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Transcript of Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J...
![Page 1: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/1.jpg)
Chapter 4 and 5 Ig study questions (Tu):• Can you name at least four ways in which CSR
and V(D)J recombination differ?• What are the substrates (what genes, what
areas) for CSR and SHM? What are the key factors?
• How does AID work? Where does it operate?• How are specific class switch events regulated?• Why are antibodies used so widely, as lab
reagents, diagnostics, therapeutics?
![Page 2: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/2.jpg)
![Page 3: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/3.jpg)
The brick?
![Page 4: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/4.jpg)
The brick? AID
![Page 5: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/5.jpg)
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.
![Page 6: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/6.jpg)
Consequently, the molecular biology of the
BCR/antibody (immunoglobulin) gene loci following activation is much more complex
![Page 7: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/7.jpg)
i.e., there are further recombination events to generate antibodies with specific effector functions (Ig isotype switching), andsite-specific mutagenesis that allows the selection of high-affinity antibodies (affinity maturation)
![Page 8: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/8.jpg)
• Immunological phenomena:-Isotype switching-Affinity maturation
Happen after stimulation of mature B cells, in the secondary lymphoid organs (LN, spleen, Peyer’s patches).
Mostly occur in germinal centers (“the germinal center reaction”)
![Page 9: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/9.jpg)
Figure 1.18
![Page 10: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/10.jpg)
• Immunological phenomenon:-Isotype switching
Molecular mechanism:-Class switch recombination
• Immunological phenomenon:-Affinity maturation
Molecular mechanism:-Somatic hypermutation
![Page 11: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/11.jpg)
Figure 5.20
• V segment generated by V(D)J recombination is fixed,but can become mutated by SHM (heavy and light)
• Class switch recombination changes out heavy chain constant regions, replacing IgM/IgD with a “switched” isotype
e.g.,IgE
![Page 12: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/12.jpg)
-Isotype switching•same Ag binding site
different heavy chain constant regions and effector properties
•Ig comes from the same (heavy and light chain) gene
loci
![Page 13: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/13.jpg)
Figure 5.16
• We return to effector functions of antibodies in Ch.10• How do you switch from IgM/IgD to other isotypes?
Heavy chain constant region defines the isotype
![Page 14: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/14.jpg)
Figure 5.3 + 5.16 + 5.18
• Not to scale and simplified• Figure above only shows protein-
coding regions. Regulatory elements in DNA provide control of recombination and transcription
![Page 15: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/15.jpg)
Figure 5.3 + 5.16 + 5.18 + 5.25
L
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When B cells leave the bone marrow, they express IgM and IgD on their surface.
During an immune response, B cells may undergo “class switching” - to give rise to B cells producing IgG, IgA or IgE.
RAG-1 & -2 NOT required
Ig Class Switch Recombination (CSR):
L
![Page 17: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/17.jpg)
During class switching, recombination occurs between switch sites.
The initial recombination will occur between the m switch site and a downstream switch site.
Since there is no switch site associated with the d constant region gene segment, the B cell cannot class switch to IgD.
Ig Class Switch Recombination (CSR):
![Page 18: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/18.jpg)
Extra-chromosomal DNAis lost/degraded
Class switching does not alter the VDJ sequence and consequently has no effect on antibody specificity.
Ig Class Switch Recombination (CSR):
![Page 19: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/19.jpg)
Ig Class Switch Recombination (CSR) is instructed
Unlike V(D)J, which is largely random
CSR is initiated by CD40L:CD40 + appropriate cytokine
IgG1 IgG2a IgG2b IgG3 IgA IgE
IL-4 + +IL-5 +IFN + +TGF + +
![Page 20: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/20.jpg)
Extra-chromosomal DNAis lost/degraded
IL-4 receptor
“sterile” transcription
Ig Class Switch Recombination (CSR):
![Page 21: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/21.jpg)
• CSR requires:– Activation-induced deoxycytidine deaminase
(AID)– Uracil DNA glycosylase (UNG)
Ig Class Switch Recombination (CSR):
![Page 22: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/22.jpg)
Properties of AID
AID
• Expressed mainly in GC B cells• Acts on single-stranded DNA
![Page 23: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/23.jpg)
• AID converts C U at switch sites• U removed by UNG, “abasic site”• Cut by endonuclease• Joining to another switch site
Ig Class Switch Recombination (CSR):
![Page 24: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/24.jpg)
Figure 5.25
![Page 25: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/25.jpg)
“sterile” transcription
Ig Class Switch Recombination (CSR):
![Page 26: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/26.jpg)
germline transcript=sterile transcript
Ig Class Switch Recombination (CSR):
![Page 27: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/27.jpg)
Evidence-
• AID-/- mice & humans • UNG knockout mice
No CSR
Ig Class Switch Recombination (CSR):
![Page 28: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/28.jpg)
Occurs during antibody response - NOT B cell maturation.
During proliferation of B cells in germinal centers, there are increased mutation rates in the sequence encoding the V regions of the BCR (Ab).
This will randomly alter (increase or decrease) the affinity of the B cell’s BCR (Ab) for the eliciting antigen.
Those B cells with increased affinity for the antigen are selected.
As a result, as an immune response proceeds, the affinity of antibody produced increases “Affinity maturation”
Somatic Hypermutation (SHM):
![Page 29: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/29.jpg)
Somatic Hypermutation (SHM):
Figure 5.24
![Page 30: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/30.jpg)
Monoclonal antibodies as drugs…
![Page 31: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/31.jpg)
Evidence-
• AID-/- mice & humans • UNG knockout mice
• Somatic hypermutation uses a similar mechanism
Somatic Hypermutation (SHM):The Germinal Center Reaction:
No CSRNO SHM
![Page 32: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/32.jpg)
From Martin & Scharff (2002)
• SHM targets V, not C region; = 1/1000 bp/division
• Transcription means RNA polymerase, not DNA polymerase (typo on page 182 of Janeway textbook.)
Im is the promoter forsterile (noncoding) transcription of Cm switch region
VDJ
Somatic hypermutation (SHM):
![Page 33: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/33.jpg)
Properties of AID• Expressed only in GC B-cells• Acts on single-stranded DNA
![Page 34: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/34.jpg)
Properties of AID• Expressed only in GC B-cells• Acts on single-stranded DNA
DNAreplication
![Page 35: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/35.jpg)
•AID converts C U at Ig V exon•U can be fixed as T by replication, or•U removed by UNG, “abasic site”•Cut by endonuclease•Error-prone repair
Somatic hypermutation (SHM):
![Page 36: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/36.jpg)
Cryosection of lymph node obtained 5 days after secondary antigen injection. Green: ki-67--dividing cells (spatially defines the germinal center). Red: IgM--primary B cell follicle. Yellow: Macrophages.
![Page 37: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/37.jpg)
Somatic Hypermutation (SHM):The Germinal Center Reaction:
Evidence for a role of AID in SH and CSR:•AID-/- mice & humans: no SH or CSR•Fibroblasts transfected with AID gene &
substrate SHM & CSROther B cell-specific functions not
required (necessary and sufficient)
![Page 38: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/38.jpg)
Figure 5.26
H L
H – heavy chain locusL – k and l light chain loci
H L
H L
H
H L
H
H
RAG-1/2
AID
AID
![Page 39: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/39.jpg)
The brick?
![Page 40: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/40.jpg)
The brick? AID
![Page 41: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/41.jpg)
The brick? AIDThe window?
![Page 42: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/42.jpg)
The brick? AIDThe window? Ig loci (light chain V region, heavy V and S)
![Page 43: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/43.jpg)
The brick? AIDThe window? Ig loci (light chain V region, heavy V and S)Al?
![Page 44: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/44.jpg)
The brick? AIDThe window? Ig loci (light chain V region, heavy V and S)Al? Natural Selection (APOBECs)
![Page 45: Chapter 4 and 5 Ig study questions (Tu): Can you name at least four ways in which CSR and V(D)J recombination differ? What are the substrates (what genes,](https://reader036.fdocuments.net/reader036/viewer/2022062309/5697bfaf1a28abf838c9d30d/html5/thumbnails/45.jpg)
Chapter 4 and 5 Ig study questions (Tu):• Can you name at least four ways in which CSR
and V(D)J recombination differ?• What are the substrates (what genes, what
areas) for CSR and SHM? What are the key factors?
• How does AID work? Where does it operate?• How are specific class switch events regulated?• Why are antibodies used so widely, as lab
reagents, diagnostics, therapeutics?