signal transduction
Post on 17-Jun-2015
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G-PROTEINS • 30 different types for different signaling
pathways • Major two types : • G-Stimulatory(Gs) : Stimulates AC• G-Inhibitory (Gi) : Inhibit AC • α – subunit of Gs and Gi are different while
β and γ are similar
1. G-Protein – Coupled Receptors(Largest family of cell surface receptors)
cAMP is a second messenger that mediates cellular responses to a variety of hormones.
G-PROTEINS
• The hormone - triggered exchange of GTP for GDP in Gi splits it into two subunits then binds α – subunit of Gs, reversing the activation of AC
• The inhibition is more stronger and effective as cellular concentration of Gi is more than Gs
NorepinephrineAdenylate cyclase
cAMP
The protein kinase that is activated by cAMP is called protein kinase A (PKA).
Low glucoselevel
Pancreas
glucagonTargets onliver cell
cAMP
glycogen converted to glucose
glucose enters blood
adenylate cyclase
• Epinephrine is also involved in the control of glucose levels
• Similar in effect to glucagon but affects primarily muscle tissue.
epinephrine
cAMP production
Activated PKA
(?) Protein phosphorylation
glycogen degradation
glucose enters blood
- adrenergic receptor Adenylate
cyclase
Characteristics of signal transduction:
• Chemical signal that results from Ligand/ receptor binding is amplified.
Many molecules of second messenger can be produced from a single hormone signal.
αi -GTP inhibits Adenylate Cyclase by binding with it and lowers intracellular concentration of c-AMP
Main action of c-AMP is to activate some protein kinases allosterically
Diseases of cellular communication
• Example - Cholera toxin • Binds to gangliosides of intestinal mucosal cells leading to
ribosylation of α- subunit of Gs and inhibition of Inherent GTPase activity and irreversible activation of G-protein :
• Results in continuous activation of adenylate cyclase & high cAMP levels in epithelial cells of the intestine.
• Causes uncontrolled release of water and sodium, leading to diarrhea
and dehydration.
• Pertussis toxin: Assignment
Two different secondary messenger systems areinvolved in adrenaline signal transduction: The action of receptors is mediated by G protein- adenylate cyclase - cAMP system;
The action of receptors are mostly related with phospholipase signal pathways and relocation of intracellular Ca++.
Phospholipase C – IP3 - Ca++ SecondMessenger System
Binding of epinephrine to the -receptorsstimulates a totally different intracellular cascade.
A membrane enzyme, phospholipase C is activated
via G-protein (, complex). Phospho inositol Inositol-3-triphosphate (IP3) 4,5-bisphosphate (PIP2) + diacelglycerol
Two secondary messengers, IP3 and DAG are generated.
PLC
The binding of IP3 and its receptor causes the opening of Ca++ channel on the endoplasmic reticulum membrane. Ca++ will be released from ER into the cytoplasm down…
Elevated intracellular Ca triggers processes like smooth ms contraction , glycogen breakdown and exocytosis
Hormones acting through Calcium • Intracellular Ca concentration is lower than
extracellular • Hrs can change it by following mechanisms : • Alter the permeability of membrane • Action of Ca-H ATPase pump • By releasing intracellular stores • Binding of Ca To CALMODULIN , calcium
dependant regulatory protein with 4 Ca binding site
Hormones acting through Calcium • Binding of Ca brings conformational change in
calmodulin and plays a role in regulating various Kinases
• Phosphodiestrase (cleaves cAMP) is regulated by Ca levels in the cells
• Intracellular ca acts as mediator of Hormone action either independently or in conjunction with cAMP
• EXAMPLE : Phosphorylase Kinase action : ASSIGNMENT
Activation of - adrenergic receptor
G protein
Phospholipase C
PIP2 IP3 + DAG
Endoplasmic reticulum opening of Ca channels
Ca++ + Calmodulin
Protein kinase C Protein P-tion …
Liver cell
- receptor
-receptor
PKA
PKC
Phosphatidylinositol (PIP2) Signal Cascades
Some hormones activate a signal cascade based on the membrane lipid phosphatidylinositol.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OH
H
H
OHH
OH
H
O
H OH
1 6
5
43
2
phosphatidyl-inositol
Kinases sequentially catalyze transfer of Pi from ATP to OH groups at positions 5 & 4 of the inositol ring, to yield phosphatidylinositol-4,5-bisphosphate (PIP2).
PIP2 is cleaved by the enzyme Phospholipase C.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OPO 32
H
H
OPO 32H
OH
H
O
H OH
1 6
5
43
2
PIP2 phosphatidylinositol- 4,5-bisphosphate
When a particular GPCR (receptor) is activated, GTP exchanges for GDP. Gqa-GTP activates Phospholipase C. Ca++, which is required for activity of Phospholipase C, interacts with (-) charged residues & with Pi moieties of the phosphorylated inositol at the active site.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OPO 32
H
H
OPO 32H
OH
H
O
H OH
1 6
5
43
2
PIP2 phosphatidylinositol- 4,5-bisphosphate
cleavage by Phospholipase C
Different isoforms of Phospholipase C have different regulatory domains, & thus respond to different signals.
A G-protein, Gq activates one form of Phospholipase C.
Cleavage of PIP2, catalyzed by Phospholipase C, yields 2 second messengers: inositol-1,4,5-trisphosphate (IP3) diacelglycerol (DG).
Diacelglycerol, with Ca++, activates Protein Kinase C, which catalyzes phosphorylation of several cellular proteins, altering their activity.
O HH 2 C
C H
H 2 C
OCR 1
O O C
O
R 2
d iacylg lycero l
O H
H
O PO 32
H
H
O PO 32 H
O H
H
H O H
O PO 32
1 6
5
43
2
IP 3 in o sito l-1 ,4 ,5 -trisp h o sp h a te
IP3 activates Ca++-release channels in ER membranes.
Ca++ stored in the ER is released to the cytosol, where it may bind calmodulin, or help activate Protein Kinase C.
Signal turn-off includes removal of Ca++ from the cytosol via Ca++-ATPase pumps, & degradation of IP3.
Ca++
ATP ADP + Pi
Ca++
IP3
calmodulin
endoplasmic reticulum
Ca++
Ca++-ATPase
Ca++-release channel
Sequential dephosphorylation of IP3 by enzyme-catalyzed hydrolysis yields inositol, a substrate for synthesis of PI.
IP3 may instead be phosphorylated via specific kinases, to IP4, IP5 or IP6. Some of these have signal roles.
E.g., the IP4 inositol-1,3,4,5-tetraphosphate in some cells stimulates Ca++ entry, perhaps by activating plasma membrane Ca++ channels.
O H
H
O H
H
H
O HH
O H
H
H O H
O H
O H
H
O PO 32
H
H
O PO 32 H
O H
H
H O H
O PO 32
(3 s teps)+ 3 P i
IP 3 inosito l
The kinases that convert PI (Phosphatidylinositol) to PIP2 (PI-4,5-P2) transfer Pi from ATP to OH at positions 4 & 5 of the inositol ring. PI 3-Kinases instead catalyze phosphorylation of Phosphatidylinositol at the 3 position of the inositol ring.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OH
H
H
OHH
OPO32
H
O
H OH
1 6
52
3 4
phosphatidyl-inositol-
3-phosphate
Head-groups of these transiently formed lipids are ligands for particular pleckstrin homology (PH) & FYVE protein domains that bind proteins to membrane surfaces. Other protein domains called MARKS are (+) charged, and their binding to (-) charged head-groups of lipids like PIP2 is antagonized by Ca++.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OH
HH
OHH
OPO32
H
O
H OH
1 6
52
3 4
phosphatidyl-inositol-
3-phosphate
PI-3-P, PI-3,4-P2, PI-3,4,5-P3, and PI-4,5-P2 have signaling roles.
Protein Kinase B (also called Akt) becomes activated when it is recruited from the cytosol to the plasma membrane surface by binding to products of PI-3 Kinase, e.g., PI-3,4,5-P3. Other kinases at the Cytosolic surface of the plasma
membrane then catalyze phosphorylation of Protein Kinase B, activating it.
Activated Protein Kinase B catalyzes phosphorylation of Ser or Thr residues of many proteins, with diverse effects on metabolism, cell growth, and apoptosis.
Downstream metabolic effects of Protein Kinase B include stimulation of Glycogen synthesis, stimulation of Glycolysis, and inhibition of Gluconeogenesis.
Tyrosine Kinase System (Complex and largely unknown)
Peptide hormones such as Growth hormone, Prolactin, Insulin and other growth factors use tyrosine kinase system for their intracellular signal transduction.
Ligand-receptor binding will activate tyrosine kinase (some times as part of the receptor) that phosphory-lates the receptors and some transducer proteins at the tyrosine residue.
Schematic Representation of Insulin Signaling Pathways
ST
AT
S
ST
AT
S
P
InsR
Ins
SHCGrb2
RasTK
Jak-2Jak-2SOS
Raf
P P
ST
AT
S
ST
AT
S
P
MAPKK
MAPK
ST
AT
S
P
The intracellular transducer proteins that are phosphorylated by tyrosine kinase include Janus kinase, Map kinase, STAT (signal transducers and activators of transcription). …
The eventual effect is altered DNA expression levels and stimulated cell division.
Communication between cells
• Multicellular organisms need a way to coordinate activities between cells.
• Most cells produce and secrete molecules to pass information to others - paracrine control.
• The signals for paracrine control are mostly growth factors and cytokines, which use tyrosine kinase pathways.
Lipid rafts: Complex sphingolipids tend to separate out from
glycerophospholipids & co-localize with cholesterol in membrane microdomains called lipid rafts.
Membrane fragments assumed to be lipid rafts are found to be resistant to detergent solubilization, which has facilitated their isolation & characterization.
Differences in molecular shape may contribute to a tendency for sphingolipids to separate out from glycerophospholipids in membrane microdomains.
Signal complexes are often associated with lipid raft domains of the plasma membrane.
Scaffold proteins as well as signal proteins may be recruited from the cytosol to such membrane domains in part by insertion of lipid anchors interaction of pleckstrin homology or other lipid-
binding domains with head-groups of transiently formed phosphatidylinositol derivatives, such as PIP2 or PI-3-P.
AKAPs (A-Kinase Anchoring Proteins) are scaffold proteins with multiple domains that bind to regulatory subunits of Protein Kinase A phosphorylated derivatives of phosphatidylinositol various other signal proteins, such as:
• G-protein-coupled receptors (GPCRs)• Other kinases such as Protein Kinase C• Protein phosphatases• Phosphodiesterases
AKAPs localize signal cascades within a cell.
They coordinate activation of protein kinases as well as rapid turn-off of signals.
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