Immuno Test 2 (Part 2) Flashcards
The Infectious- Nonself Model
- APCs recognize EVOLUTIONARY DISTANT PATHOGENS
- APCs activated via PRR
- PRR recognize PAMP on bacteris
- Upon activation, APC up-regulate COSTIMULATORY SIGNALS, process the bacterial ANTIGENS, and PRESENT them to passing T CELLS
***The PRR allow APC to DISCRIMINATE between “INFECTIOUS-NONSELF” and “NONINFECTIOUS-SELF”
Danger Theory Model
- APCs are ACTIVATED by DANGER/ DAMAGE Signals
- Danger Signals are generated by:
1) Injured Cells
2) Pathogens
3) Toxins
4) Mechanical Damage
What is the Danger Signal?
Potential Danger Signals:
1) Infection by Microbial Pathogens
2) Products of Injured (Necrosis) or Stressed Cells
3) Immunostimulatory Compounds (HEPARAN SULFATE)
4) Inflammatory Cytokines (IFN-alpha/ beta or TNF-alpha)
5) Rupture of vessels or chemotaxis of Blood-Borne Cells
- ***For POTENT ACTIVATION:
1) MHC Signal
2) CD28 to B7
Humoral Response to Danger Signals
- Activation of Complement
- Activated Immune Cells Release:
1) Chemokines
2) Leucotrienes and Prostaglandins
3) Reactive Oxygen Intermediates (ROI)
4) Nitric Oxide (NO)
Necrosis Generates Danger Signals and Inflammation
***NECROSIS: a Passice, Catabolic cell DEATH in response to EXTERNAL TOXIC FACTORS
- Necrosis is a dirty FORM of cell death characterized by SWELLING and RUPTURE of cell membrane (cell lyse) which may cause INFLAMMATION or harm other neighboring cells
**INFLAMMATION: a response to the INNATE IMMUNITY that involves activation of leukocytes and release of inflammatory mediators
- Immune cells are activated by the DANGER SIGNALS
Necrosis and Danger/ Damage Signals
1) HMGB1: High Mobility Group Box 1, is a protein passively released during Necrosis:
- Activates NF-kB pathway
- RAGE is a receptor for HMGB1
* * HMGB1 is an ENDOGENOUS DANGER SIGNAL*
2) URIC ACID is another diffusible danger signal:
- Activated NF-kB
3) HSPs are another danger signal:
- HSPs induce NF-kB pathway and release Inflammatory cytokines (TNF-alpha and IL-1beta)
HSP: Heat Shock Proteins
RAGE: Receptor of Advanced Glycation End Products
Acute Inflammatory Response
1) Increased BLOOD SUPPLY to the affected areas —> REDNESS and HEAT
2) Increased in CAPILLARY PERMEABILITY —> Leak from the BLOOD VESSELS —> SWELLING AND PAIN
3) Massive INFLUX of NEUTROPHILS
4) Arrival of MACROPHAGES (16-48hrs)
5) Macrophage mediated PHAGOCYTOSIS of debris, restoring the HOMEOSTASIS
Stages of Acute Inflammatory Response
1) DETECTION of Danger/ Damage Signal:
- VERY EARLY
- The inflammatory process begins with VASCULAR COAGULATION (clotting) and the DETECTION of pathogens or cellular injury by PATTERN RECOGNITION RECEPTORS (PRR)
- MAST CELLS release HISTAMINE
2) LEUKOCYTE RECRUITMENT and ELIMINATION Stimuli
- Signaling through PRR induces RELEASE of Inflammatory mediators which ACT ON BLOOD VESSELS (Endothelial cells) to PROMOTE recruitment of Leukocytes and EXUDATION of plasma into the DAMAGED TISSUE
- PHAGOCYTOSIS of OPSONIZED pathogens and cellular debris!!!!!
3) RESOLUTION:
- After ELIMINATION OF MICROORGANISMS and necrotic tissue, Leukocyte Recruitment ceases and apoptotic neutrophils are phagocytized by Macrophages
* *** MACROPHAGES clean cell debris using SCAVENGER RECEPTORS (another type of PRR)!!!!!!!!
4) WOUND HEALING:
- Tissue repair and remodeling involves the development of NEW BLOOD VESSELS (Angiogenesis), RESURFACING of the WOUND (Re-epithelization) and COLLAGEN DEPOSITION
* *** MACROPHAGES stimulate Fibroblasts by releasing TGF-beta!!!!!!1
Inflammation in Atherosclerosis
1) Monocytes recruites though the activated endothelium differentiate into MACROPHAGES
2) Several endogenous and microbial molecules are recognized by TLRs and ACTIVATE THE CELLS
3) Inflammatory CYTOKINES, CHEMOKINES, ROI, NO, and other inflammatory molecules are released
4) Inflammation and tissue DAMAGE OCCURS
5) MACROPHAGE accumulate LIPIDS and become FOAM CELLS
Apoptosis vs Necrosis
- Cell Death may occur by Necrosis or Apoptosis:
1) NECROSIS: is a DIRTY form of cell death characterized by SWELLING and RUPTURE of cell membrane (cell lyse) which may cause INFLAMMATION or harm other neighboring cells
2) APOPTOSIS: is a CLEAN form of cell death in which any harm done to the organism by this process is MINIMIZED!!!
- Apoptotic cells are removed WITHOUT TISSUE INFLAMMATION
- Apoptosis is NORMAL PHYSIOLOGICAL PROCESS occurring in embryonic development and immune response
- DANGER SIGNALS are NOT RELEASED
- When Ag is ELIMINATED, then only MEMORY CELLS left
Apoptosis and Anti-Inflamamtory Response
Immunological Tolerance is the process by which the Immune System does not respond to Ag*
- Apoptosis provide NO DNAGER SIGNALS —> NO Immune Response
- HMGB1 is NOT RELEASED during Apoptosis
- APOPTOSIS can render APCs into a Tolerant State
- Anti-inflammatory IL-10, TGF-beta, and Regulatory T cells may contribute to the process
Molecular Mechanisms of Apoptosis
1) Triggers:
- DNA Damage
- Cytokine Starvation
- Hypoxia
- Temperature
- Death Receptor
2) Regulators: PRO-APOPTOTIC: - Death Domain Factors - Cytochrome C - p53 ANTI-APOPTOTIC: - BCL-2 Family - Myc/ Oncogenes
3) Executioners
- Capsase 9 (Intrinsic) and Capsase 8 (Extrinsic) activate CASPASE 3 for cell death!!!!!
- Each cell express Fas, but only ACTIVATED LYMPHOCYTES express Fas LIGAND (FasL)
Caspases in Apoptosis
- Caspases are the Major Executioners
- Caspases are CYSTEINE PROTEASES
- Caspases directly and indirectly ORCHESTRATE the MORPHOLOGIC CAHNGES of the cell during APOPTOSIS
- Capsases exist as LATENT PRECURSORS
- When activated, Caspases initiate apoptosis by DESTROYING KEY COMPONENTS of the Cellular Infrastructure
Intrinsic Mitochondrial Pathway
3 Main Triggers:
1) Bcl-2 Family proteins
2) CALCIUM!!!!!!!!
3) FREE RADICALS!!!!!!!
4 Downstream Regulators:
1) CYTOCHROME C!!!!!!!!!
2) Protein Smac/Diablo
3) Apoptosis-inducing factor
4) Endonuclease G
EXECUTIONERS:
- Caspase 9
- CASPASE 3!!!!!!!
- Apaf-1
Extrinsic Apoptotic Pathway
1) Fas Ligand binds to Fas!!!!
2) FADD recruits to the intracellular portion of the Fas receptor (Death Domain)
3) Caspase 8 is ACTIVATED and activates CASPASE 3 (Caspase Cascade)!!!!!!
- Once Caspase 3 is made there is no turning back
4) APOPTOSIS
- Mitochondrial damage
- Membrane Changes
- Proteolysis
- Nuclear Condensation and DNA fragmentation
Apoptosis In Action
-
Once Viral CLEARANCE happens then the CTL Response will also decrease**
- CTL will go through APOPTOSIS when there is no more Viral Antigens
Autoimmune Lymphoproliferative Syndrome (ALPS)
*** The Majority of ALPS patients have HETEROZYGOUS MUTATIONS in the gene encoding fas
Patients have:
1) CHRONIC ADENOPATHY and/or SPLENOMEGALY in the early years of life
2) CHRONIC PERSISTENCE and ACTIVATION of BOTH T CELLS that stimulate B-CELL maturation (Abs in BLOOD)
3) The EXTENDED SURVIVAL of LYMPHOCYTES due to the DEFECTIVE Fas-mediated APOPTOSIS may allow MALIGNANT TRANSFORMATION to occur!!!!!
Overview of Immune Responses to Microbes
- Survival and Pathogenicity of microbes is influenced by the ability of microbes to EVADE or RESIST the effector mechanism of immunity
- Many microbes establish LATENT, or PERSISTENT, INFECTIONS in which the immune response controls but DOES NOT ELIMINATE the microbe and the microbe survives WITHOUT PROPAGATING the INFECTION
- Tissue INJURY and disease may be caused by the host response to the MICROBE (COLLATERAL DAMAGE) RATHER THAN by the microbe itself
- Inherited and acquired DEFECTS in innate and adaptive IMMUNITY are important causes of SUSCEPTIBILITY to INFECTIONS
What happens without Immunity?
- Lack of INNATE IMMUNE MECHANISMS leads to very quick expansion of the pathogen
- Without ADAPTIVE IMMUNITY the infection is initially controlled by Innate Immunity, however, the PATHOGEN can NOT be ELIMINATED completely
Immune protest against Extracellular Microbes
1) Antibodies are PRESENT
2) Complement is PRESENT
3) Neutrophils are PRESENT
4) Th2 are PRESENT and ACTIVE (DOMINATES!!!!!!!)
5) Th1 small number present so NOT MUCH ACTIVITY
6) Macrophages are present in a low number
7) Cytotoxic T cells are NOT PRESENT
8) NK Cells are PRESENT along with CYTOKINES
Defenses Against Extracellular Bacteria
- Complement
- Phagocytes
- Antibodies
Steps of Immune Response Gains Infections with Extracellular Bacteria (Steps 1 -6)
1) Break in epithelial surface allows BACTERIA ENTRY and PROLIFERATION
2) Surface LIPOPOLYSACCHARIDES may activate the ALTERNATIVE COMPLEMENT pathway or MANNAN-BINDING LECTIN pathway leading to BACTERIAL LYSIS
- Other complement activators operating at this stage include C-REACTIVE PROTEIN, which binds Bacterial COAT POLYSACCHARIDES
3) MAST CELL DEGRANULATION enhance BLOOD Flow!!! The increased blood flow and LOCAL EDEMA are perceived as ITCHINESS and IRRITATION in the inflamed areas
4) ROLLING, MARGINATING Neutrophils adhere to the vein wall as locally released cheekiness and bacterial derived molecules (ENDOTOXINS) ACTIVATE both the ENDOTHELIUM and the NEUTROPHILS, resulting in ADHESION BETWEEN THE TWO
5) BACTERIAL PRODUCTS (Formyl-Methionyl-Leucyl-Phenylalanine tripeptide), COMPLEMENT FRAGMENTS (C5a) and CHEMOKINES (IL-8/CXCL8) ATTRACT Neutrophils to the site (CHEMOTAXIS)!!!!!!!!
6) Opsonized Bacteria are RAPIDLY ENGULFED and KILLED by Neutrophils. DENDRITIC CELLS Engulf and Internalize Bacteria, are ACTIVATED via PATTERN RECOGNITION RECEPTORS (Toll-Like Receptors) and MIGRATE via the LYMPHATICS
Steps of Immune Response Gains Infections with Extracellular Bacteria (Steps 7-11)
7) Dendritic Cells enter the Local LYMPH NODES and moves to GERMINAL CENTER.
- Local Inflammation leads to UPREGULATION of ADHESION MOLECULES on high Endothelial VENULES of Lymph Node, and Lymphocytes enter directly from the blood
- Many lymphocytes become TRAPPED in the LOCAL INFLAMED NODE, and the consequent SWELLING, along with local HYPEREMIA, leads to the symptom of SWOLLEN PAINFUL/TENDER LYMPH NODES!!!!!
8) Th cells are RECRUITED and ACTIVATED by PROFESSIONAL APCs in the Lymph Node, and by B CELLS, promoting the production of BACTERIA-SPECIFIC ANTIBODIES
- Naive T Cells become differentiated towards Th1 and Th2, according to the DENDRITIC CELL SIGNALS
- Initially IgM CLASS AB is PRODUCED, followed by CLONAL EXPANSION and SWITCHING to other classes (IgG or IgA) for MUCOSAL PATHOGENS
9) Early ANTIBACTERIAL AB production is of the IgM CLASS.
- This relatively LOW AFFINITY interaction is enhanced by the FIVE ADHESION sites on IgM, leading to HIGH EFFICIENCY of BINDING
- IgM is a VERY POTENT Complement ACTIVATOR and OPSONIN
- Opsonized bacteria are LYSED by Complement
10) Following the resolution of the BACTERIAL INFECTION, protective mechanisms for future encounters are put in place by laying down of MEMORY CELLS and PRODUCTION OF ABs
11) In the Resolution of an infection, bacterial debris is removed by LOCAL MACROPHAGES and NEUTROPHILS, or by ANTIBODY as SOLUBLE immune complexes!!!
- IgM is not a good SOLUBLE Ab because it is a pentamer and does not go into tissue easily
- IgG is more likely to go into the tissue because of its Monomer Configuration
Immunity to Extracellular Bacteria
- EXTRACELLULAR bacteria are capable of REPLICATING OUTSIDE host cells in the blood, Conn Tissue, Epithelial surfaces, the GI Tract etc
PATHOGENIC Extracellular Bacteria have 2 MECHANISMS:
1) INFLAMMATION causes tissue DESTRUCTION at side of infection
2) Bacteria produce TOXINS which ahem diverse Pathological effects
Bacterial Toxins have 2 Categories:
1) Endotoxins:COMPONENTS OF BACTERIA CELL WALLS
2) Exotoxins: SECRETED BY THE BACTERIA
- ENDOTOXIN (LPS) of Gran Neg bacteria is a POTENT ACTIVATOR of Macrophages, Dendritic Cells, and Endothelial cells
- Many EXOTOXINS are CYTOTOXIC including:
1) DIPHTHERIA TOXIN: Shuts down protein synthesis in infected cells
2) CHOLERA TOXIN: Interferes with ion/ water transport
3) TETANUS TOXIN: Inhibits Neuromuscular transmission - Other Exotoxins interfere with Normal Cellular functions without killing cells, and yet other EXOTOXINS STIMULATE THE PRODUCTION OF CYTOKINES THAT CAUSE DISEASE
Innate Immunity to Extracellular Bacteria
PRINCIPLE MECHANISMS:
1) Complement Activation
2) Phagocytosis
3) Inflammation
Complement Activation
- PEPTIDOGLYCANS (Gram Pos Bacteria) and LPS (Gram Neg Bacteria) activate complement by the ALTERNATIVE PATHWAY
- Bacteria that EXPRESS MANNOSE on their surface may bind Mannose-Binding Lectin, which activates complement by the LECTIN PATHWAY
1) MAC lyses Bacteria (Neisseria) are particular susceptible to LYSIS
2) C3a and C5a stimulate Inflammatory responses by RECRUITING and ACTIVATING LEUKOCYTES
3) Complement Activation results in OPSONIZATION and enhanced PHAGOCYTOSIS of the bacteria
Complement Mediated Killing
1) CLASSICAL Pathway:
- Requires IgM or IgG; not exactly an INNATE IMMUNE RESPONSE
- C3 Convertase: C4bC2a
- C5 Convertase: C4bC2aC3b
2) MBL Pathway:
- Does not use C1!!!!!!
- When MBP binds MASPs are ACTIVATED
- C3 Convertase: C4bC2a
- C5 Convertase: C4bC2aC3b
3) ALTERNATIVE Pathway:
- C3 undergoes spontaneous HYDROLYSIS and C3 CONVERTASE (C3bBb) is formed presence of Factors B and D!!!!!!!!!!!!!!
- Eventually C5 CONVERTASE (C3bBbC3b) is formed!!!!
- PROPERDIN stabilizes CONVERTASE
Complement Role In Host Defense
1) COMPLEMENT-MEDIATEDKilling
- C3b binds to microbe
- Activation of late components of complement
- Formation of the MAC
- OSMOTIC LYSIS OF MICROBES
2) OPSONIZATION and PHAGOCYTOSIS
- Binding of C3b (or C4b) to microbe OPSONIZATION
- Recognition of bounds C3b by PHAGOCYTE C3b receptor (CR1!!!!!!!!)
- Phagocytosis of microbe
Phagocytes and Inflammation
- Neutrophils and Macrophages use surface MANNOSE RECEPTORS and SCAVENGER RECEPTORS to recognize extracellular bacteria, and they use FC RECEPTORS and COMPLEMENT RECEPTORS to recognize bacteria opsonized with ABS and COMPLEMENT PROTEINS
- Microbial products activate TOLL-LIKE RECEPTORS (TLRs) on Phagocytes and other cells
- MANNOSE and SCAVENGER receptors PROMOTE PHAGOCYTOSIS of the MICROBES
- TLRs stimulate the MICROBIAL ACTIVITIES of the Phagocytes (mainly TLRs)
- The FC and COMPLEMENT RECEPTORS promote PHAGOCYTOSIS and ACTIVATION of the phagocytes
- Activated phagocytes SECRETE CYTOKINES, which induce Leukocyte infiltration into site of infection (Inflammation)
- The RECRUITED LEUKOCYTES ingest and DESTROY the BACTERIA
Opsonization and Phagocytosis Components
- CR1 is a common receptor for Complement
- CR2 binds to C3d and only needs a small amount of C3d to activate the Complement Cascade
- CR1-4 are Complement Receptors
- Fc receptors bind to the FC REGION of the Antibodies
Opsonization and Phagocytosis Steps
1) Recognition and Attachment
- Microbes bind to phagocyte receptors
2) Engulfment
- Phagocyte membrane zips up around microbe
3) Killing and Degradation
- PHAGOSOME of ingested microbe fuses with a LYSOSOME to form a PHAGOLYSOSOME
- The ROS, NO in the Phagolysosome cause for the microbe to be DEGRADED
- PRR-mediated PHAGOCYTOSIS IS SHOW!!!!!
- Complement and Ab-mediated Phagocytosis has the same mechanism
Microbicidal Mechanisms of ROI and NO
Harm Effectos of ROI:
1) Damage of DNA
2) Lipid Peroxidation
3) Oxidations of Amino Acids in proteins
4) Oxidative Inactivation of enzymes by Oxidation of co-factors
Peroxynitrite (ONOO-) Causes Apoptosis or Necrosis
1) ONOO- causes NECROSIS by:
- Lipid Peroxidation
- Protein Oxidation
- Protein Nitration
- Inactivation of enzymes
2) ONOO- causes APOPTOSIS by:
- Caspase activation by the release of Cytochrome C from the mitochondria
- AIF activation from the mitochondria
- Peroxynitrite is a result of L-arginine losing a NO and converting to Citrulline!!!!!!!
- NO + O2- = ONOO-
Innate Immune Evasion by Extracellular Bacteria
1) Inhibition of Complement Activation
- Many bacteria
2) Resistance to Phagocytosis
- Pneumococcus, Neisseria meningitidis
3) Scavenging of reactive oxygen Species
- Catalse- Positive Staphylococci
Role of Abs in Elimination of Microbes
1) Neutralization of Microbes
2) Opsonizatoin and Phagocytosis of microbes
- Fc gamma Receptor on Phagocyte
3) Antibody dependent cellular Cytotoxicity
- NK cells
4) Lysis of Microbes
5) Phagocytosis of Microbes Opsonized with Complement Fragments (C3b)
6) Inflammation
Neutralization of Bacteria and Toxins by ABs
1) Antibody blocks BINDING of Microbe and INFECTION of cell
2) Antibody blocks INFECTION of ADJACENT cell
3) Antibody blocks BINDING of TOXIN to CELLULAR RECEPTOR
Ab Mediated Phagocytosis
**IgG**
1) Opsonization of Microbe by IgG
2) Binding of Opsonized Microbe to Phagocyte Fc Receptors (FcGammaRI)
3) Fc receptor signals activate phagocyte
4) Phagocytosis of Microbe
5) Killing of ingested Microbe
Fc Receptors and Phagocytic Cells
- Phagocytes like Macrophages and Neutrophils express Fc receptors for IgG (FcGammaRI) and IgA (FcAlphaRI)!!!!!!!!!!!!
- FcgammaRI and FcalphaRI interact with Ab-Ag complexes only
- Abs bind to PATHOGEN or TOXIN to form COMPLEXES
- COMPLEXES are removed by PHAGOCYTIC CELLS (Neutrophils, Monocytes, and Macrophages) via FcGammaRI and FcAlphaRI
EXCEPTION:
- MAST CELLS, Eosinophils, and Basophils express FcEpsilonRI that most strongly binds IgE!!!!!!!
- FcEpsilon is unique because it bids FREE IgE (NO AG REQUIRED)!!!!!!!!
IgG
* 200 Binding Strength*
- FcGammaRI!!!!!!!
Cell Type:
- Macrophages
- Neutrophils
- Eosinophils
- Dendritic Cells
- Effect of IgG ligation:
- Uptake
- Stimulation
- Activation of Respiratory Burst
- Induction of Killing
IgE
* 20,000 Binding Strength*
- FcEpsilonRI!!!!!!!
Cell Type:
- Mast Cells
- Eosinophils
- Basophils
- Effect of IgE ligation:
- Secretion of Granules
IgA1, IgA2
* 20 Binding Strength*
- FcAplhaRI!!!!!!!
Cell Type:
- Macrophages
- Eosinophils
- Neutrophils
- Effect of IgE ligation:
- Uptake
- Induction of Killing
Bacterial Strategies to Evade Immunity
1) Antigenic Variation:
- Bacterium or virus alters its surface proteins in order to evade a host immune response
- Neisseria gonorrhoeae
- Escherichia coli
- Salmonella typhimurium
2) Inhibition of Complement:
- Bacterial capsules, decay of C3 Convertase, Blocking MAC formation (VITRONECTIN)
- Many bacteria
3) REsistance to Phagocytosis:
- Interfere with Complement activation and/or deposition at bacterial surface, injection of anti-phagocytic effectors into the cell
- Pneumococcus
4) Scavenging ROI:
- CATALSE Positive pathogens deactivate the Peroxide radicals and thus survive unharmed within the host
- Catalase- Positive Staphylococci
Diversity in Immune System
1) Immunoglobulins
2) T Cell Receptors
3) Major Histocompatibility
- A combination of shuffling of gene segments by SOMATIC RECOMBINATION and High MUTATIONAL VARIABILITY for some segments, leads to a LARGE GENETIC DIVERSITY for ANTIBODY PRODUCTION
- Theoretically 10^14 different possible antibodies
Development of Immunity
- During development, individual segments of certain genes are rearranged into different combinations, producing immune cells that contain different genetic information such that each is adapted to attack ONE PARTICULAR ANTIGEN
- Mechanism requires relatively FEW GENES to begin with but provides an enormous diversity of cells capable of responding to an equally ENORMOUS NUMBER of FOREIGN Substances
Immune Response B cells
- Humoral Immunity = B Cells
- Each Initial B cell undergoes genetic rearrangement so that is it will recognize and respond to ONLY ONE KIND OF FOREIGN ANTIGEN
- Each AB produced by plasma cell, will only respond to and BIND TO A SPECIFIC ANTIGEN. Receptors specific for a given ANTIGEN will also be produced on the surface of B cells
- Immune response occurs when appropriate Antigen binds to one of these receptors
Immune Response T cells
- Cellular Immunity = T Cells
- They develop receptors on the cell surface that response to a unique Antigen
- T cells work in concert with Macrophages, which initially recognize an Antigen and which also present pieces of the antigen to appropriate T cells
- In order for T cells to function properly they must bind to a SPECIFIC ANTIGEN and bind to SELF ANTIGEN, a MAJOR-HISTOCOMPATABILITY COMPLEX
- The bidding to the MHC ANTIGEN helps to mark that particular T cell as one that has UNDERGONE SELECTION during embryonic development WITHIN the THYMUS
Immune Response Proteins
1) Immunoglobulins
2) T Cells Receptors
3) Major Histocompatibility Antigens (HLAs)
- Same plasma cell can secrete one or more kinds of immunoglobulins (different isotope or classes), but they will all respond to the same antigen
Isotype: Classes of Ab
Idiotype: Affinity for Ag
Immunoglobulin Structure
Immunoglobulins (Antibodies):
- Synthesized by B cells
- Synthesized and secreted by Plasma Cells:
- Terminally differentiated B cells
- IMMUNOGLOBULINS with specificity for an Antigen are referred to as ANTIBODIES
Immunoglobulin Structure
- Consist of FOUR POLYPEPTIDES linked by DISULFIDE BONDS:
- Two Identical HEAVY CHAINS (VDJC)
- Two Identical LIGHT CHAINS (VJC)
Light Chains
- Immunoglobulin molecule contains two identical light chains (isotopes):
1) Either TWO IDENTICAL KAPPA Chains:
- Encoded on Chromosome 2!!!!!!
2) Or TWO IDENTICAL LAMBDA Chains:
- Encoded on Chromosome 22 - NEVER ONE OF EACH TYPE!!!!!!!!!!!!!!1
Light Chain Domains:
- Variable (VL) domain (110AA) - Constant (CL) domain (110AA)
Heavy Chains
- Immunoglobulin molecule contains two IDENTICAL HEAVY CHAINS
- Encoded on Chromosome 14
5 TYPES of HEAVY CHAINS (Isotypes):
- Mu, Delta, Gamma, Epsilon, and Alpha - M,D,G,E,A
Heavy Chain DOMAINS:
- One Variable (VH) Domain
- Three or Four Contant (CH) Domains
- D,G, and A chains have THREE CONSTANT DOMAIN (CH1-3)!!!!!!!!!!!!!!!
- M and E have an ADDITIONAL FOURTH CONSTANT DOMAIN (CH1-4)!!!!!!!!!!!!!!!!!!!!!
Antigen Receptors of Lymphocytes
B Cell Receptors: (IgM is first Ab produced)
- Immunoglobulins
- May be MEMBRANE BOUND
- May be SECRETED
T Cell Receptors:
- TCRs
- Exist only as MEMBRANE BOUND RECEPTORS
TCR Maturation
1) Double NEGATIVE (CD4-/CD8-) Seen in the SUBCAPSULAR CORTEX region of the THYMUS!!!!!!
2) T cells then rearrange gene coding TCR and express both becoming DOUBLE POSITIVER (CD4+/CD8+), and they are seen in the CORTEX of the THYMUS!!!!
3) T cells whose receptors bind self-MHC molecules lose expression of either CD4 or CD8 and INCREASE levels of expression of TCR. These are called SINGLE POSITIVE T CELLS (CD4+/CD8-) or (CD4-/ CD8+) and are seen in the MEDULLA of the THYMUS!!!!!!!!
How Helper T Cells Help
1) A Macrophage (APC) processes phagocytised Ag that is then bound to CLASS II MHC and presented to T HELPER CELLS (Cell- Mediated Immunity)!!!!!!!!!
2) Activated Th cells divide by Mitosis to Incr population of Th cells at site of Ag presentation
3) Th cells INTERACT with B cells in the presence of APC to INDUCE 1) Immediate access of B cells to FREE Ag in Extracellular space 2) PROLIFERATION of B Cells
4) B cells with CELL SURFACE-SPECIFIC Immunoglobulin arrive at the site to RAPIDLY NEUTRALIZE free Ag
5) B cells DIFFERENTIATE into PLASMA CELLS that Secrete Immunoglobulin to BLOCK FREE Ag (Humoral Immunity)
Mechanisms for Generating Ag Receptor Diversity
1) Somatic Recombination
- COMBINATIONAL DIVERSITY
2) Junctional Diversity
3) Somatic Hypermutation
Germline Organization
1) Ig H CHAIN (Chromosome 14)
- C (mu) is always the first CONSTANT REGION expressed!!!!! (IgM produced first)
2) Ig Kappa Chain Locus (Chromosome 2)
- Has no D REGIONS!!!!
- Has ONLY ONE CONSTANT REGION
3) TCR Beta Chain Locus (Chromosome 7)
- Has D REGION
4) TCR Alpha Chain Locus (Chromosome 14)
- Has NO D REGION!!!!
SOmatic Recombination
Expression of an Ig Heavy Chain involved TWO GENE RECOMBINATION EVENTS:
1) D-J JOINING 2) V to DJ COMPLEX
- Intervening gene segments are DELETED
- Recombined gene is TRANSCRIBED
- VDJ SEGMENT IS SPLICED onto the FIRST HEAVY CHAIN RNA!!!!!!
- The mRNA is TRANSLATED to produce HEAVY CHAIN PROTEIN
- Ig LIGHT CHAIN (and TCR Alpha/ Beta Chains) follow the SAME sequence EXCEPT in the Ig LIGHT CHAINS and the TCR Alpha chain a V gene COMBINES DIRECTLY with a J GENE SEGMENT
- Light chains and TCR Alpha have NO D REGIONS!!!!!!!!
Light Chains: VJC
- B cells commits to Light Chain
- One V brought next to One J region gene
- RECOMBINATION
- V gene PROMOTER is brought CLOSE to ENHANCER between J and C regions
- mRNA TRANSLATION and L is removed as protein is transported to ER
- Light Chain and Heavy Chain ASSEMBLE IN ER!!!!!!!!!
Gene Segments
- Genes coding for the different chains are composed of Segments
- Segments for the Human Kappa Gene:
- 30-35 V (variable) segments
- 5 J (Joining) segments
- 1 C (Constant) segment
Kappa/ Lambda Gene Organization
1) Lambda Light Chain
- Have 4 Constant Regions!!!!!!!
2) Kappa Light Chain
- Has 1 Constant Region!!!!!!!!!
Kappa Light Chain Genes
1 Constant Region!!!!!!
- Each Variable Region gene as two Exons:
1) One LEADER exon
2) One VARIABLE exon - Joining region between V and C
Lambda Light Chain Genes
4 Constant Regions!!!!!!
- One for EACH SUBTYPE!!
- Each variable region as TWO Exons:
- One LEADER
- One for MOST VARIABILITY
- Joining region between V and C!!!
Light Chains
- Most variable of all the regions in the light and heavy chains is the Fab region
- This VARIABILITY is produced through JUNCTIONAL RECOMBINATION
1) Some Genes in the V region can be deleted (PERMANENT)
2) Then the V and J regions will join together
3) An mRNA TRANSCRIPT will be made from this Light chain segment
4) Then there is a DELETION in the mRNA bringing the VJ and C gene TOGETHER!!!!!!!!
* Ig Light Chains and TCR Alpha Chains LACK D REGIONS therefore the V region recombines DIRECTLY with J!!!!!!**
Formation of Ig Heavy Chains
1) Germline DNA at Ig H Locus
2) SOMATIC RECOMBINATION (D-J Joining) in TWO B CELL Clones
3) Then Transcription occurs so the Primary RNA is formed
4) The Primary RNA is spliced to form the mRNA!!!!!
5) The mRNA is TRANSLATED into Ig Mu (M) Chains in TWO B Cell Clones
Valence: How many Immunoglobulins Ag can bind to. Most Immunoglobulins have a Valence of 2
IgM is secreted in the PENTAMER FORM and when it is Membrane bound it is in DIMER FORM
Somatic Recombination
- Gene REARRANGEMENT that is occurring in SOMATIC CELLS
Combinational Diversity:
- TCR = 3 x 10^6 possibilities
- Ig = 10^6 possibilities
Junctional Diversity:
- TCR 10^16 possibilities
- Ig 10^11 possibilities
Heavy Chain VDJC
1) DJ Rearrangement
2) VDJ Rearrangement
3) TRANCRIPTION (Primary RNA)
Junctional Diversity
Diversity of Ag Receptor is Incr by:
1) COMBINATIONAL DIVERSITY
- Due to SOMATIC RECOMBINATION
- Involves V(D)J RECOMBINASE!!!!!!!!!
- Limited by Number of available V,D, and L gene segments
2) JUNCTIONAL DIVERSITY
- Almost UNLIMITED
- Due to:
- REMOVAL of Nucleotides from V, D, and J segments by EXONUCLEASE at time of Recombination
- TERMINAL DEOXYRIBONUCLEOTIDY TRANSFERASE (TdT)!!!!!!!!!!
V(D)J Recombinase
Collection of Enzymes:
- Expressed in IMMATURE B and T Lymphocytes
- Recognizes DNA sequences that FLANk all Ag Receptor V,D, and J gene segments
- BRINGS THESE SEGMENTS CLOSE TOGETHER
- Constists of:
- Recombinase-Activating Gene (RAG)-1
- Recombinase- Activating Gene (RAG)-2 - EXONUCLEASE cuts DNA at the ENDS of the segments
- LIGASES repair the Breaks
Recombination SWITCH Sequence:
- Found next to every VDJ Segment
- Consists of THREE Elements:
- A HEPTAMER
- A NONAMER
- A SPACER of 12 or 23 bp
- RAG-1 and 2 bind to 12 bp and 23 bp SPACER
