Transcript Janeway`s Immunobiology - University of Utah School of Medicine

Chapter 4 and 5 Ig study questions (Th):
• How does the immune system recognize a
diverse universe of possible antigens?
• How do antibodies simultaneously recognize a
huge variety of antigens and carry out a limited
number of effector functions?
• What is V(D)J recombination? What do the letters
(and parentheses) signify?
• What is the 12/23 rule?
• What are RAG-1/2 and TdT? What do they
recognize/do?
• What molecular changes are required to switch
from membrane-bound to secreted Ig?
Ig Structure and Function
Lecture – Sept. 8 & 13, 2016
Dean Tantin, PhD, Department of Pathology
Division of Microbiology & Immunology
University of Utah School of Medicine
JMRB 5200K, 7-3035
[email protected]
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Abcam
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• Epitope: the portion of the target antigen (Ag) bound
• Never before seen epitopes can be recognized
• Too many epitopes to allow direct encoding in the genome
VH
CH1
VL
CL
Fab = Fragment
antigen binding
Fc = Fragment
crystallizable
CH2
CH3
Why name it
F(ab’)2?
Figure 4.4
• Ig domain is a
recurring protein
structure in
immunology
• Immunoglobulin
superfamily
• This is how the
textbook will
denote:
heavy
light
Figure 4.3
• A quick preview of the T cell
receptor (TCR).
• Structurally related to
immunoglobulin.
• Does not recognize free
antigen. Instead recognizes
peptide antigens in the
context of presentation by the
major histocompatibility
complex (MHC) molecules.
Figure 4.13
One difference between Ig
and TCR: Ig recognizes Ag in
native form whereas TCR
recognizes Ag as peptide
presented by MHC molecules.
These three
regions vary.
“They are know as
hypervariable
regions” or
“complementaritydetermining
regions”.
Figure 4.3
Figure 4.6
AA
AA
• Hypervariable regions=CDRs,
complementary determining regions
Figure 4.7
Another difference between Ig
and TCR: TCR solely functions
as an antigen-specific receptor,
whereas Ig encodes both
antigen receptor and the major
effector molecule. Effector
functions are mostly mediated
by the IgH constant regions.
Antigen (Ag)
recognition
here
Business end of the
molecule is here,
encoded by heavy
chain (IgH) C regions.
IgH C regions specify
the isotype: IgM, IgD,
IgG, IgE, or IgA
Review. So far…
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IgG is example
4 polypeptide chains
Modular construction
Ig superfamily
Flexibility in hinge region
Heavy chain constant
region defines the isotype.
Figure 4.1
Your first look at the immunoglobulin genes
• This is just heavy chain. There
is also k and l light chain.
• Modular construction, just like
Ig proteins!
• Here mainly concerned with the
5’ end of the locus, and at the
end of the lecture with the 3’
end.
Related to Fig.5.5, but more complex
VH
C H1
C H2
C H3
C H4
Your first look at the immunoglobulin genes
•
•
•
V=variable
(D)=diversity
(heavy chain only)
J=joining
• This is just heavy chain. There
is also k and l light chain.
• Modular construction, just like
Ig proteins!
• Today will mainly be concerned
with the 5’ end of the locus, and
Tuesday with the 3’ end.
Related to Fig.5.5, but more complex
VH
C H1
C H2
C H3
C H4
• This is how your Ig loci look
while you are a developing
embryo, and in every cell of
your body except B cells
• “Gene segments”
• In B cells, either k or l locus
is used, not both
• Heavy chain defines
isotype: Cm=IgM
Figure 5.5
• No need to memorize this chart.
Know that there are many V
segments, several J segments; D
segments restricted to IgH/TCRb
• Not to scale
• Figure above only shows proteincoding regions. Regulatory
elements in DNA provide control
of e.g. recombination
Figure 5.5, 5.4
Figure 4.1
Figure 5.3
• Basically Ig recombination only in the B lineage
Figure 4.6, 4.7
• CDR 1 and 2 encoded in V
segment
• CDR3 made up of the junction
between gene segments
• Questions?
Figure 5.5, 4.7
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• Recombination signal sequences (RSSs, asterisks) mark
potential sites of V(D)J recombination.
• Recognized by the RAG-1 and RAG-2 proteins.
Figure 5.5
• Recombination signal sequences (RSSs) are recognized
by RAG (recombination activating gene) proteins-1 and -2
5’
5’
3’
3’
• 12mers/23mers do not recombine with themselves, only with RSSs with
the other spacer (the “12/23 rule”)
• ∴ V segments never recombine with other V segments
Figure 5.6
• You are looking at an Ig light chain, either k or l…
-or-
Figure 5.7
✔ -or- ✔
and ✔
or…
• “Direct repeat” RSSs leads to joining and deletion of the intervening DNA
• “Inverted repeat” RSSs leads to joining and inversion of the intervening
sequence. You will have to study the topology of this figure to see it.
Figure 5.7
• This is the
basis of the
12/23 rule
Figure 5.8
• Variability in the cut
generates “P-nucleotide
varariability”
• TdT=terminal deoxyribonucleotidyl transferase
• TdT is lymphocyte specific
• TdT introduces additional
nucleotides at the coding
junctions, and increasing
diversity “N-nucleotide
variability”
• Lack of RAG proteins, or
DNA-PK, or ArtemisSCID
(C=combined, meaning B
and T cells affected)
Figure 5.8
No need to
memorize this
chart.
combinatorial
diversity
junctional
diversity
Figure 5.15
+ somatic hypermutation
In B CELLS ONLY
(next lecture)
A difference between Ig and
TCR: TCR solely functions as
an antigen-specific receptor,
whereas Ig encodes both
antigen receptor and the major
effector molecule. Effector
functions are mediated by the
IgH constant regions.
Consequently, the
molecular biology of the
BCR/antibody
(immunoglobulin) gene
loci following activation
is much more complex
i.e., gene expression from the
Ig loci is increased many fold.
In plasma cells (effector B
cells), Ig accounts for 50% of
all gene expression. This
does not happen with T cell
receptor.
Conversion to Ab secretion changes cellular moprhology
Resting B
cell:
Antibody secreting
(plasma) cell:
Atlas of Blood Cells
i.e., there are specific
molecular mechanisms that
change immunoglobulin from
membrane-bound to secreted
forms. This does not happen
with T cell receptor.
B cells secrete antibody upon stimulation
cell surface
• B cells become activated by signaling through B cell Ag
receptor (BCR) complex, plus costimulatory signals.
• Same thing at other heavy chain regions.
• SC: secretion coding; MC: membrane coding
Figure 5.22
B cells secrete antibody upon stimulation
cell surface
• B cells become activated by signaling through B cell Ag
receptor (BCR) complex, plus costimulatory signals.
• Same thing at other heavy chain regions.
• SC: secretion coding; MC: membrane coding
Figure 5.22
i.e., there are further
recombination events to
generate antibodies with
specific effector functions (Ig
isotype switching), and
site-specific mutagenesis that
allows the selection of highaffinity antibodies (affinity
maturation)
Read the Chapters over the weekend!
I will be relying on figures from your textbook
less in the next lecture. Reading the chapters
therefore becomes more of an additional
resource.
Chapter 4 and 5 Ig study questions (Th):
• How does the immune system recognize a
diverse universe of possible antigens?
• How do antibodies simultaneously recognize a
huge variety of antigens and carry out a limited
number of effector functions?
• What is V(D)J recombination? What do the letters
(and parentheses) signify?
• What is the 12/23 rule?
• What are RAG-1/2 and TdT? What do they
recognize/do?
• What molecular changes are required to switch
from membrane-bound to secreted Ig?