrhea  dehydration. 3. Pertussis toxin: inhibits Gαi by preventing GDP displacement by GTP  whooping cough, histamine hypersensitivity, and lowered blood glucose level. 4. The drug forskolin (anti-asthmatic): by direct stimulatory effect. 5. PKA and PKC (directly or indirectly). 6. Calmodulin. Hormones receptors intracellular signaling pathways  Hormones acting through cAMP system: 1. Angiotensin-II (vascular smooth muscle). 2.PGE1 & Catecholamines (α2, β receptors). 3. Vasopressin (V2receptor). 4. FSH, LH 5. ACTH. 6. TSH 7. HCG 8.PTH. 9. Calcitonin. 10.Muscarinic 2,4. 11. Somatostatin & Secretin. 12. Glucagon. Hormones receptors intracellular signaling pathways d) cGMP as 2nd messenger:  Exists in 2 forms: o Trans-cellular: e.g.: for ANP and for gastro-intestinal polypeptide. o Intracellular (soluble, heme-containing): activated by NO and NO-containing compounds.  Important in: 1. Vision in both rods and cone cells. 2. cGMP-activated channels. 3. cGMP-dependent kinase (PKG). Hormones receptors intracellular signaling pathways e) Kinase/Phosphatase 2nd messenger system:  Phosphorylation of the target’s tyrosine, serine or threonine residues.  Tyrosine kinase (TKase): o The receptor itself may have intrinsic TKase (membrane-bound) or the hormone-receptor complex (H-R) may activate cytoplasmic TKase. MLCK (smooth muscle contraction) Phosphorylase kinase CaM kinase I (synaptic) Calmodulin-dependant CaM kinase II (synaptic & Ca+2-ATPase) Serine or CaM kinase III (protein synthesis) threonine kinases CaM kinase IV PLC  PIP2 dependent PKC (7 subspecies) cAMP-dependent kinases (PKA) Cyclic nucleotide-dependent cGMP-dependent kinases (PKG) Hormones receptors intracellular signaling pathways e) Kinase/Phosphatase 2nd messenger system: - Intrinsic (trans-membrane): Tyrosine o Insulin, IGF-1, EGF, PDGF Macrophage-CSF receptors kinases - H-R  cytoplasmic: o GH, prolactin, erythropoietin (cytoplasmic) Conclusion There are different methods of chemical transmission between cells The endocrine system is one of the regulatory systems of the body. Human body contains several endocrine glands. Both the endocrine and nervous systems are interconnected. Hormones have different characteristics. Hormones may be protein, polypeptides, steroids or Tyrosine derivatives. Hormone receptors may be trans-membrane, cytoplasmic or nuclear. Conclusion Trans-membrane receptors act through ion channels, GPCR or enzyme-linked. Ion channels have different configurations. GPCRs are classified according to the G-protein into small and heterotrimeric. Hormone-activated enzymes may be part of the receptor or cytoplasmic. 2nd messengers may be Ca+2, PLC, cAMP, cGMP, or different kinases. References Guyton and Hall, 13th edition. Unit II(6). Ganong’s review of medical physiology 25th ed. Section I(5). Thank You