SIGNAL TRANSDUCTION AND MECHANISM OF HORMONE

ACTION

Dr.S.Chakravarty, MD

Learning objectives • List the various types of receptors in the cells

• Differentiate the features of water and fat soluble hormones

• List the various second messengers involved in signal transduction

• Describe the G protein and its coupled receptor in detail

• Differentiate intrinsic and extrinsic tyrosine kinase mediated pathways of signal transduction

• Describe the mechanism of insulin release and the structure of insulin receptor

Types of signalling

Receptors for various hormones

• Intracellular (transcription factors) – type 1 hormones

• Cell surface – type 2 hormones

Types of hormones based on solubility

Effect Days Minutes to hours

Type -1 hormones –intracellular receptors

Type 2 hormones- have membrane receptors

Growth factors!

G protein coupled receptor:

• Primary receptor for most of type 2 hormones.

• Link receptors to second messengers

• Serpentine -7 channel transmembrane receptor

• Coupled to G proteins.

• 4 types of G proteins – Gs, Gi, Gq and G12

• 3 heterotrimers of G proteins – αβγ subunits.

GPCR – Plasma membrane coupled receptor

G-α β

γ ATP

C -AMPProtein Kinase A

Enzyme Enzyme

P

CREB

CREB

P

CRE

GDP(Inactive)

Adenylate Cyclase

Cell membrane

Cytosol

Plasma

G – protein coupled Receptor cAMP mediated pathway

GTP(active)

cAMP response element binding protein

cAMP response element

Second messenger is cAMP

c-AMP pathway cont..

PKA is also known as cAMP-dependent protein kinase

Mechanism of action of c-AMP on protein kinase-A

PKA is also known as cAMP-dependent protein kinase

Hydrolysis of c-AMP by Phosphodiesterase

c-AMP 5’-AMP

Active Inactive

H2O

Phosphodiesterase

(-)

Methyl Xanthines(Caffeine, Theophylline,

Theobromine)

Various types of G-proteins

Classes and Functions of Selected G Proteins1

Class or Type Stimulus Effector Effect

Gs

αs

αOlf

Glucagon, α-adrenergics

Odorant

Adenylyl cyclaseCardiac Ca2+, Cl–, and Na+ channelsAdenylyl cyclase

Glyconeogenesis, lipolysis, glycogenolysis

Olfaction

Gi

αi-1,2,3

α0

αt

Acetylcholine, α2-adrenergicsM2 cholinergicsOpioids, endorphins

Light

Adenylyl cyclasePotassium channelsCalcium channelsPotassium channelscGMP phosphodiesterase

Slowed heart rateNeuronal electrical activity

Vision

Gq

αq

α11

M1 cholinergicsα1-Adrenergicsα1-Adrenergics

Phospholipase C-β1

Phospholipase C-β2

Decreased Muscle contractionand decreased Blood pressure

G12

α12Thrombin Rho Cell shape changes

Table 42–2. Subclassification of Group II.A Hormones

HORMONES THAT STIMULATE ADENYLYL CYCLASE

HORMONES THAT INHIBIT ADENYLYL CYCLASE

ACTH Acetylcholine

ADH α2-Adrenergics

α1-Adrenergics Angiotensin II

Calcitonin Somatostatin

CRHFSH

GlucagonhCGLH

LPHMSHPTHTSH

Protein Phosphorylation by protein kinases

• Serine / threonine phosphorylation:

• Protein kinase A and C and G attach phosphate groups to hydroxyl groups of serine and threonine – to phosphorylate proteins

Type- 3 Second messenger as Calcium and phosphatidyl inositol

Calcium is required for : • Skeletal muscle contraction

• Blood clotting

• Secretion of various hormones and enzymes

• Membrane excitability

• Enzymatic activity.

IMP

Action of phospholipase C

Important aspect in Metabolic regulation

Enzyme Enzyme P

Protein kinase

Phosphatases

ATP ADP

H2O

GlucagonEpinephrine

Insulin P

ADP RIBOSYLATION OF PROTEINS

ADP-ribosyltransferase

ADP-ribosyltransferases

ADP RIBOSYLATION OF PROTEINS

• Cholera toxin – causes ADP ribosylation of GαS

& subunit and inhibits intrinsic GTPase activity – prolonged action of adenylate cyclase – watery diarrhea

• Pertussis toxin – causes ADP ribosylation of G α i-2 and prevents dissociation of α i-2 from βγ subunit .Gαs activity in such cells unopposed.

Usmle!

Two more toxins for USMLE !

• Diphtheria toxin – causes ADP ribosylation of eEF-2 and inhibits protein synthesis by inhibiting translocation of ribosomes

• Tetrodotoxin – is a neurotoxin which acts on fast sodium channels in the neuronal membrane and inhibit action potential in the neurons.(pufferfish, porcupinefish, ocean sunfish or mola, and

triggerfish, )Usmle!

membrane bound Guanyl

cyclase

GTP C-GMP GTP

Protein kinase G

Vasodilation (smooth muscle)

NO ANF Cyclic GMP pathway

Soluble Guanyl cyclase

5’-GMP

Phosphodiesterase

Anti anginal Drugs

+

Guanyl cyclase – cGMP (class II)

• Membrane bound Guanyl cyclase – Atrial natriuretic peptide (ANP).

• Soluble Guanyl cyclase – nitrates, nitroprusside, NO.

• Sildenafil (Viagra) – phosphodiesterase V inhibitor

• Remember :-Theophylline, caffeine – phosphodiesterase inhibitors.

Drugs acting through NO pathway

• Sodium nitroprusside – arterial and venodilator

• Nitroglycerine – veins • Hydralazine – arterial

• Artery – decreased TPR and BP• Vein – decreased preload and cardiac output

Receptor tyrosine kinase Insulin and growth factors

Tyrosine kinases – phosphorylate proteins at hydroxyl groups of tyrosine amino acid

With intrinsic tyrosine kinase activity

• Insulin receptor• IGF -1 • EGF, PDGF, FGF

• MAP kinase RAS protein

Without intrinsic tyrosine kinase

• GH, Prolactin • Erythropoietin • Cytokines

• Janus kinase STAT (signal transducers and

activators of transcription)

I

Cell membrane

Beta subunit

Alpha subunit

GLUT-4

G

GG

GI

Phosphorylation of Insulin Receptor substrates

GLUT-4

G

G

G

PHOSPHORYLATION OF TYROSINE RESIDUES

DIFFERENT PATHWAYS

One Cause for insulin resistance

Serine phosphorylation instead of tyrosine phosphorylation of insulin receptor– mediated by inflammatory mediators like TNF-alpha, interleukins, free fatty acids etc.

Translocation of GLUT-4 receptor

• Insulin dependent translocation

• Well fed state/abundant glucose in blood

• PI-3 kinase pathway

• Exercise induced translocation:

• During exercise – no insulin

• AMPK pathway

Pathway G-protein Enzyme Second messenger

Protein kinase

C-AMP Gs or Gi Adenylate cyclase

C-AMP PK –A

C-GMP Single channel Guanyl cyclase C-GMP PK - G

PIP-2 Gq Phospholipase C IP-3, Ca2+ and DAG

PK - C

Insulin and GFs IRS and SH2 domains

•Tyrosine kinase •Protein phosphatase

Mechanism of action of steroid hormones

SRESteroid receptor

HSP 90Steroid hormone( Cortisol)

In the absence of the steroid hormone cortisol, Glucocorticoid receptor resides in the cytosol complexed with several chaperone proteins including Hsp90

Foye’s Medicinal Chemistry, 7th Edition by Lemke

Nf-kb pathway

p50 p65

Ikb

Cytokines, TNF a, free radicals, viruses

IKKcomplex

P +

Nucleus

Ubiquitin proteosome pathway

Steroids inhibit this pathway and act as potent anti-inflammatory agents

(Inhibitor of Κb)

IκB kinase

Very large family of Nuclear receptor proteins

DBD – DNA BINDING DOMAINLBD – LIGAND BINDING DOMAIN HINGE REGION - Highly variable region that provides flexibility to the receptor, so it can assume different DNA-binding conformationsAF-1 transactivation domain referred

Convergence of pathways

CREB binding protein & p300 coactivator

Function of CBP/P300 protein

• Histone acetyl transferase activity

• Recruitment of RNA pol II to the promoter region of the gene of interest

• Helps in transcription and translation of proteins.

MCQ 1

MCQ 2

The phosphorylase kinase-associated regulatory protein identified by the letter A in Figure 3-4 is a calcium-binding protein. Which of the following proteins represents this regulatory subunit?

MCQ 3

Thank you