Signal Transduction

2. SIGNAL TRANSDUCTION
• Any process occurring within cells that convert one
kind of signal/stimulus into another type
• It is also known as cell signaling in which the
transmission of molecular signals from a cell’s
exterior to its interior
• Signals received by cells must be transmitted
effectively into the cell to ensure an appropriate
response
• This step is initiated by cell surface receptors which
triggers a biochemical chain of events inside cell
creating a response

3. The Three Stages of Cell Signaling
• Earl W. Sutherland discovered how the hormone
epinephrine acts on cells
• Sutherland suggested that cells receiving signals
went through three processes
–Reception
–Transduction
–Response

4. • Signal transduction usually involves multiple
steps
• Multistep pathways can amplify a signal: A
few molecules can produce a large cellular
response
• Multistep pathways provide more
opportunities for coordination and regulation
of the cellular response
• Like falling dominoes, the receptor activates
another protein, which activates another, and
so on, until the protein producing the
response is activated

5. SIGNAL TRANSDUCTION
In this case the receptor protein is a
membrane protein
Ligand
Often turning on or off
enzyme activity

6. The extracellular signal molecule (ligand) that binds to
the receptor is a pathway’s “first messenger”
Second messengers are small, nonprotein, water-
soluble molecules or ions that spread throughout a cell
by diffusion
Second messengers relay the signal from one
location to another (such as plasma membrane to the
nucleus)  cascade of events/changes occur within the
cell
Second messengers participate in pathways initiated
by GPCRs and RTKs
Cyclic AMP and calcium ions are common second
messengers

7. Protein Phosphorylation and
Dephosphorylation
• In many pathways, the signal is transmitted by a
cascade of protein phosphorylations
• Protein kinases transfer phosphates from ATP
to protein, a process called phosphorylation

8. Protein phosphatases remove the
phosphates from proteins, a process
called dephosphorylation
This phosphorylation and
dephosphorylation system acts as a
molecular switch, turning activities on
and off or up or down, as required

9. Receptor
Signaling molecule
Activated relay
molecule
Inactive
protein kinase
1 Active
protein
kinase
1
Active
protein
kinase
2
Active
protein
kinase
3
Inactive
protein kinase
2
Inactive
protein kinase
3
Inactive
protein
Active
protein
Cellular
response
ATP
ADP
ATP
ADP
ATP
ADP
PP
PP
PP
P
P
P
P i
P i
P i

10. CLASSIFICATION OF INTERCELLULAR
COMMUNICATION
Intercellular signaling is subdivided into the following classifications:
Autocrine signals target the cell itself. Sometimes autocrine cells can target
cells close by if they are the same type of cell as the emitting cell. An
example of this are immune cells.
Paracrine signals target cells in the vicinity of the emitting cell.
neurotransmitters represent an example.
Endocrine signals target distant cells. Endocrine cells produce hormones
that travel through the blood to reach all parts of the body.
Juxtacrine signals target adjacent (touching) cells. These signals are
transmitted along cell membranes via protein or lipid components integral
to the membrane and are capable of affecting either the emitting cell or
cells immediately adjacent.

11. Activated relay
molecule
Inactive
protein kinase
1 Active
protein
kinase
1
Active
protein
kinase
2
Active
protein
kinase
3
Inactive
protein kinase
2
Inactive
protein kinase
3
Inactive
protein
Active
protein
ATP
ADP
ATP
ADP
ATP
ADP
PP
PP
PP
P
P
P i
P i
P i
P

12. AUTOCRINE SIGNALING

13. PARACRINE SIGNALING

14. ENDOCRINE SIGNALING

15. JUXTACRINE SIGNALING

16. Features of signal transduction
• Specificity: Signal molecules fits binding site on its
complementary receptor, other signals do not
• Affinity: High of receptors for signal molecules
• Amplification: Signal receptor activates many
molecules of second enzyme which activates many
molecules of third enzyme and so on.
• Desensitization: Feedback circuit shuts of the
receptor or removes it from the cell
• Integration: Two signals with opposite action on
second messenger  regulatory outcome results
from integrated output from both the receptors

17. EXTERNAL SIGNALS  TRANSDUCED TO  A PHARMACOLOGICAL
RESPONSE
LIGAND BINDING DOMAIN
RECEPTOR
FACTOR DOMAIN
1. Ligand binds to receptor Conformational change
2. Change in Transducer protein i.e. G-proteins
3. T.P activate Amplifier enzymes
Second messengers in cytoplasm
Small molecules or ions which directly or indirectly mediate cellular
response

18. 18
Receptors relay signals via intracellular SIGNALING CASCADES

19. RECEPTOR TYPES
19
Receptors can be defined by their location:
Intracellular receptors – located within the cell
Cell surface receptors or membrane receptors –
located on the plasma membrane to bind a
ligand outside the cell

20. Ligands
e.g., steroid hormones
e.g., nitric oxide
e.g., epinephrine
e.g., insulin

21. Receptors in the Plasma Membrane
• Most water-soluble signal molecules bind to
specific sites on receptor proteins that span the
plasma membrane
• There are three main types of membrane
receptors:
G protein-coupled receptors
Receptor tyrosine kinases
Ion channel receptors

22. Intracellular Receptors
• Intracellular receptors proteins are found in
the cytosol or nucleus of target cells
• Small or hydrophobic chemical messengers
can readily cross the membrane and activate
receptors
• Examples of hydrophobic messengers are the
steroid and thyroid hormones of animals
• An activated hormone-receptor complex can
act as a transcription factor, turning on specific
genes

23. Intracellular
Receptor

24. 24

25. GPCRs
As the name suggests, this RECEPTOR is coupled with G-
Proteins
GPCRs are integral membrane proteins known to possess
seven membrane-spanning domains or transmembrane helices.
The receptors span the cell membrane 7 times.
These sense molecules outside the cell and activate signal
transduction inside the cell.
Humans express over 800 GPCRs which are responsible for
every aspect of human biology from vision, taste, sense of smell,
sympathetic and parasympathetic nervous functions,
metabolism, and immune regulation to reproduction
~45% of all pharmaceutical drugs are known to target GPCRs.

27. G Proteins

28. Structure of G Protein
G proteins, also known as guanine nucleotide-binding
proteins, involved in transmitting signals and function
as molecular switches
Their activity is regulated by factors that control their
ability to bind to and hydrolyze GTP to GDP
 When they bind GTP, they are 'on', and, when they
bind GDP, they are 'off ‘
G–protein-regulated effectors include enzymes such as
adenylyl cyclase, phospholipase C, cyclic GMP
phosphodiesterase (PDE6), and membrane ion
channels selective for Ca2+ and K+

29. • The G protein heterotrimer is
composed of :
• Guanine nucleotide-binding α sub
unit(which confers specific recognition to
both receptors and effectors)
• Associated dimer of β and γ
subunits that helps confer membrane
localization of the G protein heterotrimer
by prenylation of the subunit
• In the basal state of the receptor
-heterotrimer complex, the α subunit
contains bound GDP and the –α GDP: βγ
complex is bound to the unliganded
receptor
α subunit
β subunit
γ subunit

30. G-PROTEIN SUBUNITS
The G protein family is comprised of
23 α subunits (which are the products of 17
genes)
 7 β subunits, and
12 γ subunits.
The α subunits fall into four families (Gs, Gi, Gq,
and G12/13) which are responsible for coupling
GPCRs to relatively distinct effectors.

32. GTP
GTPase

33. GASES
NO
H2S
CO
HYDROPHOBIC
Diacylglycerol
Phosphatidylinositol
s
HYDROPHILIC
cAMP
cGMP
IP3
Ca2+.
TYPES OF SECOND MESSENGERS

34. Cyclic AMP
• Cyclic AMP (cAMP) is one of the most widely used
second messengers
• A second messenger is a substance that is released in
the cytoplasm following activation of a receptor
• It is non-specific and can generate a variety of
responses in the cell
• Adenylyl cyclase, an enzyme in the plasma membrane,
converts ATP to cAMP in response to an extracellular
signal

35. Adenylyl cyclase Phosphodiesterase
Pyrophosphate
AMP
H2O
ATP
P i
P
cAMP

36. Cyclic AMP is synthesized by adenylyl cyclase under the
control of many GPCRs; stimulation is mediated by the Gs
subunit, inhibition by the Gi subunit
There are nine membrane-bound isoforms of adenylyl
cyclase (AC) and one soluble isoform found in mammals
Cyclic AMP generated by adenylyl cyclases has three
major targets in most cells:
• cyclic AMP dependent protein kinase (PKA)
• cAMP-regulated guanine nucleotide exchange
factors
• PKA phosphorylation, a transcription factor
termed CREB (cAMP response element binding
protein)

38. Gs cAMP Dependent Pathway
Stimulate

39. Gs cAMP Dependent Pathway
The Gs alpha subunit of the stimulated G protein
complex exchanges GDP for GTP and is released from
the complex
In a cAMP-dependent pathway, the activated Gs alpha
subunit binds to and activates an enzyme
called adenylyl cyclase, which, in turn, catalyzes the
conversion of ATP into cAMP
Increases in concentration of the second
messenger cAMP may lead to the activation of an
enzyme called protein kinase A (PKA).

40. Cholera Toxin
Cholera is an infection of the small intestine caused by
the bacterium Vibrio cholerae
Mechanism:
• When cholera toxin is released from the bacteria in the
infected intestine, it binds to the intestinal cells known as
enterocytes
• Toxin enters, where it activates the G protein Gs through an
ADP-ribosylation reaction that acts to lock the G protein in its
GTP-bound form, thereby continually stimulating adenylate
cyclase to produce cAMP
• Over-activation of cytosolic PKA  phosphorylates the cystic
fibrosis transmembrane conductance regulator (CFTR)
chloride channel proteins leading to ATP-mediated efflux
of chloride ions and leads to secretion of H2O, Na+,K+,
and HCO3
- into the intestinal lumen.

42. Gi cAMP Dependent Pathway
Gi mainly inhibits the cAMP dependent pathway by inhibiting
adenylate cyclase activity, decreasing the production of cAMP from
ATP, which, in turn, results in decreased activity of cAMP-
dependent protein kinase
 Ultimately Gi activation has an an opposite on cAMP-
dependent protein kinase
 Moreover, when Gi receptors gets activated, they release
activated G-protein βγ- subunits from inactive heterotrimeric G
protein complexes
 Gβγ dimeric protein interacts with GIRK channels opening them
 resulting in hyperpolarization of the cell
These receptors are primarily found on heart as well as in brain.

43. Gi cAMP Dependent Pathway

44. Terminators of cAMP-Induced
Physiologic Responses
Phosphodiesterases
Phosphatases

45. PHOSPHODIESTERASES
Hydrolyze cAMP to 5′-AMP
rapid turnover of cAMP signal termination
of biologic process after the removal of
hormonal stimulus
11 known members

46. methylated xanthine derivatives
Ex. Caffeine
-increase intracellular cAMP and
mimic or prolong the actions of
hormones
Phosphodiesterase Inhibitors

47. PHOSPHATIDYLINOSITOL-DERIVED
SECOND MESSENGERS
• Phosphatidylinositol ( PI) is a negatively charged
phospholipid and a minor component in eukaryotic cell
membranes.
• The inositol can be phosphorylated to form
– Phosphatidylinositol-4-phosphate (PIP)
– Phosphatidylinositol-4,5-bis-phosphate (PIP2)
– Phosphatidylinositol-3,4,5-trisphosphate (PIP3)
• Intracellular enzyme phospholipase C (PLC),hydrolyzes
PIP2 which is found in the inner layer of the plasma
membrane. Hydrolysis of PIP2 yields two products:
– Diacylglycerol (DAG)
– Inositol-1,4,5-trisphosphate (IP3)
PHOSPHOINOSITDES

49. MODE OF ACTION
• Peptide and protein hormones like vasopressin, TSH, and
neurotransmitters like GABA bind to GPCRs
• This activate the intracellular enzyme phospholipase C
(PLC).
• PLC in turn cleaves PIP2 to yield two products – DAG and
IP3.
• Both of these products act as second messengers.
• Cleavage of PIP2 by PLC is the functional equivalent of
the synthesis of cAMP by adenylyl cyclase

50. Gq Protein Coupled Receptor
Gq protein is a heterotrimeric protein subunit
that activates phospholipase C (PLC)
PLC in turn hydrolyzes Phosphatidylinositol
4,5-bisphosphate (PIP2) to diacyl glycerol
(DAG) and inositol trisphosphate (IP3) signal
transduction pathway
DAG acts as a second messenger that activates
Protein Kinase C (PKC) and IP3 acts on calcium
channels to release calcium from stores and
phosphorylation of some proteins

51. G protein
EXTRA-
CELLULAR
FLUID
Signaling molecule
(first messenger)
G protein-coupled
receptor
Phospholipase C
DAG
PIP2
IP3
(second messenger)
IP3-gated
calcium channel
Endoplasmic
reticulum (ER)
CYTOSOL
Various
proteins
activated
Cellular
responses
Ca2
(second
messenger)
Ca2
GTP
Gq

52. • Low cytoplasmic Ca++ at rest (10–100 nM)
• To maintain this low concentration, Ca2+ is actively pumped
from the cytosol to the extracellular space and into
the endoplasmic reticulum (ER)
• Certain proteins of the cytoplasm and organelles act as
buffers by binding Ca2+
• Signalling occurs when the cell is stimulated to release
calcium ions (Ca2+) from intracellular stores, and/or when
calcium enters the cell through plasma membrane ion
channels.
Calcium as a 2nd Messenger

53. Calcium acts as a second 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 Calcium causes profound
conformational changes in calmodulin that increase
calmodulin`s affinity for its effector molecules.
Calmodulin, when activated, causes contraction of
smooth muscles

54. Calcium as a 2nd Messenger

55. WHAT IS A TYROSINE KINASE
• A tyrosine kinase is an enzyme that can
transfer a phosphate group from an ATP to
a tyrosine residue in a protein
• Tyrosine kinases are important mediators of
the signaling cascade, determining key roles
in diverse biological processes like growth,
differentiation, metabolism and apoptosis in
response to external and internal stimuli.

56. Structure
Four common structural features shared
among RTKs:
 Extracellular ligand-binding domain
 Single transmembrane domain
 Cytoplasmic tyrosine kinase domain(s)
 Regulatory domains

57. Classification of RTK
 Receptor tyrosine kinases (RTKs)-The RTK family includes
the receptors for insulin and for many growth factors such as:
 Epidermal growth factor (EGF)
 Fibroblast growth factor(FGF)
 Platelet-derived growth factor (PDGF)
 Vascular endothelial growth factor(VEGF)
 Nerve growth factor (NGF)
 Nonreceptor tyrosine kinases (NRTKs)
 Src
 Janus kinases (Jaks)
 Abl

58. • When a growth factor binds to the
extracellular domain of an RTK,
its dimerization is triggered with other
adjacent RTKs.
• Dimerization leads to a rapid activation of the
protein's cytoplasmic kinase domains, the first
substrate for these domains being the
receptor itself.
• The activated receptor as a result then
becomes autophosphorylated on multiple
specific intracellular tyrosine residues

59. Signal transduction
• The phosphorylation of specific tyrosine residues within the
activated receptor creates binding sites for Src homology 2 (SH2)
domain- and phosphotyrosine binding (PTB) domain-containing
proteins.
• Specific proteins containing these domains
include Src and phospholipase Cγ
• Phosphorylation and activation of these two proteins on
receptor binding lead to the initiation of signal
transduction pathways
• Other proteins that interact with the activated receptor act
as adaptor proteins and have no intrinsic enzymatic activity of
their own.
• These adaptor proteins link RTK activation to downstream signal
transduction pathways, such as the MAP kinase signalling
cascade.

60. Signaling
molecule (ligand)
2
1
3 4
Ligand-binding site
 helix in the
membrane
Tyrosines
CYTOPLASM Receptor tyrosine
kinase proteins
(inactive monomers)
Signaling
molecule
Dimer
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
Tyr
P
P
P
P
P
P
P
P
P
P
P
P
Activated tyrosine
kinase regions
(unphosphorylated
dimer)
Fully activated
receptor tyrosine
kinase
(phosphorylated
dimer)
Activated relay
proteins
Cellular
response 1
Cellular
response 2
Inactive
relay proteins
6 ATP 6 ADP

62. The insulin receptor (IR) is a transmembrane receptor that is activated
by insulin, IGF-I, IGF-II and belongs to the large class of tyrosine
kinase receptors.