Transcript B Cell - Biotechnology

Lecture 15
B Cell Activation and
Antibody Production
Overview of B Cell
Development,
Activation,
Antibody Production
B Cell Antigens
Antigens
Thymus Dependent Antigens
Thymus Independent Antigens
Dependent Upon Helper T Cells
to Induce Antibody Production
Does Not Need Helper T Cells
to Induce Antibody Production
Proteins
Polysaccharides, Lipids
B Cell Responses to ThymusDependent Antigens (T CellDependent Antibody Responses)
Primary and Secondary Antibody Responses
Phases of the Humoral Immune Response
A T-Dependent Antigen Must Contain Both
B and T Cell Epitopes
Antigen
T Cell Epitope B Cell Epitope
LINKED RECOGNITION
Follicles
(B Cells)
(T Cells)
Activation of B Cells by Antigen and Complement
1. Biochemical Signals
2. Endocytosis of Antigen
Antigen Recognition Phase of T-Dependent
Antibody Response
Interactions of B Cells with Helper T Cells
TEM Picture
B Cell
B Cell
T Cell
Initial Contact
T-B Conjugate
Note the broad area of membrane contact between
B and T Cells.
Helper T Cell-Dependent Activation
Of B Lymphocytes
B-Cell Activation by
Thymus-Dependent Antigens
C’R
Cytokines
Linked Recognition
Activated B Cells
(Following Interaction with TH Cells
Extra-follicular Site
Antibody Secreting
Cells
Antibodies
Follicle
Germinal Center
Late Events in T Cell-Dependent
Antibody Responses-Germinal
Center Reaction
• Affinity Maturation
– Somatic Hypermutation
• Generation of Memory B Cells
Somatic Hypermutation and Affinity
Maturation of Antibodies
Affinity maturation is the process that
leads to increased affinity of antibodies
for a particular antigen as a result of
somatic mutation in the Ig genes followed
by selective survival of B cells producing
the antibodies with the highest affinity
Affinity Maturation in Antibody Responses
Selection of High Affinity
B Cells in Germinal Center
Phases of the Humoral Immune Response to
T-Dependent Antigen
Anatomy of Humoral Immune Responses
Antibody Isotype
Switching
Isotype Switching Under the Influence of Helper
T Cell-Derived Cytokines
Mechanism of Ig Isotype Switching
CD4 T Cell-Dependent Effects in
Antibody Responses
• Memory B Cell Development
• Isotype Switching
• Affinity Maturation
ThymusIndependent
Antigens
B-Cell Activation by
Thymus-Independent and Dependent Antigens
Most TI antigens are polyvalent and induce maximal
Crosslinking of membrane Ig on B cells, without a
Need for T cell help.
Features of Antibody Responses to
T-Dependent and T-Independent Antigens
Antibody Response to T-Dependent
Antigens
• Role of Helper T Cells
– Cytokines
– CD40/CD40L interactions
• Isotype Switching
– Switch Recombination
– Cytokines and Isotypes
• Affinity Maturation
– Somatic hypermutation
– Selection for B cells which produce High Affinity
Antibodies
• Memory B Cells
Antibody Effector
Functions
Effector Functions of Antibodies
Neutralization of Microbes by Antibodies
Neutralization of Toxins by Antibodies
Opsonization of Microbes by Antibodies
Antibody-Dependent Cellular Cytotoxicity (ADCC)
Functions of Complement
Complement-Mediated Lysis of E. coli
Alive
Killed
Cellular Interactions in Immune Responses
The Immune
Response:
A Summary
WHY can immune system recognize so
many different epitopes??
Antibody heavy and light chains are
composed of gene segments
Variable regions are unique
A limited variety of constant region sequences
are used
They must be rearranged into functional genes
before they can be transcribed
p. 106
Organization of Ig genes
Germline DNA- gene segments surrounded by
noncoding regions
These are rearranged to form functional genes
Light chains- V, J and C segments
Heavy chain- V, D, J, C
V regions rearrange first
A single V can rearrange to more than one C
Multigene families
 or 
In humans: 40 V, 5 J, 1 C
Similar number of  genes in humans;
this is rare in mice
Heavy-chain gene families are similar but more
complex (D segment)
CH regions formed from exons
p. 111
One of many possible combinations
Heavy chain DNA
D-J and V-DJ rearrangements must occur
separately
On a mature B cell, both mIgM and mIgD
are expressed on the cell surface
How does rearrangement occur?
Each V, D and J is flanked by RSS
(Recombination signal sequences)
Mechanism is controlled by RAG-1 and RAG-2
proteins and an enzyme TdT
If any of these proteins is defective no mature
B cells can form; nor T cells
p. 112
“Junctional flexibility” contributes to diversity
p. 115
But not all rearrangements are “productive”
B cells are diploid and contain chromosomes
from both parents
However, heavy chain genes are rearranged
from only one chromosome, as are light chain
genes.
Therefore, any one B cell will contain
one VH and one VL (antigen specificity)
How? Allelic exclusion (Yancopoulos and Alt, 1986)
Model for allelic exclusion
p. 116
Generation of antibody diversity
(why are there so many possible antigen
combining sites?)
Multiple germline gene segments
In human germline:
51 VH, 27 D, 6 JH
40 V, 5 J 
30 V , 4 J
Combinatorial V-J and V-D-J joining
57 V X 27 D X 6 J= 8262 possible combinations
for VDJ joining
40 V X 5J = 200 possible V
120 possible V
8262 X (200+120) = 2.64 X 106 possible
combinations
Without taking into account other sources of
diversity
Junctional flexibility in V-J or V-D-J junction
Additional nucleotides added at junctions
(P or N addition), if a single-stranded
region is created during the joining
process
Somatic hypermutation
mutations occur AFTER rearrangement
tends to occur in CDR regions
affects antigen affinity (tends to increase):
“affinity maturation”
occurs in B but not T cells
Class switching
After antigen stimulation heavy-chain DNA can
rearrange so VDJ joins to any isotype
Cytokines help determine the isotype
IgG2a or IgG3 (mice): IFN-
IgM: IL-2, IL-4, IL-5
IgE: IL-4
p. 122
Membrane-bound or secreted?
Alternative splicing, p. 124
Mature B cells express both mIgM and mIgD
No switch site between C and C
The VDJCC contains 4 polyadenylation sites
mIgM or mIgD can be generated depending
on which polyadenylation site is used
Regulatory elements of transcription
Promoters
Enhancers
Gene silencers
Gene rearrangement brings enhancers close
to the promoter they influence
Why aren’t Igs produced in B cells?
In T cells a protein may bind to the -enhancer
and prevent V-J joining
Arrangement of immunoglobulin genes (and
formation from exons) and greatly
facilitated formation of genetically
engineered antibodies