This Slide gives you a idea about the subject Cellular and Molecular pharmacology where the cell signalling, secondary messengers and its intracellular signalling pathways has been celarly explained

1. SECONDARY
MESSENGERS AND
INTRACELLULAR
SIGNALING PATHWAYS
A.Venkatraman
M Pharm 1styear 1st semester
Pharmacology
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2. DIFFERENCE BETWEEN PRIMARY AND
SECONDARY MESSENGERS
PRIMARY MESSENGER
 Extracellular factors like
hormones, or neurotransmitters
such as epinephrine, growth
hormones and serotonin
 PEPTIDE HORMONES and
neurotransmitters are hydrophilic
 Physically cannot cross the
phospholipids bilayer
SECONDARY MESSENGER
 These are intermediate molecules
like cyclic AMP or cyclic GMP
 When hormones bind to the target
cell receptor, the cell release or
creates this intermediates which
then stimulate cell responses
 Pathway involve SIGNAL
TRANSDUCTION PATHWAYS

3. SECONDARY
MESSENGERS:
 Secondary messengers are
molecules that relay signals from
receptors on the cell surface to
target molecules inside the cell
 They greatly amplify the strength
of the signal ,cause some kind of
change in the activity of the cell
 They are a component of cell
signaling pathways
 EARL WILBUR SUTHERLAND
jr., discovered secondary
messengers, for which he won the
1971 Nobel prize
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4. IMPORTANCE OF SECONDARY
MESSENGERS
 Short lived intracellular signaling molecules
 Elevated concentration of secondary messengers leads to rapid alteration in
the activity of one or more cellular enzymes
 Removal or degradation of secondary messengers terminate the cellular
response
 Four classes of secondary messengers include
Cyclic nucleotides
Membrane lipid derivatives
Ca2+
Niric oxide or carbon monoxide
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5. TYPES OF SECONDARY MESSENGERS
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6. CAMP
SIGNALING
PATHWAY
 CAMP, a major target of G-Protein signaling in
mammalian cells discovered in 1958 by earl
sutherland
 Sutherland found that the action of
epinephrine was mediated by an increase in
the intracellular concentration of cAMP,
leading to the concept that cAMP is a second
messenger in hormonal signaling
 cAMP is formed from ATP by the action of
adenylyl cyclase which is stimulated by Gs and
degraded to AMP by cAMP phosphodiesterase
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7. BASIC CONCEPT
 Effects of cAMP in animal cells are mediated
by the action of cAMP dependent protein
kinase, or protein kinase A
 Protein kinase A is a tetramer consisting of
two regulatory and two catalytic subunits.
 Cyclic AMP binds to the regulatory subunits,
leading to their dissociation from the catalytic
subunits.
 The free catalytic subunits are then
enzymatically active and able to
phosphorylate serine residues on their target
proteins
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8. Regulation of glycogen metabolism by
epinephrine
 Receptor stimulation by epinephrine leads to G protein mediated activation of
adenylyl cyclase.
 cAMP activates protein kinase A, which consists of two regulatory (R) and two
catalytic (C) subunits in its inactive form.
 Binding of cAMP to the regulatory subunits induces a conformational change
that leads to dissociation of the catalytic subunits, which are then
enzymatically active.
 Protein kinase A activates phosphorylase kinase and phosphorylase kinase
activates glycogen phosphorylase, which catalyzes the breakdown of glycogen
to glucose1phosphate.
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10. Cyclic AMP-inducible gene expression
 Receptor stimulation leads to G protein mediated activation of adenylyl
cyclase, synthesis of cAMP, and activation of protein kinase A.
 The free catalytic subunit of protein kinase A translocates to the nucleus and
phosphorylates the transcription factor CREB [ CRE binding protein]
 It leads to the recruitment of coactivators and expression of cAMP inducible
genes
 cAMP plays important roles in controlling
Proliferation
Survival
Differentiation of wide variety of animal cells
Learning and memory
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12. Regulation of protein phosphorylation by
protein kinase A and protein phosphatase 1
 Protein phosphorylation is rapidly reversed by the action of protein
phosphatases
 These protein phosphatases serve to terminate the responses initiated by
receptor activation of protein kinases.
 EXAMPLE ;the serine residues of proteins that are phosphorylated by protein
kinase A are usually dephosphorylated by the action of a phosphatase called
protein phosphatase 1
 The levels of phosphorylation of protein kinase A substrates (such as
phosphorylase kinase and CREB) are thus determined by a balance between
the intracellular activities of protein kinase A and protein phosphatases.
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13. EFFECTS OF cAMP
 cAMP are mediated by protein kinase A
 CAMP as a secondary messenger involved in sensing smells
 The odorant receptors in sensory neurons in the nose are G protein-coupled
receptors that stimulate adenylyl cyclase, leading to an increase in
intracellular cAMP.
 cAMP in this system directly opens Na+ channels in the plasma membrane,
leading to membrane depolarization and initiation of a nerve impulse.
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14. CGMP AS SECONDARY MESSENGER:
 It is an important messenger in animal cell
 Cyclic GMP is formed from GTP by guanyl cyclase and degraded to GMP BY
phosphodiesterase
 Guanyl cyclase are activated by nitric oxide and carbon monoxide as well as
peptide ligands
 Stimulation of these guanyl cyclase leads to elevation level of cGMP which
mediate responses such as blood vessel dilation
 The action of cGMP is activated is mediated by the activation of cGMP-
dependent protein kinases.
 It also regulates ion channels and phosphodiesterases
 Nitric oxide stimulates the synthesis of cGMP
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15. ROLE OF cGMP IN
PHOTORECEPTION
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It is responsible for converting the visual signals received as light
to nerve impulses
The photoreceptor in rod cells of the retina is a G-protein coupled
receptor called RHODOPSIN
Absorption of light by retinal activates the G-protein coupled
receptor RHODOPSIN.
The alpha subunit of Transducin then stimulates cGMP
phosphodiesterase ,leading to a decrease in intracellular levels of
cGMP
This change in cGMP levels in retinal rod is translated to a nerve
impulse by direct effect of cGMP in ion channels.

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17. Ca2+ a secondary messengers
 Many cells respond to extracellular stimuli by altering their intracellular
calcium concentration
 Ca++ acts as secondary messenger in two ways:
It binds to an effector molecule such as an enzyme ,activating it;
It binds to an intermediary cytosolic calcium binding protein such as
CALMODULIN
 The binding of ca++ causes profound conformational changes in calmodulin
that increase calmodulin's affinity for its effector molecules
 Calmodulin ,when activated causes contraction of smooth muscles.
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19. NITRIC
OXIDE(N0)
Nitric oxide (NO) acts as a secondary messenger
because it is free radical that can diffuse through the
plasma membrane and affect nearby cells
It is synthesized from arginine and oxygen by the NO
synthase
It activates soluble guanyl cyclase, which when
activated produces another secondary messenger,
cGMP
It is toxic in high concentration, but is the cause of
many other function like relaxation of blood vessels,
apoptosis.

21. PHOSPHATIDYLINOSITOL-DERIVED
SECONDARY MESSENGERS
 Phosphatidylinositol(PI) is a negatively charged phospholipid and a minor component in
eukaryotic cell membranes.
 The inositol can be phosphorylated to form
1. Phosphatidylinositol-4-phosphate(PIP)
2. Phosphatidylinositol-4-5-bis-phosphate(PIP3)
3. Phosphatidylinositol-3,4,5-triphosphate(PIP3)
 Intracellular enzyme PHOSPHOLIPASE C (PLC) hydrolyzes PIP2 which is found in the inner
layer of the plasma membrane .
 It yields two products
1. Diacylglycerol(DAG)
2. Inositol-1,4,5-triphosphate(IP3)

23. DIACYLGLYCEROL
 Diacylglycerol stimulates protein kinase C activity by greatly increasing the
affinity of the enzyme for calcium ions
 Protein kinase C phosphorylates specific serine and threonine residues in
target proteins.
 Known target proteins include calmodulin, the glucose transporter, HMG-CoA
reductase, cytochrome P450

24. INOSITOL TRIPHOSPHATE(IP3)
 This soluble molecule diffuse through the cytosol and binds to receptors on
the endoplasmic reticulum causing the release of calcium ions into the
cytosol
 The rise in intracellular calcium triggers the response
 Example ; the calcium rise is needed for NF-AT (the "nuclear factor of
activated t cells") to turn on the appropriate genes in the nucleus

25. Phospholipase C: IP3-DAG pathway
 Activation of phospholipase Cβ (PLcβ) by the activated GTP carrying α subunit
of Gq hydrolyses the membrane phospholipid phosphatidyl inositol 4,5-
bisphosphate (PIP2) to generate the second messengers inositol 1,4,5-
trisphosphate (IP3) and diacylglycerol (DAG).
 The primary action of IP3 is facilitation of Ca2+ mobilization from
intracellular organellar pools
 While DAG in conjunction with Ca2+ activates protein kinase C (PKc) which
phosphorylates and alters the activity of a number of functional and
structural proteins
 Cytosolic Ca2+ is a veritable messenger: combines with calmodulin (CAM) to
activate myosin light chain kinase (MLCK) inducing contraction, and another
important regulator calcium-calmodulin protein kinase (CCPK).

28. MITOGEN ACTIVATED PROTEIN
KINASE(MAPK) SIGNALING
 The MAP kinase pathway refers to a cascade of protein kinases that are highly
conserved in evolution and play central roles in signal transduction in all
eukaryotic cells ranging from yeasts to humans.
 MAPK is an enzyme that translocates signals to nucleus and activates many
transcriptional factors
 It phosphorylates many different protein that regulates expression of cell
cycle
 The importance of Ras in intracellular signaling was then indicated by
experiments showing that active Ras directly induces proliferation of normal
mammalian cells.

29. Regulation of Ras proteins

31. Induction of immediate-early genes by
ERK

32. JAK STAT SIGNALING PATHWAY
 The kinases associated with cytokine receptors include members of the Janus
kinase (or JAK) family, which consists of four related nonreceptor tyrosine
kinases.
 The key targets of the JAK kinases are the STAT proteins (signal transducers
and activators of transcription)
 Stimulation of cytokine receptors leads to recruitment of STAT proteins, which
bind via their SH2 domains
 The STAT proteins are phosphorylated by JAK family kinases.
 Pairs of phosphorylated STAT dimerize and translocate to the nucleus to
regulate gene transcription resulting in a biological response.
 Many cytokines, growth hormone, prolactin, interferons, etc. act through this
type of receptor.

33.  The STAT proteins are transcription
factors with SH2 domains that
mediate their binding to
phosphotyrosine containing
sequences.
 In unstimulated cells, STAT proteins
are inactive in the cytosol.
 Stimulation of cytokine receptors
leads to the binding of STAT
proteins to phosphotyrosine
binding sites on the receptor,
where they are phosphorylated by
the receptor associated JAK
tyrosine kinases.
 The phosphorylated STAT proteins
then dimerize and translocate to
the nucleus, where they activate
the transcription of target genes

34. CONCLUSION
 Signal transduction pathways allows cell to respond to environmental signals
 In these pathways signals are amplifies
 This signal amplification is brought by secondary messengers like cAMP ,cGMP,
Ca ions,IP3,DAG,NO
 Secondary messengers essentially serve as chemical relays from the plasma
membrane to the cytoplasm thus carrying intracellular signaling pathways .

35. REFERENCES
 The Cell, A Molecular Approach. Geoffrey M Cooper, eighth edition,2019
 KD. Tripathi. Essentials of Medical Pharmacology, Seventh edition
 The Pharmacological Basis of Therapeutics, Goodman and Gillman‘s
 Karp, Gerald, cell and molecular biology, sixth edition, john Wiley and sons,
inc.