Chapter 5 Antigen Recognition

by T lymphocytes
 • B cells and T cells recognize antigen through
variable antigen specific receptors
• B cells make Ab

B cell and T • T cells interact with other cells

• B cells bind epitopes on intact molecules, such
cell as proteins, carbohydrates and lipids
comparison • T cell receptors bind peptide antigens that are
derived from pathogen
 T Cell Receptor (TCR)
• Membrane bound
• Two different polypeptide chains= one antigen
binding site
• Gene rearrangement responsible for variability
– same mechanism used for immunoglobulins
• No somatic hypermutation or isotype switching
• TCR similar to Fab of immunoglobulin
• TCR= α chain and β chain
• Millions of different T cell receptor in healthy
human
TCR binding site

• Hypervariable regions on V
domains of α chain and β chain
have three loops CDR1, CDR2, and
CDR3
• TCRs have only one binding site
for antigen
• TCR recognition of antigen is
always in context of major
histocompatibility (MHC) complex
Generation
of T cell
diversity

When and where does this

happen?
 Diseases
associated with
RAG proteins
• Severe combined immunodeficiency disease (SCID)- one cause is
lack of RAG proteins
• “combined” because lack functional B and T cells
• “severe” because compared to immunodeficiencies that
lack only B cells
• Must have bone marrow transplants or die in infancy from
common infections
• Omenn syndrome- RAG protein with partial enzymatic activity
due to missense mutations
• Must have bone marrow transplants or die in infancy from
common infections
 α:β T Cell
receptor complex
• In ER α:β heterodimer associate with four
invariant membrane proteins
• CD3 complex -CD3γ, CD3δ, CD3ε (chromosome
11)
• Fourth protein is ζ chain (chromosome 1)
• Important for transmitting signal by
associating with intracellular signaling
molecules
• Nonfunctional CD3δ or CD3ε results in
immunodeficiency because transport of T cell
receptors is impaired. T cells have low number
receptors and impaired signaling
 γ:δ T cells
• Similar structure to
α:β receptor but
two different
protein chains
• T cells express one
or the other, not
both
 • δ gene segments located within α chain locus on chromosome 14
• Rearrangement of α chain locus = deletion of δ chain locus
• γ chain locus on chromosome 7
• Fewer variable gene segments in γ chain and in δ chain so γ:δ T

g and d
cell receptors are less diverse
• Off set by increased junctional diversity in δ gene and more
potential combinations

chain •
•
2 D segments incorporated between V and J
Junctional diversity seen between 2 D segments, V and

loci D segments and D and J segments

location
 • α:β T cells in birds, mammals, fish all recognize
peptide antigen bound to MHC molecule
•
Functional •
γ:δ T cell function not conserved
Major differences between γ:δ T cells in humans and
difference mice
• Similarities
between T • More abundant in tissue than in circulation

cell • Share properties with NK cells and other

lymphocytes
receptors • Recognition doesn’t depend on MHC molecules
and bound peptide, but can be involved
 T cell repertoire
more diverse than
Immunoglobulin’s
repertoire
 • TCR recognizes short peptide (8-25 aa)
produced from intracellular degradation
of pathogen’s proteins
• Pathogen can be phagocytized and killed
and its proteins processed or pathogen
such as Influenza virus can infect cell and
resulting viral proteins can be processed
Antigen • Major histocompatibility complex
presentation molecule (MHC)- binds peptide and then
to T cell MHC molecule transports peptide to
surface of cell
 Two classes MHC
• MHC class I present antigens from intracellular organisms
• Peptides from intracellular organisms degraded in the cytoplasm are delivered to ER
• MHC class I molecules in ER bind peptide and transport it to the surface of the cell

• MHC class II present antigen from extracellular pathogens

• MHC class II molecules travel to endosomal vesicles and bind peptide produced from
pathogens degraded by lysosomal enzymes.
• MHC class II molecule then transports peptide to cell surface

Cytotoxic T cells (CD8)

MHC class I
intracellular infections
Helper T cells (CD4)
MHC class II
extracellular infections
T cell co-receptors

• Cooperate with TCR in recognizing peptide:MHC

complex
• CD4 binds site on MHC II
• Single polypeptide
• 4 extracellular immunoglobulin like domains
• CD8 binds site on MHC I
• Heterodimer of two chains
• Each chain has one extracellular
immunoglobulin like domain
 Function of T cells
• CD8 T cells kill infected cells
• CD4 T cells help activate tissue macrophages
• increase phagocytic ability
• secrete cytokines and chemokines = drive adaptive immune response

• CD4 T cells help B cells

• B cells make high affinity Ab

• HIV selectively infects CD4 T

cells because it uses CD4 as a
receptor for entry into the cell
• Infection progresses resulting
in less circulating CD4 T cells
• Can become so low that
infected individual cannot fight
off other infections
 Structures of MHC
molecules

MHC class I molecule

• Transmembrane heavy chain (α chain)

• Three extracellular domains (α1, α2, α3)
• Peptide binding site formed by α1 and α2 folding (domains farthest
from membrane)
• α3 domain and β 2-microglobulin support
• Small protein (β 2-microglobulin)

MHC class II molecule

• Two transmembrane chains

• Each contributes one domain to peptide binding site

• Each contributes one domain for support

• MHC class I α3 domain also important for
binding CD8 coreceptor

• MHC class II β2 domain binds CD4 coreceptor

 MHC molecule
peptide binding
site
• Promiscuous binding specificity
• MHC class I
• length of peptide limited by pockets at ends of
peptide binding groove
• 8, 9, 10 aa in length- 9 aa most common
• Most bind peptide with hydrophobic or basic residue
at C terminal end depending on if MHC class I
binding pocket is complimetary to hydrophobic or
basic aa side chains
• MHC Class II
• Ends not pinned into pockets
• Peptides can be longer and more variable in length
compared to peptide binding MHC Class I
• 13-25 aa usually, but can be much longer
Peptide bound to MHC class I Peptide bound to MHC class II
 Trafficking of peptides

• Translation in cytoplasm- inefficient

• Improperly folded cell proteins targeted
to degradation and recycling
• Viral proteins in virally infected cells also
subject to degradation
• Peptides from degradation, traffic to ER
and bind MHC class I molecules.
• Peptide:MHC class I exported to plasma
membrane through Golgi
• Healthy cells= all peptides are self
• Infected cells= some peptides are from
pathogen (non-self)
• Material that comes into cells through
phagocytosis is broken down into
peptides in the lysosomes
• In the endosome peptides bind to MHC
class II molecules
• No extracellular infection= all self
peptides
• Extracellular infection= some non-
self peptides
MHC Class I
 Proteasome

• Proteasome can be modified in infected cells to

favor production of peptides that bind MHC class I
molecules
• IFNγ secreted by NK cells
• Alternative subunits replace β subunits
• Increased cleavage after
hydrophobic residues results in increased
number of peptides with hydrophobic or
basic aas at C terminus
Transporter associated with
antigen processing
• Transporter associated with antigen processing (TAP)
• Heterodimer TAP-1 and TAP-2
• Requires ATP
• 8+ aa long peptides with a hydrophobic or basic C-terminal region
• Most do not successfully bind MHC class I molecule, but are
transported out of ER into cytosol

• Bare lymphocyte syndrome type I (MHC class I deficiency)- non functional

TAP protein
• No peptides transported into ER
• Individuals have < 1% normal level of MHC Class I molecules on
surface
• Weak CD8 T-cell response to viruses

• Chronic respiratory infections from young age

 Peptide loading of
MHC class I
molecules

• MHC class I heavy chains

enter ER and bind calnexin
• Calnexin- C-type lectin-
Chaparone protein that
retains bound protein in ER
until subunits have folded
and associated correctly
• MHC class I heavy chain
released from calnexin once it
has folded, formed disulfide
bonds, and bound β2-
microglobulin
• Incorporated into peptide-
loading complex
 Peptide
loading
complex
• Tapasin- bridging protein brings heterodimer
close to TAP
• Extracellular Ig-like domains of tapasin bind to
α2 and α3 domains of MHC class I molecule
• TAP and tapasin interact via membrane
spanning region
• Result is peptide-binding groove is positioned
to receive peptide from TAP
• Erp57 stabilizes peptide loading complex
physically by interacting with calreticulin and
enzymatically protects disulfide bond of α2
domain from being broken by chemical
reduction
 Peptide bound MHC class I molecule

• Open conformation of peptide groove reduces overall

strength of peptide binding site
• Tight binding peptide = conformational change occurs
in the MHC class I molecule
1. Conformational change causes break in bond with
tapasin
2. MHC class I molecule leaves peptide loading complex
and exits ER in membrane enclosed vesicle
3. traffics through Golgi and is transported to cell surface
 Peptide
Editing
• Peptide editing- “trying on”
peptides
• Endoplasmic
reticulum aminopeptidase (ER
AP)- remove amino terminal
amino acids from peptides
that are too long, but
C terminus binds tightly
• Recycling of MHC class I
molecule that has lost
its peptide
MHC Class I- movie
MHC Class II
 • MHC class II molecules display
extracellular peptides
• Endosomes, phagosomes, lysosomes
peptides
Peptides • Leprosy and tuberculosis- use
presented vesicular system as protected site for
by MHC intracellular growth and replication-
No presentation of peptides by MHC
class II class I or MHC class II
molecules
 Invariant Chain

• Invariant chain- identical in all

individuals
• Prevent MHC class II
molecules from
binding peptides in the
ER
• Stabilizes
conformation of MHC
class II molecule until it
binds peptide
• Deliver MHC class II
molecules to
specialized endocytic
vesicles where they
bind peptide
 MHC class II
compartment (MIIC)

Cathepsin S and other

proteases cleave invariant
chain into CLIP-class II
invariant chain peptide
When a peptide binds tightly
there is a conformational
change in MHC class II
molecule and a release of
HLA-DM
MHC class II molecule
with bound peptide leaves
MIIC in vesicle that transports
it to plasma membrane
HLA-DO bound to HLA-
DM prevents HLA-DM from
releasing CLIP
IFN-γ increases expression of
HLA-DM but not HLM-DO
Important during infection
MHC class II- movie
 Antigen cross
presentation
• Cross-presentation- antigens on class II
pathway of antigen presentation can be
transferred to class I pathway of antigen
presentation
• Virally infected hepatocytes in liver are
phagocytized and the MHC class I
molecules and peptides are transferred
to the membranes of intracellular
vesicles of the Kupffer cell and then
transported to the cell surface
• Viral components from endocytic
vesicles go into cytoplasm for
degradation in proteasome and then
follow usual MHC class I pathway
 MHC expression

• Most cells express MHC

class I molecules
• Erythrocytes lack MHC
class I molecules
• MHC class II molecules
expressed only on B cells,
macrophages and dendritic
cells (professional antigen
presenting cells)
T cell receptor
recognizes
both peptide
and MHC
molecule

• CDR3 loops of T
cell receptor α and
β chains grasp side
chains of one of
the aa in middle of
peptide
• CDR1 and CDR2
loops contact α
helices of MHC
molecule
 • Human MHC = human leukocyte antigen complex
(HLA)
• Stable genes, no rearrangement
• Inherited diversity
• Gene families- multiple similar genes for
MHC I α chain, MHC II α chain, MHC II β
Diversity chain
• Genetic polymorphism- multiple alternative
of MHC forms of a gene in a population
• Highly polymorphic- numerous alleles
molecules • Monomorphic- genes with no
polymorphism
in humans • Oligomorphic- few alleles
• Importance of high genetic polymorphism =
most children inherit a different allele from each
parent – heterozygous vs. homozygous
• MHC class I isotypes (6)

• A, B, C- polymorphic

• Fx- present Ag to CD8

T cells

• Ligands for receptors

on NK

• E, G- Oligomorphic

• Ligands for NK cell

receptors

• F

• Chaparone

• MHC class II isotypes (5)

• DP, DQ, DR- polymorphic

• Present peptide Ag
directly to CD4 T cells

• DM, DO- oligomorphic

• Supervise peptide
loading of DP, DQ, DR
• Alleles of highly polymorphic MHC
genes have 1-50 aa substitutions
• Substitutions are located in the
domains that bind peptide and
interact with T cell receptor
• Anchor residues- aa that anchor
peptide to MHC molecule, little
variation within aa
• Peptide binding motif- combination
of anchor residues that bind
particular MHC isoform
MHC class I molecule = 2 and 9 typical anchor residues
MHC class II molecule = anchor residue less defined because of heterogeneity in length

The number of peptide binding motifs is limited so MHC allotypes that differ by only
a few AA often bind overlapping population of peptides

The greater the difference in sequence between 2 MHC allotypes the greater the
difference in peptide sequences they bind.
 MHC Restriction

•T cell that responds to a

peptide presented by one MHC
allotype will not respond to
another peptide bound by the
same MHC allotype or to the
same peptide bound to another
MHC allotype
 Heterozygote
advantage

• Haplotype- combination of HLA

alleles found on a given
chromosome 6
• Balancing selection-
evolutionary selection that acts to
maintain a variety of phenotypes in
a population
• Gray circle= total number antigenic
peptides from a pathogen that can
be presented by human MHC class
I and II molecules
• Yellow circle= subpopulation of
peptides that can be presented by
the MHC class I and II molecules
encoded by the genes of particular
MHC haplotypes
Exposure to pathogens selects for MHC
polymorphism

Directional Selection
Formation of new variants
of HLA class I and II alleles
• Point mutations
• Recombination of alleles of the same gene
• Recombination of alleles of different genes in the
same family
 MHC
heterozygosity
and
progression of
HIV-1 to AIDS

• Certain families of HLA

alleles associated with
slow progression of HIV
to AIDS, other families of
HLA alleles associated
with rapid progression
• Almost all correlations are
with HLA I
• Makes sense because
immune response is
killing virus infected cells Red- heterozygous for highly polymorphic class I and class II loci
by cytotoxic CD8 T cells
Yellow- homozygous for one locus
Blue- homozygous for two or three allelles
 • Autologous- self-MHC isoforms
• Allogeneic- all other MHC isoforms
• AlloreactiveT cells- T cells that respond to
complexes of peptide and allogeneic MHC
class I and II molecules
• Alloreaction- powerful T cell response
Terms against donor organ due to allogeneic
antigen
• HLA type- combination of HLA alleles that a
person has
• Alloantibody- any antibody raised in one
member of a species against
an allotypic protein from another member
of the same species