immu2011: t + b cells

antigen

a substance recognised by lymphocytes

foreign to body

b cells

recognise free floating or membrane bound antigen

t cells

recognise peptides on MHC molecules

MHC molecules

surface molecules presenting peptide antigens

naive t cell

t cells from thymus, not yet exposed to antigens

t cell receptor

recognises a specific pathogen antigen

lymphocyte recirculation

movement of naive lymphocytes through lymphoid tissues

t cells re-circulate to continue to encounter foreign antigens

antigen encounter

bringing T cells and antigens to lymphoid tissues for interaction

maximising antigen encounter

DCs are strategically located in periphery where they capture antigens, become activated, and migrate to lymph node where they present antigen to naive T cell

lymph node

DC travel from periphery to the _____.

HEV = specialised vessels which connect lymph to blood

only DCs can activate naive T cells

antigen processing

breaking down captured proteins into peptides for MHC presentation

afferent lymphatic vessel

where the antigen enters from periphery and is transported to the lymph node where T cell are location

DC migration

via lymphatic vessels

during transport DC become activated and mature, allowing them to present antigen to T cells

antigenic determinants

small parts of pathogen molcules which are recognized by T cell receptors, couple aa’s long

AKA epitope

antigen recognition + presentation

1. antigen captured (by resident cells + DCs)

2. DCs carry antigen to lymph node via vessels, during transport DC mature allowing them to present to T cells

3. antigen presentation to receptors on T cells

antigen processing

proteins broken down into smaller peptides

peptides loaded onto special surface molecules (MHC) so T cells can “see” them

epitope

amino acids recognised by T cell receptors

MHC

major histocompatibility complex

responsible for displaying antigens to T cells

HLA molecule

human leukocyte antigen, another term for MHC molecules

the MHC in humans, HLA genes located on chromosome 6

polymorphic

having variations between individuals, highly expressed in MHC genes

MHC class 1

expressed by all uncleared cells

presents peptides to CD8+ t cells

MHC class 2

expressed only on antigen presenting cells (eg, DC)

presents peptides to CD4+ t cells

graft acceptance

determined by MHC

• identical MHC locus = acceptance

• dissimilar MHC = graft rejection

expression of MHC

co dominant

for each MHC - 1 allele from each parent

3 class loci for each class 1 gene = HLA-A, HLA-B, HLA-C - as co-dominant, each DC expresses 6 MHC class 1 types

3 class loci for class 2 → HLA-DP, HLA-Dq, HLA-DR

peptide binding cleft

site on MHC molecules where peptides are accommodated for T cell recognition

class 1 MHC locus

• HLA-A

• HLA-B

• HLA-C

3 loci, codominant expression = 6 molecules expressed on cell surface

class 2 MHC locus

1. HLA-DR

2. HLA-DQ

3. HLA-DP

3 loci, codominant expression = 6 molecules expressed on cell surface

MHC class 1 structure

• 1 alpha chain

• a1 and a2 is where polymorphic residues are, they also form a closed peptide binding cleft

• cleft may accomodate aa 8-11 long

• a3 is invariant, contains site for CD8+ t cells only, can only activate CD8 t cells

• a chain is non-covalently associated w/ B2 microglobulin, a protein which is encoded by gene outside MHC

CD8

T cell co receptor binding to MHC class 1, restricting CD8+ t cell recognition

CD4

t cell co-receptor binding to MHC class 2, restricting CD4+ t cell recognition

MHC class 2 structure

• an alpha and beta chain

• a1 and b1 = polymorphic residues, form (open conformation) binding cleft

• cleft accommodates 10-30 aa’s

• b2 = binding site for CD4+ only

• a2 and b2 chains anchor MHC to membrane of APCs (DCs, B cells, macrophages)

antigen processing pathways

processes where MHC class 1 presents intracellular antigens while class 2 presents extracellular antigens

note - DC may express class 1 or class 2 MHC

proteasome

cellular complex degrading proteins into peptides for MHC presentation

TAP

transporter associated with moving peptides into ER for MHC loading

tapasin

molecule assisting in the correct folding of MHC class 1 in ER

phagolysosomes

vesicle formed by fusion of phagosome with lysosome, degrades molecules

CLIP

invariant chain with CLIP occupies the peptide binding cleft in newly synthesised class 2 molecules, CLiIP sequence keeps molecule stable and blocks other peptides form binding until MHC class 2 is fully synthesized

ubiquitin

tagging protein for degradation in the proteasome during MHC class 1 pathway

DM

protein in late endosomes exchanged CLOP for higher affinity peptides in class 2 pathway

novel prize for MHC

awarded to peter doherty and rolf zinkernagel for MHC resistrction work

bare lymphocyte syndrome

condition caused by TAP deficiency leading to susceptibility to infections

MHC class 1 pathway

• proteins sourced from cytosol

• proteins are unfolded and tagged with ubiquitin for degradation

• proteasome shed the proteins into peptides - enhanced by inflammatory cytokines (TNF, IL-1)

• peptides actively moved by TAP into ER

• in ER class 1 molecules are synthesised - a chain (folding assisted by chaperone molecules), b2 microglobulin

• newly formed class 1 MHC molecule binds to TAP via tapasin

• peptides are loaded into the MHC class 1 groove

• peptide loaded MHC class 1 exits to the golgi to be packaged into exocytic vesicles

• presentation to CD8+ MHC class 1 restricted lymphocytes

MHC class 2 pathway

1. extracellular antigens (bacteria, parasites, fungi) recognised by PRR

2. internalised by phagocytosis into phagosome/endosomes

3. endosomes fuses w/ lysosome, which contains proteolytic enzymes + acidic pH

4. degrades proteins into peptides

AT same TIME

• a and b chains of MHC 2 are being synthesised in ER

• invariant chain (Ii) with CLIP occupies binding cleft in newly synthesised class 2

• class 2 transported out of ER via golgi in a exocytic vesicle (brings MHC class 2 and degraded proteins together)

• enzymes in the late endosomes/lysosomes also contain a class 2 MHC-like protein called DM

• class 2 molecules loaded with antigen and transported to cell membrane

• presentations to CD4+

humoral immunity

a type of adaptive immune response primarily involving antibodies, which are proteins produced by B cells

involves macromolecules in “humours” i.e bodily fluids secreted antibodies, complement proteins, some antimicrobial

B cell role

production of antibodies

common lymphoid progenitor

neutralise + eliminate EC microbes + microbial toxins

antibodies can’t enter cells, so IC microbes not eliminated, need cell mediated immunity

plasma cells

activated B lymphocytes producing antibodies, memory b cells

X linked agammaglobulinemia

genetic disorder affecting B cell development, causing susceptibility to infections

severe recurrent infections, particularly in response to encapsulated bacteria (b cells particularly good at eliminating these)

poor vax response - as no antibodies

vaccination response

immune systems reaction to provoke antibody production

b cell cascade of events

antigen recognition ( b cells) → proliferation (activated) → differentiation (plasma cells) → memory

b cell origin

begin life as a cell surface bound receptor (b cell receptor/BCR), also known as bound antibody

antigen specific receptors

b cells have diverse __________.

can recognise ~10^11 antigens

receptors generated by somatic recombination of antigen receptor gene segments

note - for innate cells, PRR only recognise ~1000 via germline encoded receptor

B cell receptor (BCR)

antibody precursor on b cell surface

somatic recombination

process generating diverse antigen receptors

bursa of fabricus

site of b cell development in birds

b cell receptor structure

immunoglobulin, 2 binding sites

• membrane bound

• constant region (purple) + variable regions (other colours)

• variable region binds to antigen

• ~10^11 possible BCRs, space constraints dictate that frequency of cells specific for one antigen is very low (~1:100 - 1:10,000)

• after activation - cells undergo clonal expansion so response is slower than innate response

TCR = t cell receptor

• membrane bound

• constant = purple, variable = other

BCR vs. TCR

• genetically related (similarities between heavy chain and B chain, light chain and alpha chain)

• structurally and functionally unique

• antigen receptors of any one cell all have one specificity

• each cell expresses thousands of this one receptor on their cell surface

TCR - antigen recognition

• recognise processed peptide antigen, presented by MHC molecules

• linear epitopes

• TCR binds to both peptide and MHC

BCR - antigen recognition

• recognise ‘free’ antigens or antigens ‘delivered’ by other cells

• linear + conformational epitopes

• can be protein, lipids, carbs, nucleic acids (native form)

• more recognition than t cells

antibody isotopes

different antibody classes with distinct functions

IgG

most abundant antibody in human serum

antibody structure

4 polypeptide chains assembled in Y shape

• 2 identical light (L) chains - approx 25 kDa

  • light chain = 1 variable + 1 constant domain

• 2 identical heavy (H) chains - approx 50kDa

  • 1 variable + 3-4 constant domains

• 2 variable regions make up ‘antigen binding’ site

• hinge gives flexibility - allows variable regions to come closer together or further apart to bind epitopes

papain

a protease enzyme from papaya

used to dissect antibodies

cleaves molecule into two Fab fragments and one Fc region

Fab region

fragment - antigen binding region

Fc region

fragment, crystallizable region, binds to cell surface receptors (Fc receptors) and some proteins of complement

IgM

original antibody isotope, always first one produced

low affinity so assemble for 10 binding sites

abundance of isotypes

IgM - ~5%, M heavy chain

IgG - ~80%

IgA - ~15%

IgE - traces

IgD - ~0.25%

class switching

requires CD4 helper T cells to switch from IgM to other antibody classes

constant domain for each isotope is the same, different isotypes have different heavy chains which creates different effector functions

neutralisation

process where antibodies bind to pathogens to prevent entry

IgM, IgG, IgA (lumen of mucosal organs) can act as neutralisers

mucosal immunity

immune defence in GI, respiratory, urogenital tracts

IgA antibody, diners held together with J chain

IgA binds to poly-Ig receptor

opsonisation

process of tagging pathogens for phagocytosis

IgA defence mechanisms

complex is endocytosed into epithelial cells and transported across cell (transcytosis)

released into lumen by proteolytic cleavage

then neutralises microbes + prevents their entry

opsonisation - antibodies

“antibody dependent cell phagocytosis”

• IgG + IgA - Fc regions bind to Fc receptors (FcyR or FcaR) on phagocytes

ADCC

antibody-dependant cell-mediated cytotoxicity (cell killing mechanism)

ADCC process

• IgG binds to foreign antigens on surface of cells (infected cells)

• NK cells express FcyR3 that engages with IgG bound cell

• receptor mediated signals elicit release of granules containing toxic proteins

• Fc region binds to Fc receptors on NK cells (IgG)

IgE

antibody involved in allergic responses and parasitic defence

activates mast cells + basophils

Antibody + Helminths

• IgE combats parasites

• helminth (worms) are big and cannot be phagocytosed

  IgE opsonsises worms, FceRI on eosinophils binds IgE inducing eosinophil activation + release of cytoplasmic granules that kills the parasite

IgE role

• binds to FceRI on mast cells + basophils, coating with IgE = sensization

• antigen needs to bind to 2 or more FcR bound IgE to trigger cross linking of FcR + cell activation

• triggers release of inflammatory mediators (histamine)

• allergic responses, also physiological responses to protect from venoms + toxins

complement + antibodies

classical pathway activated by IgM + IgG antibodies

IgG1 and IgG3 most regular activators

lymphocyte development

process of B and T cell lineage commitment and receptor gene rearrangement

many selection events

selection events - lymphocyte development

process that preserves cells with useful receptors and eliminates self reactive ones

b cell development

process during which b cells develop and mature

pro b cell

cell starting point in b lymphocyte development

V(D)J recombination

process of gene segment rearrangement to create B cell receptor variability

VDJ recombinase

enzyme regulating gene rearrangement in immature B and T cells

junctional diversity

process adding random nucleotides to increase BCR diversity

positive selection

process determining if B cell receptor is functional and in frame

after all cutting and lighting of gene segments, is the VDJ segment in frame with constant region Cm segment), can it make a functional protein

no BCR = no survival signals

negative selection

process eliminating self-reactive B cells

did BCR that binds to self-antigen too strongly, if self reactive, B cells rearrange the light chain in a process called receptor editing

failing this step results in deletion

making a diverse BCR

1. rearrangement of gene segments by V(D)J recombination

   • random rearrangement of one V (variable) gene segment, one D segment (Ig H chain only) and one J gene segment to make a single V(D)J exon that will code for the variable region of the protein

2. introduce random nucleotides at the junctions between VDJ = junctional diversity

   • greatly increase diversity of receptors produced, with just VDJ = ~3×10^6 possible BCR, with JD = ~10^11 BCRs

   • addition of random nucleotides catalyses by TdT (lymphocyte specific enzyme)

3. positive + negative selection

4. migration to spleen as an immature B cell, differentiation into mature B cell

pro b cell BCR

Ig H chain (heavy chain)

Ig k/

pro B cell - BCR structure

Ig H locus (heavy chain) = variable region (V, D, J segments) + constant region - c

Ig K/¥ locus = variable region (V, D, J segments) + constant region (c ) - light chain

VDJ recombination - key points

1. ordered - D gene segment fuses to J gene segment, then V to DJ

2. its mediated by an enzyme only expressed in immature B and T cells - VDJ recombinase, cut DNA and bring segments close together

B1 cells

innate type of B cells

follicular B cells

major subset of B cells, responsible for antibody production

express surface bound IgM and IgD

recirculation of B cells

movement of mature (naive) B cells between lymphoid organs in search of antigens

BCR cross linking

trigger for B cell signalling upon antigen binding

b cells in lymph nodes

b cells located in follicles

T independent response

B cell response without T cell help, produces short lived plasma cells

T dependent response

B cell response requiring T cell assistance for class switching + affinity maturation

germinal centre reaction

process where B cells proliferate and undergo affinity maturation