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BIOCHEMISTRY GENERAL MEDICINE HORMONAL REGULATION regulation… ·  · 2008-10-15BIOCHEMISTRY GENERAL MEDICINE HORMONAL REGULATION RNDr. Zden ěk DVO ŘÁK, ... Peptide hormone (natriuretic

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  • BIOCHEMISTRY

    GENERAL MEDICINE

    HORMONAL REGULATION

    RNDr. Zdenk DVOK, PhD.Department of Medical Chemistry and BiochemistryFaculty of Medicine, Palacky University Olomouc

  • HORMONAL REGULATION

    Compounds involved in the co-ordination of metabolic activities of various organsand tissues cellular signalling; signal transduction

    HORMONES NEUROTRANSMITTERS GROWTH FACTORS CYTOKINES

    Extracellular signalling substances Synthesis in one class of cell

    transmission

    TARGET CELLS

    HORMONES Synthetized by specific tissues ENDOCRINE GLANDS Secreted directly into the BLOODSTREAM and carried to their sites of action Specifically alter METABOLIC ACTIVITIES of TARGET CELLS (remotefrom secretory organ) Active at very low concentrations (pM M) Rapidly metabolized SHORT-LIVED EFFECT

  • SIGNAL TRANSDUCTION

    SIGNAL

    ULTIMATECELLULARRESPONSE

    HORMONE RECEPTOR

    TRANSDUCTOR

    EFFECTOR

    SECOND MESSENGER

  • INTEGRATION AND CONTROLOF METABOLIC PROCESSES

    Senzory inputs from the environment

    CENTRAL NERVOUS SYSTEM

    Hypothalamus

    Anterior pituitary Posterior pituitary

    Primary target

    OxytocinVasopresinProlactinFolicule

    Stimulatinghormone

    Luteinizinghormone

    SomatotropinCorticotropinThyrotropin

    Secondary target

    Thyroid Adrenalcortex

    PancreaticIslet cells

    Adrenalmedula

    Ovary Testis

    TestosteroneProgesterone

    EstradiolInsulin

    GlucagonSomatostatin

    CortisolCorticosterone

    Aldosterone

    ThyroxineTriiodo-thyronne

    EpinephrineNor-

    epinephrine

    MusclesLiver

    Manytissues

    LiverMuscles

    Reproductive organsLiverMusclesHeart

    Mammaryglands

    SmoothMuscle;

    Mammaryglands

    Arterioles

    Ultimate target

  • CLASSIFICATION OF HORMONES ACCORDING TO STRUCTURE

    1. Derived from aminoacids - epinephrine (adrenaline)- nor-epinephrine- thyroxine- triiodthyronine epinephrine

    2. Peptides and aminoacids - insulin; glucagon- liberins; oxytocin; vasopresin- ADH; ACTH

    oxytocin

    3. Steroids - cortisol; aldosterone- progesterone; estradiol; testosterone

    estradiol4. Eicosanoids - prostaglandins; leucotrienes

    - prostacyclines; thromboxanes

    arachidonic acid

  • CLASSIFICATION OF HORMONES ACCORDING TO MODE OF ACTION

    1. Hormone DOES NOT TRAVERSE plasma membrane of a target cell amino acids; peptides hydrophylic compounds binding of hormone to the RECEPTOR at the CELL SURFACE hormonal response inside the cell changes in AFFINITY of key proteins/enzymes ACTION THROUGH: - second messengers

    - activation of enzyme activity of cytosolic receptor domain- opening ion channels

    SHORT-TERM action; ULTRA-RAPID response

    2. Hormone TRAVERSES plasma membrane of a target cell steroid and thyroid hormones lipophylic compounds binding of hormone to the INTRACELLULAR RECEPTOR HORMONE-RECEPTOR complex binds to DNA and triggers TRASCRIPTION ofspecific genes changes in LEVEL of key proteins/enzymes LONGTERM action; SLOW response

  • RECEPTORS

    proteins located in plasma membrane or in interior of the cell

    2 binding sites - hormone binding site- a component of signal transduction system

    HORMONE BINDING(saturable; M.-M. kinetics)

    Extensive conformationchange of receptor

    ACTIVATION OF SECONDBINDING SITE

    GRGR

    GRE

    PP

    GRGR

    TR RAR

    Peptides, AA

    steroids

    retinoids

    thyroid

  • HORMONE

    Several classesof receptor

    Identical hormonebinding site

    Different secondbinding site

    Various physiological effects(different tissue)

    ADRENERGIC RECEPTORS1 coupled to phosphatidyl inositol cascade2, 1, 2 coupled to adenylate cyclase cascade

    1 salivary gland K+; H2O secretion2 pancreatic cells secretion

    - muscle glycogenolysis1 heart rate; contraction force

    - adipocyte lipolysis2 liver glycogenolysis

    increase in receptor density in the cell = increased cellular response

  • GTP- binding proteins = G - proteins guanyl-nucleotides proteins TRANSDUCTORS = carriers of excitation signal from RECEPTOR to EFFECTORwithin plasma membrane

    peripheral membrane proteins (located on cytosolic side) TRIMER = it consists of , and subunits subunit is binding site for GDP/GDP; GTPase activity

    large extracellular ligand binding domain formation of hormone-receptor complex conformation change of the receptor transduction of signal inside the cell

    receptor interacts with G-protein exchange of GDP for GTP in -subunit

  • release of -(GTP) subunit activation of effector

    hydrolysis of GTP to GDP release of -GDP from effector re-association with and subunits

  • Inhibition of GTPase activity of -GTP subunit results in irreversibile inactivationof effector and consequently in uncontrolled cellular response

    Example: Vibrio cholerae toxin inhibits GTPase activity in enterocytes that inturn leads to permanent activation of the effector (adenylate cyclase). As theresult; the secretion of Na+ and H2O is uncontrolled and severe dehydratation oforganism is developed.

    G - protein signalling

    G - proteins

    GS - stimulatory

    Gi - inhibitory

    Interaction withdifferent receptors

    Stimulus Receptor G-prot. Effector Response

    Epinephrine -adrenergic r. Gs Adenylate cyclase Glycogen breakdownSerotonin Serotonin r. Gs Adenylate cyclase Behavioral sensitizationLight Rhodopsin Transducin cGMP phosphodiest. Visual excitationIgE-antigen complex Mast cell IgE r. GPLC Phospholipase C Secretionf-Met peptide Chemotactic r. GPLC Phospholipase C ChemotaxisAcetylcholine Muscarinic r. Gk Potassium channel Slowing pacemaker activity

  • SECOND MESSENGERS amplified intracellular signals products of activated effector action small molecules or ions allosteric effectors

    cAMP - cyclic adenosine monophosphate cGMP - cyclic guanosine monophosphate DAG - 1,2-diacylglycerol IP3 - inositol-1,4,5-triphosphate Ca2+ - calcium (free or bound to calmodulin)

    SIGNAL TRANSDUCTION SYSTEMS

    ADENYLATE CYCLASE system PHOSPHATIDYLINOSITOL system TYROSINE KINASE system GUANYLATE CYCLASE system STEROID/THYROID/RETINOID SIGNALLING

  • ADENYLATE CYCLASE (AC) SYSTEM

    AC is activated via G-protein effector = membrane AC

    AC converts ATP to cAMP elevation of cytosolic cAMP cAMP is second messenger

    cAMP action cAMP is allosteric activator of proteinkinases activated proteinkinases phosphorylate target proteins phosphorylated proteins (enzymes) have altered functions cellular response

    degradation of storage fuels HCl secretion by gastric mucosa aggregation of blood platelets

  • Obrzek IX/5

  • ACTIVATION OF PROTEIN KINASE A (PKA) BY cAMP

    heterotetramer dissociates active catalytic subunits of PKA are released

    PKA - 2 catalytic and 2 regulatory subunits cAMP binds to regulatory subunits

    PKA phosphorylates cytosolic proteins PKA may enter nucleus and phosphorylate TFs

    phosphorylated TFs recruit co-activators gene expression is triggered

  • Hormones using cAMP as second messenger

    Calcitonin Chorionic gonadotropin Corticotropin Epinephrine Follicle-stimulating hormone Glucagon Luteinizing hormone Nor-epinephrine Lipotropin Melanocyte-stimulating hormone Parathyroid hormone Thyroid-stimulating hormone Vasopressin

    cAMP signalling

  • GUANYLATE CYCLASE SYSTEMExtracellularside

    Cytosolicside

    PP

    P

    P

    Peptide hormone(natriuretic factor)

    PP

    P

    P

    cellular response e.g. NO synthesis

    GTP

    cGMP

    P

    hydrolysis byphosphodiesterases

    inactivation loss of cellular response

    inhibited by VIAGRA prolonged cell response

  • PHOSPHATIDYL INOSITOL PHOSPHATE (PIP) SYSTEM

    principal molecule in the signalling isphosphatidyl inositol-4,5-bis-phosphate (PIP2)

    membrane receptor trasmits signaland activate effector PI3 kinase

    PI3K phosphorylates PIP2 to PIP3 phosphatidyl inositol-3,4,5-tris-phosphate (PIP3)

  • PIP3 recruits tyrosin kinase (BTK) andphospholipase C (PLC)

    BTK phosphorylates PLC

    activated PLC cleaves PIP2 to 2 fragments 2 second messengers!!!

    IP3 = inositol-1,4,5-triphophate DAG = 1,2-diacylglycerol

  • IP3 binds calcium channel in ER channels open and releases Ca2+ in cytosol

    Ca2+ binds protein kinase C (PKC) PKC then goes to plasma membrane

    PKC is activated when both, i.e.calcium and DAG are bound

  • PKC phosporylates its target proteins

    PIP

    Protein Kinase C

    Important role in controlof cell division and proliferation

    Phosphorylates variety of targetproteins; e.g.: insulin receptor;glucose carrier; CYP P450;tyrosine hydroxylase etc.

    PIP system is activated via variety of stimuli; e.g. ACTH, epinephrine,Neurotransmitters, growth factors, antigens Inactivation of PIP system: IP3 IP2

    PIP system mediates variety of effects; e.g. Glycogenolysis in liver cells;Histamine secretion by mast cells; Serotonine release by blood platelets;Insulin secretion by pancreatic islet cells; Smoth muscle contraction;Epinephrine secretion by adrenal chromaffin cells; visual transduction

  • calmodulin

    Ca2+

    Endoplasmicreticulum

    Ca2+

    Ca2+

    Ca2+ is released from ERin response to hormonesor neurotransmitters

    Ca2+

    calmodulinCa2+

    Ca2+

    Ca2+

    Ca2+Complex

    Calmodulin-Ca2+

    Transinet increase ofintracellular Ca2+ favorsformation of complex

    inactiveenzyme

    calmodulinCa2+

    Ca2+

    Ca2+

    Ca2+activeenzyme

    substrate product

    Calmodulin-Ca2+ complexis an essential componentof many Ca2+ dependentenzymes

  • TYROSINE KINASE (TK) SYSTEM

    membrane receptor contains effector domain effector = tyrosine kinase (TK); protein kinase specific for phosphorylationof tyrosine residue second messenger = O ? x IRS extracellular signals insulin, nerve growth factor; epidermal growth factor

    insulin

    P

    P

    P

    autophosphorylation

    ATP ADP

    catalytic activity of TK switched onby insuline binding enhanced activity independent

    of insuline binding

  • TYROSINE KINASE (TK) SYSTEM

    hormone binds TK = H-R complex = TK activation

    TK phosphorylates target proteins (e.g. phosphatases; amino acidtransporter; glucose transporter etc.) = CELLULAR RESPONSE

    autophosphorylation of TK ensures that TK activity is switched on even inabsence of hormone = long term effect of insulin

    insulin has general growth-promoting properties it acts as metabolic activatorand growth factor almost in all cells in the body it acts

    SHORT-TERM effects = polysaccharide and fat synthesis LONG-TERM effects = nucleic acud and protein synthesis

  • 1

    1

    2

    2

    3

    3

    4

    4

    5

    5

    membrane receptors

    OUTSIDE CELL

    Hormonalsignals

    INSIDECELL

    Cyclic AMP Cyclic GMP Ca2+ DAG Proteins

    G-prot G-prot G-prot G-prot Tyrosinkinase

    secondmessengers

    Nitricoxide

    IP3

    PKA PKG Calmodulin PKC Ser/Thr kinases

    Dedicatedkinases

    Multifunctionalkinases

    Protein substrates Protein substrates

    Protein substratesOther phospholipases

  • steroid/thyroid receptors intracellular localization cytosolic - glucocorticoid; estrogen; androgen; progesterone; mineralocorticoid(undergo nucle-cytosolic traslocation) nuclear retinoid (RARs, RXRs), vitamine D, thyroid

    no second messenger

    DNA is the effector

    binding to DNA triggering gene expression = cellular response enhanced synthesis of specific proteins

    STEROID AND THYROID HORMONES ACTION

    Hormone

    Receptor

    +

    Hormonetraversplasma

    membrane

    Outsideof cell

    cytosol ornucleus

    Nucleartranslocation

    HR complexbinding to DNA

    cellular response

  • Plasma-boundsteroid hormoneFree steroid hormone

    HR

    HR

    HR HR

    hsp

    HR

    HR

    DNA

    HRtranscription

    mRNAmRNA

    translation

    Proteinsynthesis

    NEW PROTEIN

    Biological response

  • TR, VDR and RARs regulation of transcription: Hormone receptor (HR) is dimerizedand bound to DNA at hormone response element site. Without the ligand, transcriptionis inactive due to the interaction of HR with co-repressor. When hormone binds to HR,the bound co-repressor dissociates leading to an interaction between co-activator andHR. These regulatory changes result in increased transcription.

    HR HR

    ligand

    DNA

    RNA transcriptionCo-repressor

    Co-activator

    TranscriptionIniciationkomplex

    Hormone response element

    PROMOTER REGION

  • Hormone class Target organ Target gene

    Glucocorticoids Liver tyrosine aminotransferasetryptophan oxygenase-fetoprotein (down-regulation)metallothionein

    Liver, Retina glutamine synthetaseKidney Phophoenolpyruvate carboxykinase (PEPCK)Oviduct OvalbuminPituitary Pro-opiomelanocortin

    Estrogens Oviduct Ovalbumin; Lysozyme

    Progesterone Oviduct Ovalbumin; OvidinUterus Uteroglobine

    Androgens Prostate AldolaseKidney -glucuronidaseOviduct Albumin

    1,25-dihydroxyvimin D3 Intestine Calcium-binding protein

    Thyroid hormones Liver Carbamoyl phosphyte synthetase; Malic enzymePituitary Growth hormone; Prolactin (down-regulation)

    Target organs and genes for steroid and thyroid hormones

  • HORMONE ACTION AND CARCINOGENESIS

    Alteration of structure of any component in signal transduction system

    Loss of metabolic control in a cell

    Transformation of normal cell

    G-proteins insulin receptor thyroid/steroid hormone receptors

    DAG

    Analogs encoded by specific VIRALONCOGENES = proteins which bindhormone (HR complex) but LACKSWITCH OFF mechanism (lack of GTPaseactivity; permanent TK activity; tightbinding of hormone

    Analogs (PHORBOL ESTERS) = causeactivation of protein kinase C = stimulationof tumor formation (mainly in presenceof carcinogen)

    Loss of metabolic control; uncontrolled cell growth; transformationof a normal cell to a cancer cell