1. Sheet #14
Lecture (Signal Transduction part I)
Lecture Date :
Done By :- إِيناس العـمايرة 
Edited By :- سيبويه سياج
Doctor :- Nabil Amer 

2. Cell Communication – Signal Transduction
External signal is received and converted to another form to elicit a response 2

3. Signal Transduction & G Protein-coupled Receptors
Topics
Signal Trans.: From Extracellular Signal
to Cellular Response
Cell-Surface Receptors & Signal
Transduction Proteins
G Protein-coupled Receptors (GPCRs):
Structure and Mechanism
GPCRs That Regulate Ion Channels
GPCRs That Regulate Adenylyl Cyclase
GPCRs That Regulate Cytosolic Calcium 3

4. Learn the G protein cycle of reactions involved in GPCR signaling.
Learning Objectives
Learn the general properties of signaling molecules (ligands), cell- surface receptors, & intracellular signal transduction components.
Learn the G protein cycle of reactions involved in GPCR signaling.
Learn the epinephrine receptor signal trans pathway used for control of glycogen degradation.
Learn about the GPCR-stimulated IP3/DAG signaling pathway.

5. General Principles of Signal Transduction
refers to the overall process 
of converting extracellular 
signals into intracellular 
responses .
Key players in signal transduction are :
1.signaling molecules.
 2.receptors.
3.signal transduction proteins.
4.second messengers. 
5.effector proteins.  5

6. 2 General principles of transduction :
(1) synthesis of signaling molecule .
(2)Releasing of these molecules from vesicles to Ext space .
(3)Transport of Signaling molecule to target cell .
*signaling moleculeإلى أن يرتبط بها الـ inactiveتكون في حالة  (protein receptors )المُستقبِلات 
(4)Activation of receptor protein by binding of the signaling molecule .
(5)Several modifications occur inside the cell (Initializing of signal transduction pathways + forming of 2nd messengers).
(6) The formed effector protein leads to changes in cellular functions in one of the two :
A.Modification of cellular metabolism .
B.Modification of gene expression .
(7)Inactivation of the receptor protein .
(8)Removing of signaling molecules . 2

7. Cells respond to signals by changing the activity of existing enzymes (fast) and/or the levels of expression of enzymes and cell components (slow) by gene regulation 
(Steps 7a & 7b). 
Receptors and signal transduction systems have evolved to detect and respond to hormones, growth factors, drugs & neurotransmitters. 
*Fast : Because of the presence of the enzyme .
Signals affect either the existing enzyme (Stimulating or activating it) or gene synthesis .

8. Intercellular Communication

9. Intercellular Communication
Communication between cells requires: ligand: the signaling molecule. receptor protein: the molecule to which the receptor binds -may be on the plasma membrane or within the cell 9

10. 2 Sheet page (2): ++Extra : GH ≡ Growth hormone.
Receptors are of two types : A. Membrane (cell-surface) receptors *found on the plasma membrane. B. Intracellular or “nuclear” receptors *found within the cell . Ligands based on their nature : A. Hydrophilic ligands; binds with cell-surface receptors (i.e. peptide hormones; GH , FSH , LH, ACTH ,Glucagon and Epinephrine). B. Hydrophobic ligands; which pass the membrane and reach the nucleus (their effect site) e.g: Steroid hormones, Vitamin D.
++Extra :
GH ≡ Growth hormone. 
FSH ≡ Follicle stimulating hormone. 
LH ≡ Luteinizing Hormone .
ACTH ≡ Adrenocorticotropic hormone . 2

11. Will bind to the membrane receptor (out of the cell).
Sheet Note :
Will bind to the membrane receptor (out of the cell).
Will enter the cell then binds to the receptor in order to produce cellular response 11

12. Receptors have a globular (not fibrous) structure .
Structure and function of receptors
Receptors have a globular (not fibrous) structure .
Globular proteins acting as a cell’s ‘letter boxes’.
Located mostly in the cell membrane.
Receive messages from chemical messengers coming from other cells.
Transmit a message into the cell leading to a cellular effect.
Different receptors specific for different chemical messengers. 
Each cell has a range of receptors in the cell membrane making it responsive to different chemical messengers.(Each cell has varying numbers of receptors which means that it doesn't contain only single receptor , and these receptors perform functions that differ between cells and that depends on the function of the cell itself ).  12

13. Mechanism
Receptors contain a binding site (hollow or cleft in the receptor surface) that is recognised by the chemical messenger.
Binding of the messenger involves intermolecular bonds.
Binding results in an induced fit of the receptor protein.
Change in receptor shape results in a ‘domino’ effect.
Domino effect is known as Signal Transduction, leading to a chemical signal being received inside the cell .
Chemical messenger does not enter the cell. It departs the receptor unchanged and is not permanently bound 13

14. Overall process of receptor/messenger interaction
Sheet : Binding of the ligand leads to conformational changes in the receptor and that's called Induced Fit M E R M M E R R
Signal transduction
Binding interactions must be:
       - strong enough to hold the messenger sufficiently long    
         for signal transduction to take place.
       - weak enough to allow the messenger to depart(Leave) .
Implies a fine balance
Drug design - designing molecules with stronger binding interactions results in drugs that block the binding site – antagonists*(Drugs mainly inhibit the binding of the ligand to the receptor .Instead , they bind to the receptor). 14

15. Messenger binding
Sheet Note
Ser, Phe, Asp are three important amino acids found in the binding site .
Ser: -OH containg a.a 
Asp: Acidic a.a 
Phe : Nonpolar a.a
  Bonding forces (Between the ligand and the receptor).
Ionic
H-bonding
van der Waals
vdw
interaction
Example:
Phe
H-bond
Binding site
Ser O H
ionic
bond
Asp
CO2
Receptor 15

16. How does the Binding Site Change Shape? Substrate binding
Bonding forces
Induced fit - Binding site alters shape to maximize intermolecular bonding.
Phe
Ser O H
Asp
CO2
Phe
Ser O H
Asp
CO2
Induced
Fit
Intermolecular bonds not optimum length for maximum binding strength
Intermolecular bond lengths optimised 16

17. The specific action of an enzyme with a single substrate can be explained by:
1.Induced Fit (Change of receptor shape once the binding of ligand occurs)
More common
2.Lock and key model .(مفتاح فقط يدخل في قفل واحد).
"If we imagine that the enzyme is a lock and it has its own shape, only the correctly shaped keys (substrates) that fit the lock(enzyme) are capable of opening (binding to) it ".
Differences between the two mechanisms :
Lock and Key states that there is no change needed and that only a certain type will fit.
Induced fit states that the active site(binding site) will change to help to substrate fit. 3

18. External signals are converted to
Internal Responses
 Cells sense and respond to the environment .
Prokaryotes: chemicals
Humans: 
Light  - rods & cones of the eye
sound – hair cells of inner ear
chemicals in food – nose(smell) & tongue(taste).
Cells communicate with each other
1.Direct contact
2.Chemical signals   18

19. General principles 1. Signals act over different ranges.
2.  Signals have different chemical natures.     
  3.  The same signal can induce a
        different response in different cells.
Note#4:
Remember the effect of epinephrine in the liver and adipose tissues ; epinephrine in the liver stimulates (Glycogenolysis )while in adipose tissues it stimulates (Glycolysis).
4.  Cells respond to sets of signals.
5.  Receptors relay signals via
intracellular signaling cascades. 19

20. Cells detect signal & respond
1º messenger
Effector
Enzymes
Target
Enzymes
2º messengers
Signal transduction: ability of cell to translate
   receptor-ligand interaction into a change in behavioror gene expression 20

21. Relay molecules in a signal transduction pathway
Primary
Messenger
Secondary
Messengers
Target
Enzymes
EXTRACELLULAR
FLUID
Receptor
Signal
molecule
Relay molecules in a signal transduction pathway
Plasma membrane
CYTOPLASM
Activation
of cellular
response
Reception 1
Transduction 2
Response 3
Cascade Effect 21

22. 4 Cascade of signaling events are :
1.Reception involves : Signaling molecule (Ligand )and the receptor .
Notice : the ligand is also called 1°(primary) or first messenger.
2.Transduction (Signal Transduction) involves : 2°(Secondary) or second messengers. "Transduction event converts the signal or Ligand to a form that can bring the response "
3.Response (Cellular response) : The cell respond in different ways :
A. Gene modification 
B. Target enzyme 
                                              4

23. - activates an enzyme activity, processes 100 substrates /second
Each protein in a signaling pathwayAmplifies the signal by activating multiple copies of the next component in the pathway.
 - activates an enzyme activity, processes 100 substrates /second
primary signal
*Primary enzyme activates 100 target enzymes
*Each of the 100 enzymes activates an additional 100 downstream target enzymes.
*Each of the 10,000 downstream targets activates 100 control factors so rapidly have 1,000,000 active control factors 23

24. A Signal cascade amplification .
I epinephrine --->{Response} 10^8 Glc-1-phosphate 
The transduction stage ends with the activation of 10^6 molecule of  Glycogen phosphorylase that breaks down glycogen into Glc subunits to produce Glc-1-phosphate. 

25. For better understanding : Signaling Cascade : a series of chemical reactions which are initiated by a stimulus (first messenger) acting on a receptor that is transduced to the cell interior through second messengers (which amplify the initial signal) and ultimately to effector molecules, resulting in a cell response to the initial stimulus. 5

26. Receptors relay signals via intracellular SIGNALING CASCADES26

27. Main Types of Receptors
ION CHANNEL RECEPTORS
G-PROTEIN-COUPLED RECEPTORS
KINASE-LINKED RECEPTORS
INTRACELLULAR RECEPTORS

28. 4.enzyme-linked receptor
1.Cell-surface receptors -large and/or hydrophilic ligands
2.ion-channel-linked receptor
3. Trimeric G-protein-linked receptor.(composed of subunits that are activated by binding of signaling molecule ).    
    
4.enzyme-linked receptor  
   (tyrosine kinase)  28

29. There are different types of cellular receptors ; and they are : 1
There are different types of cellular receptors ; and they are : 1. Cell surface receptors (Membrane Receptors): They act in a signaling pathway by receiving (binding to) extracellular molecules. 2. Ion-Channel linked receptors (Either closed or opened) 3.GPCR (G-protein coupled receptors ): Composed of subunits that are activated by signal molecule . 4.Enzyme Linked Receptors : They have two important domains* : A.Extracellular Domain . B.Intracellular (Catalytic ) Domain ; on the cytoplasmic side of the receptor. (Most notably is Tyrosine Kinase ) 5.Intracellular Receptors *The doctor said that they are composed of 2 subunits ; it isn't accurate since the number of subunits differ widely between different types of receptors , while all enzyme linked receptors are of 2 domains (Ext + Int). 6

30. Receptor protein is part of an ion channel protein complex
Signal transduction
 Control of ion channels
Receptor protein is part of an ion channel protein complex
Receptor binds a messenger leading to an induced fit
Ion channel is opened or closed
Ion channels are specific for specific ions (Na+, Ca2+, Cl-, K+)
Ions flow across cell membrane down concentration gradient.
Polarises or depolarises nerve membranes
Activates or deactivates enzyme catalysed reactions within a cell 30

31. Ion channel receptors Examples: 1. Muscle Contraction
Cellular response
Gate open
Gate close
Ligand-gated
ion channel receptor
Plasma Membrane
Signal molecule (ligand)
Gate closed
Ions
Examples:
1. Muscle Contraction
2.Nerve Cell communication 31

32. Review: Na+ Cl- - + Remember the Na+/K+ ATPase (Na+/K+ pump)?
[Na+] inside ~10mM; outside ~150mM
[K+] inside ~150mM; outside ~5mM
cell has membrane potential ~ -60mV 
-60mV K+ A-
Na+
Cl- - + 32

33. Intercellular Communication
How ?

34. Intercellular Communication
There are four basic mechanisms for cellular communication: 1. Direct contact. 2. Paracrine signaling. 3. Endocrine signaling. 4. Synaptic signaling. 34

35. Intercellular Communication
Direct contact (Surface to surface Contact)– molecules on the surface of one cell are recognized by receptors on the adjacent cell. 35

36. Intercellular Communication
Paracrine signaling – signal released from a cell has an effect on neighboring cells
*local (Cells secrete local chemical mediators to affect neighbouring cells ). 
ex.  nitric oxide, 
histamines,prostaglandins 36

37. Intercellular Communication
Endocrine signaling – hormones released from a cell affect other cells throughout the body
long distance (Cells or glands secrete hormones that travel through bloodstream to target cells that are distributed widely throughout the body )
ex. Estrogen, Thyroxine(Pituitary Gland), GH(Growth Hormone),Epinephrine . 37

38. Intercellular Communication
Synaptic signaling – nerve cells release the signal (neurotransmitter) which binds to receptors on nearby cells 38

39. Synaptic Signaling :
Pre-Synaptic cell : From which the neurotransmitter is released into the synaptic gap (Synaptic Cleft).
Post-Synaptic cell : The target cell that is affected (stimulated) by the released neurotransmitter .
((Neurotransmitter : PRE To POST )).
After the wanted effect (by NT) is achieved, the activity of NT is terminated by one of the two ways :
A. Reuptake mechanism .
B. Degradation (by an enzyme that is in the synaptic gap).
Ex: Acetylcholine  7

40. Intercellular Communication
When a ligand binds to a receptor protein, the cell has a response. Signal transduction: the events within the cell that occur in response to a signal. Different cell types can respond differently to the same signal. 40

41. Receptor Types
There are 3 subclasses of membrane receptors: 1. channel linked receptors – ion channel that opens in response to a ligand 2. enzymatic receptors – receptor is an enzyme that is activated by the ligand 3. G protein-coupled receptor – a G-protein (bound to GTP) assists in transmitting the signal 41

42. 42

43. Intercellular Communication
A cell’s response to a signal often involves activating or inactivating proteins. Phosphorylation is a common way to change the activity of a protein. protein kinase – an enzyme that adds a phosphate to a protein(Phosphorylation). phosphatase – an enzyme that removes a phosphate from a protein(Dephosphorylation). 43

44. Remember : Ser ,The and Tyr are –OH containing amino acids 44

45. Signal Transduction Components: Kinases/Phosphatases
Proteins that participate in intracellular signal transduction fall into two main classes--protein kinases/phosphatases and GTPase switch proteins. Kinases use ATP to phosphorylate amino acid side-chains in target proteins. Kinases typically are specific for tyrosine or serine/threonine sites. Phosphatases hydrolyze phosphates off of these residues. Kinases and phosphatases act together to switch the function of a target protein on or off . 45

46. Dual-Specifity Kinase : A kinase that can act as both Ser/Thr kinase .
Kinases - Phosphorylation
Phosphatase - Dephosphorylation
*Tyrosine-OH : Tyr-Kinases
**Serine-OH
Ser/Thr-Kinases
**Threonine-OH
„dual specificity“ Kinases

48. There are about 600 kinases and 100 phosphatases encoded in the human genome. Activation of many cell-surface receptors leads directly or indirectly to changes in kinase or phosphatase activity. Note that some receptors are themselves kinases (e.g., the insulin receptor).

49. 49

50. Insulin Receptor : Heteromeric receptor ; composed of 2 alpha subunits and 2 beta subunits . *The alpha subunit is entirely extracellular and contains insulin-binding domain . *The beta subunit has an intracellular domain that expresses Tyr kinase activity . Binding of Insulin molecules to the receptor leads to the dimerization and activation of beta chains. **Notice ; insulin binding results in Tyr phosphorylation of the beta subunit . *Phosphorylated proteins activate glycogen synthase;that converts Glc into Glycogen. Insulin is an anabolic hormone ;responsible for Glycogen Formation , increases potassium uptake , increases amino acid uptake , and stimulates glucose uptake while Acetylcholine ,epinephrine and cortisone are catabolic hormones that work in the opposite of Insulin ; degradation of glycogen (To provide us with energy). **Notice : Glycogen synthase enzyme (Stimulated by Insulin) is the opposite in action of Glycogen phosphorylase enzyme (Stimulated by Epinephrine , Glucagon , and cortisone )** *Glycogen biosynthesis and its degradation will be taken later on . 8

51. Growth hormone receptor(Works on the cell surface).
Tetrameric complex constructed in presence of growth hormone  GH
GH binding
&
dimerisation OH
Binding
of kinases HO OP PO
ATP
ADP
Activation and
phosphorylation
GH receptors
(no kinase activity) OH HO
kinases
Kinase active site
opened by induced fit
Growth hormone binding site
Kinase active site

52. Intracellular receptors
Chemical messengers must cross cell membrane
Chemical messengers must be hydrophobic
Example-steroids and
steroid receptors
CO2H
H2N
Steroid
binding region
Zinc
DNA binding region
(‘zinc fingers’)
Zinc fingers contain Cys residues (SH)
Allow S-Zn interactions

53. Intracellular Receptors
steroid hormones -have a nonpolar, lipid-soluble structure -can cross the plasma membrane to a steroid receptor -usually affect regulation of gene expression An inhibitor blocks the receptor from binding to DNA until the hormone is present. 53

54. Intracellular receptor Mechanism
Co-activator
protein
Receptor
Cell
membrane
DNA
Receptor-ligand
complex
Dimerisation
Messenger
1. Messenger crosses membrane
2. Binds to receptor
3. Receptor dimerisation
5. Complex binds to DNA
6. Transcription switched on or off
7. Protein synthesis activated or inhibited
4. Binds co-activator protein

55. Intracellular Receptors
A steroid receptor has 3 functional domains: 1. hormone-binding domain. 2. DNA binding domain. 3. domain that interacts with coactivators to affect gene expression. 55

56. 56

57.  Binding of Hormone-receptor complex to DNA molecule in the nucleus can increase synthesis or decrease(or repress) synthesis of the effector substance.
Steroid hormone receptor can be either in the cytoplasm or in the nucleus(Like thyroid Hormone receptor )and some steroid hormone receptors are found on the membrane of the target cell .
Remember : Receptor-hormone complex acts in the nucleus to enhance or inhibit transcription . 9

58. END PART I