B cell activation & organization of Immunoglobulin genes Flashcards by Hugo Siu

Overview of B cell development

List the following from most immature to most mature

( Pro - B cell, Mature B cell, Activated B cell, Lymphoid cell, Immature B cell, Pre - B cell, Hematopoietic stem cell )

Hematopoietic stem cell > Lymphoid cell > Pro - B cell > Pre - B cell > Immature B cell > Mature B cell > Activated B cell

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Organization and Expression of Immunoglobulin genes

What does V, D, J, C stands for?

V: Variable
D: Diversity
J: Joining
C: Constant

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Organization and Expression of Immunoglobulin genes

What is the structural difference between light - chain & heavy - chain?

Light chain : V - J arrangement

Heavy chain : V - D - J arrangement

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Organization and Expression of Immunoglobulin genes

What are the two light chain gene?

Kappa & Lambda

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Organization and Expression of Immunoglobulin genes

What is the order in rearranged Kappa - chain DNA?

( Intron x2 , Joined VJ gene segment, constant region, Leader exon )

Order :
Leader > Intron > Joined VJ gene segment > Intron > Constant region

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Organization and Expression of Immunoglobulin genes

Which sequence in light chain is cleaved in RER?

Leader sequance

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Organization and Expression of Immunoglobulin genes

Which of the following determines the class of antibodies?

(V, D, J, C)

C : Constant

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Organization and Expression of Immunoglobulin genes

What are the difference in rearranged Kappa - chain DNA & rearranged Heavy - chain DNA?

Kappa :
-> Joined VJ gene segment
-> A constant region

Heavy :
-> Joined VDJ gene segment
-> A series of C gene segments
( For antibody class )

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Organization and Expression of Immunoglobulin genes

What is the two class of antibodies that is expressed initially in primary RNA transcription?

C(mu) : IgM
C(delta) : IgD

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Organization and Expression of Immunoglobulin genes

What is the process that determines whether IgM or IgD is produced?

RNA splicing

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Organization and Expression of Immunoglobulin genes

Does a mature B cell that expresses both IgM & IgD have the same identical antigenic specificity, and why?

Yes

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What are the 2 mechanisms of variable - region DNA rearrangements?

RSS ( Recombination signal sequences )
-> direct recombination

VDJ recombinases
-> joining gene segment

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Organization and Expression of Immunoglobulin genes

What is the 2 types of RSSs? ( Recombination signal sequences )

One turn RSS
Two turn RSS

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Organization and Expression of Immunoglobulin genes

What are the 2 -amers in RSS?
And whats the properties of it?

Heptamer
-> Palindromic

Nonamer
-> AT - rich

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Organization and Expression of Immunoglobulin genes

What is the “ One - turn / two - turn joining rule “ about ?

One - turn RSS can only join with two - turn RSS

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Organization and Expression of Immunoglobulin genes

What is the recombinase enzyme that can recognize RSSs?

RAG - 1 / 2
( Recombination - activating gene )

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Organization and Expression of Immunoglobulin genes

What is the enzyme that cleave the hairpin structure and trim a few nucleotides from the coding sequence?

Endonuclease

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Organization and Expression of Immunoglobulin genes

What is the enzyme that adds nucleotides to the cut ends of coding sequence of heavy chain?

TdT
( Terminal deoxynucleotidyl transferase )

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Organization and Expression of Immunoglobulin genes

What is the enzyme that repair and ligate the coding sequences?

DSBR enzymes
( Double - strand break repair )

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Does the RAG enzymes cleave both strand of the DNA at the junctures of the RSS and the coding sequence?

Nope, they cleave one strand of DNA

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Organization and Expression of Immunoglobulin genes

What is P - addition and which 2 enzymes are involved?

Cleavage of hairpin by endonuclease
-> Sites for addition of P - nucleotides
- -> DSBR enzymes add complementary nucleotides

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Organization and Expression of Immunoglobulin genes

What is N - addition and which 3 enzymes are involved?

Cleavage of hairpin by endonuclease
-> Sites for addition of P - nucleotides
- -> DSBR enzymes add complementary nucleotides
- - -> TdT adds N - nucleotides

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Organization and Expression of Immunoglobulin genes

Does N - addition happens in both light chain and heavy chain?

Nope
It only happens in heavy chain

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Organization and Expression of Immunoglobulin genes

What is junctional flexibility?

The selection of which nucleotide in two coding sequences (e.g. V & J / D & J) to be joined together can be completely random.

E.g. V ( ATG ) & J ( CGC )
-> ATGCGC
-> ATGGC
-> ACGC
-> ATCGC
etc

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Organization and Expression of Immunoglobulin genes What is the disadvantage of junctional flexibility?

It can produce stop codons which interrupt translation

Organization and Expression of Immunoglobulin genes What are the 5 things that creates antibody diversity?

V(D)J rearrangement P - region nucleotide addition N - region nucleotide addition Junctional flexibility Somatic hypermutation

Organization and Expression of Immunoglobulin genes Which of the following occurs in bone marrow / germinal center? V(D)J rearrangement P - region nucleotide addition N - region nucleotide addition Junctional flexibility Somatic hypermutation

Bone marrow : V(D)J rearrangement P - region nucleotide addition N - region nucleotide addition Junctional flexibility Germinal center : Somatic hypermutation

Organization and Expression of Immunoglobulin genes What is the mutation rate in somatic hypermutation? 10 ^ (??) / bp / mutation

10 ^ (-3) / bp / mutation

Organization and Expression of Immunoglobulin genes Somatic hypermutation guarantees at least one mutation per every (??) division

Two

Organization and Expression of Immunoglobulin genes What is allelic exclusion?

Allelic exclusion ensures single antigenic specificity

Organization and Expression of Immunoglobulin genes In allelic exclusion, what is first produced in progenitor B cell?

D and J component of VDJ heavy chain

Organization and Expression of Immunoglobulin genes In allelic exclusion, which is the first allele checked?

Mu heavy chain (Mu allele #1)

Organization and Expression of Immunoglobulin genes In allelic exclusion, what does Mu heavy chain (Mu allele #1) inhibits & induces?

Inhibits rearrangement of the Mu allele #2 Induces Kappa rearrangement

Organization and Expression of Immunoglobulin genes In allelic exclusion, what is the second allele checked after Mu heavy chain (Mu allele #1) is productive?

Kappa light chain (Kappa allele #1)

Organization and Expression of Immunoglobulin genes In allelic exclusion, what happens when both Mu heavy chain (Mu allele #1) & Kappa light chain (Kappa allele #1) are productive alleles?

Antibodies are successfully produced

Organization and Expression of Immunoglobulin genes In allelic exclusion, what does Mu heavy chain (Mu allele #1) & Kappa light chain (Kappa allele #1) inhibits & induces?

Inhibits -> rearrangement of the Kappa allele #1 -> Lambda rearrangement Induces nothing

Organization and Expression of Immunoglobulin genes In allelic exclusion, what happens if Kappa light chain (Kappa allele #1) is nonproductive?

Kappa light chain (Kappa allele #2) is checked

Organization and Expression of Immunoglobulin genes In allelic exclusion, what if both Kappa light chain (Kappa allele #1 & #2) are nonproductive?

Lambda light chain ( Lambda allele #1) is checked

Organization and Expression of Immunoglobulin genes In allelic exclusion, what does Mu heavy chain (Mu allele #1) & Lambda light chain (Lambda allele #1) inhibits & induces?

Inhibits rearrangement of Lambda allele #2

Organization and Expression of Immunoglobulin genes In allelic exclusion, what happens if both Mu allele are nonproductive?

Apoptosis

Organization and Expression of Immunoglobulin genes In allelic exclusion, what happens if both Kappa allele & Lambda allele are nonproductive?

Apoptosis

Organization and Expression of Immunoglobulin genes Which two class of membrane bound antibodies are simultaneously expressed in mature naïve B cell?

mIgM & mIgD

Organization and Expression of Immunoglobulin genes What are the difference between sIgM & mIgM?

sIgM -> Contains hydrophilic portion that are encoded by S exon of C(mu) mIgM -> Contains both hydrophilic & hydrophobic portion that are encoded by M1 & M2 exons of C(mu) - -> Hydrophilic : Outside + Cytoplasm - -> Hydrophobic : Membrane

Organization and Expression of Immunoglobulin genes What processing action in RNA transcription causes the difference between sIgM & mIgM?

RNA splicing

Organization and Expression of Immunoglobulin genes Which of the two organelles is essential for sIg production?

RER & Golgi

Organization and Expression of Immunoglobulin genes What are the 4 things that happen in RER for sIg production?

Translation of heavy chain & light chain Cleavage of Leader sequence Assembly of heavy chain & light chain to form monomer Assembly of monomers to form homodimer

Organization and Expression of Immunoglobulin genes What are the 2 things that happen in Golgi for sIg production?

Post translational modification of homodimer -> Addition of oligosaccharide Packaged by secretory vesicles

Organization and Expression of Immunoglobulin genes What is the difference in Secretion between sIg & mIg

Same thing happens in RER & Golgi However -> Hydrophobic part of mIg becomes embedded on the membrane of secretory vesicle -> Fusion of secretory vesicle with cell membrane causes mIg to stay on the membrane instead of being released

Organization and Expression of Immunoglobulin genes What are the 3 types of regulatory elements in Ig genes?

Promoters Enhancers Silencers

Organization and Expression of Immunoglobulin genes Where can promoters be found and what does it do?

200 bp upstream from transcription initiation site Promotes initiation of RNA transcription in a specific direction

Organization and Expression of Immunoglobulin genes Where can enhancers be found and what does it do?

upstream / downstream of genes ( in this case VDJ ) It can stimulate promoter to initiate transcription regardless its position / direction

Organization and Expression of Immunoglobulin genes Where can silencer be found and what does it do?

upstream or downstream of genes ( in this case VDJ ) It can inhibit / repress gene transcription regardless its position / direction

Organization and Expression of Immunoglobulin genes Does each V ( heavy chain ) & V ( light chain ) segment has a promoter & leader sequence?

Yes

Organization and Expression of Immunoglobulin genes Which of the following DNA does not contain silencers? Heavy chain DNA Kappa chain DNA Lambda chain DNA

Lambda chain DNA

Organization and Expression of Immunoglobulin genes Explain the mechanism in class switching ( from IgM -> IgA ), Including the following words Switch region Constant exon region Switch recombinase

In C gene after VDJ genes There are switch region right before every different constant exon region (e.g. S(IgM) C(IgM) S (IgD) C(IgD) ) When B cell desires to switch IgM class to IgA Switch recombinase aids in forming a loop from S(IgM) -> C(IgE) ) ( According to order IgM/D/G/E/A ) Removal of the loop brings S(IgA) in proximity to VDJ genes, allowing transcription of VDJ C(IgA)

Organization and Expression of Immunoglobulin genes What is the switch factor (cytokine)?

IL - 4

B - cell Maturation, Activation & Differentiation How long can plasma cell survive?

1 - 2 weeks

B - cell Maturation, Activation & Differentiation What is the function of IL - 7?

It down regulates the adhesion molecules

B - cell Maturation, Activation & Differentiation What triggers the expression of IL - 7 receptor on Pre - B cells?

Binding of c-Kit -> SCF *c-Kit -> Bone marrow stromal cell SCF -> Pro - B cell

B - cell Maturation, Activation & Differentiation What is the protein that help transduce activating signals of mIg on B cell ?

Ig - alpha / Ig - beta heterodimers

B - cell Maturation, Activation & Differentiation What are the two signals that drive B cells into & through the cell cycle?

Ag - mIg CD40L

B - cell Maturation, Activation & Differentiation What does Ag - mIg induce?

Expression of MHC - 2 & B7 on B cell

B - cell Maturation, Activation & Differentiation What is the purpose of expressing MHC - 2 on B cell?

Recognization of MHC - 2 by Th cell activates Th cell

B - cell Maturation, Activation & Differentiation What is the purpose of expressing B7 on B cell?

Allow B7 - CD28 interactions which co - stimulate Th cell with MHC - 2 recognition

B - cell Maturation, Activation & Differentiation What happens when Th cell is activated by the co - simulation?

Th cell begins to express CD40L -> Interaction between CD40 & CD40L (2)

B - cell Maturation, Activation & Differentiation What happens after both signals is present & active?

B cells express cytokines receptor Th cell release cytokines -> IL - 2 -> IL - 4 -> IL - 5 Signals supports progression of B cell -> DNA synthesis -> Differentiation

B - cell Maturation, Activation & Differentiation Which of the following cytokine induce class switch to IgG2a / IgG3? IL - 2 IL - 4 IL - 5 IFN - gamma TGF - beta

IFN - gamma

B - cell Maturation, Activation & Differentiation Which of the following cytokine induce class switch to IgA / IgG2b? IL - 2 IL - 4 IL - 5 IFN - gamma TGF - beta

TGF - beta

B - cell Maturation, Activation & Differentiation Which of the following cytokine induce class switch to IgE / IgG1? IL - 2 IL - 4 IL - 5 IFN - gamma TGF - beta

IL - 4

B - cell Maturation, Activation & Differentiation Which of the following cytokine induce class switch to IgM? IL - 2 IL - 4 IL - 5 IFN - gamma TGF - beta

IL - 2 IL - 4 IL - 5

Match the following IFN - gamma TGF - beta Th1 IL - 4 IL - 2 Th2 IL - 5

Th1 -> IFN - gamma -> TGF - beta Th2 IL - 2 IL - 4 IL - 5

B cell activation & organization of Immunoglobulin genes Flashcards

(71 cards)