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Chapter9B

Lymphocyte

Xing-cheng WEI ( 韦星呈 )Room 323, Building of Basic Medicine

Department of Immunology, Tel.62215671(office)

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Functions of AbFunctions of Ab（ 1 ） Neutralization（ 2 ） Opsonization（ 3 ） Complement activation （ 4 ） Mediation of ADCC

Binding of C1q to Ag-Ab Complex

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Patient of Bruton’s hypogammaglobulinemia

(B cell deficiency)

A 22 month-old male with Bruton’s hypogammaglobulinemia accompanied by vaccinia necrosum. Note the large necrotic area at the vaccination site. Progressive vaccinia occurs because of an immune defect in B cells and Ig.

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Mechanisms of Bruton’s hypogammaglobulinemia

Bruton's tyrosine kinase (abbreviated Btk or BTK) also known as tyrosine-protein kinase BTK is an enzyme that is encoded by the BTK gene. BTK is a kinase playing a crucial role in B-cell development.

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Development Surface MoleculesSubpopulations Functions

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Sec.1Maturation of

B

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[Review][Review] One One HLA gene only gives one peptide chainHLA gene only gives one peptide chain

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[Review] Specificity of an Ab/BCR is determined by V

region of the Ag-receptor

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I.Genomic Organisation of Ig Genes

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Genomic Organisation of Ig H-chain and L-chain Genes

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II. Generation of Diversity

of Ig Genes II. Generation of Diversity

of Ig Genes

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1. V(D)J Recombination1. V(D)J Recombination

2. V(D)J Junction2. V(D)J Junction

3. Somatic Hypermutation3. Somatic Hypermutation

[Three Mechanisms][Three Mechanisms]

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1.V(D)J Recombination

(In Bone Marrow)

1.V(D)J Recombination

(In Bone Marrow)

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V(D)J RecombinationV(D)J Recombination

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(1) Ig H-chain Gene Rearrangement

DH1-25 JH 1-6 CVH 1-40

Recombination occurs at the level of DNA which can now be transcribed to form a H-chain

peptide

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(2) Ig L-chain Gene Rearrangement

Germline

Vk Jk Ck

Spliced mRNA

Rearranged1°transcript

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DV

RSS and the 12-23 Rule

D JV

23 12

97 79

V D

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23-mer

12-mer

Loop of intervening

DNA is excised

• Heptamers and nonamers

align back-to-back

• The shape generated by the

RSS’s acts as a target for

recombinases

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V1 V2 V3 V4

V8V7

V6V5

V9 D J

V1 D J

V2

V3

V4

V8

V7

V6

V5

V9

• An appropriate shape can not be formed if two 23-mer flanked elements

attempted to join (i.e. the 12-23 rule)

Molecular explanation of the 12-23 rule

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TCR

Organisation of TCR genesL & V

x70-80 C

TCR

D1 J1 x 6 C1 D2 J2 x 7 C2

TCR genes segmented into V, (D), J & C elements(VARIABLE, DIVERSITY, JOINING & CONSTANT)Closely resemble Ig genes (~IgL and ~IgH)

This example shows the mouse TcR locus

J x 61

L & Vx52

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TCR gene rearrangement

Spliced TcR mRNA

Germline TcR

Vn J CV2 V1

Rearranged TcR1° transcript

Rearrangement very similar to the IgL chains

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Rearranged TcR 1° transcript

Spliced TcR mRNA

L & Vx52 D1 J C1 D2 J C2

Germline TcR

TCR gene rearrangement

D-J Joining

V-DJ joining

C-VDJ joining

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[Concept][Concept]

(1) Antigen receptor genes consist with many V,D, and J gene segments next to RSS.

(1) Antigen receptor genes consist with many V,D, and J gene segments next to RSS.

(2) Recombinase (RAG1/2) recognize RSS and cooperate with other enzymes to mediate rearrangement of V(D)J segments.

(2) Recombinase (RAG1/2) recognize RSS and cooperate with other enzymes to mediate rearrangement of V(D)J segments.

V(D)J recombination V(D)J recombination

[Result][Result] Multiple V, D, and J gene segments may combine randomly, so as to generate a great number of different combinations of Ig V region.

Multiple V, D, and J gene segments may combine randomly, so as to generate a great number of different combinations of Ig V region.

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2. V(D)J Junction

(In Bone Marrow)

2. V(D)J Junction

(In Bone Marrow)

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V D JTCGCATATAGCGTATA

(1) Addition of P Nucleotides at Junction

TCG CATATAGCGT ATA

TCGCACATATAGCGTGTATA

*P: Palindromic (Added by Polymerase)

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V D JTCGCCGTTATATAGCGGCAATATA

X X X X

X X X X

X X X X

Germline-encoded nucleotides

Palindromic (P) nucleotides - not in the germline

Non-template (N) encoded nucleotides - not in the germline

(2) Addition of N Nucleotides at Junction

*N: Non-template encoded (Added by TdT).

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Junction Diversity[Concept] Junction Diversity[Concept]During V (D) J recombination, one to several nucleotides can be added to or removed from the DNA end generated by recombinase, which increase antigen receptor diversity greatly.

During V (D) J recombination, one to several nucleotides can be added to or removed from the DNA end generated by recombinase, which increase antigen receptor diversity greatly.

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3.Somatic Hypermutation

(In Germinal Center)

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Somatic Hypermutation[Concept]

In proliferating germinal center B cells that finished V(D)J recombination, the Ig V genes undergo point mutation at a very high rate, which plays a very important role in affinity maturation. FR1 FR2 FR3 FR4CDR2 CDR3CDR1

Amino acid No.

Variability80

100

60

40

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20 40 60 80 100 120

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Estimates of Diversity

V D J V(D)J Junction

Repertoire

So Hy

Ig(BCR)

H 40 25

6 6000

8.4×106 3×105 ～ 1011 1×106

40 - 5 200

30 - 4 120

TCR

70 - 61 42705.8×106 ＞ 3×105 ～ 1012 -

52 2 13 1352

12 - 5 602160 ? ? -

4 3 3 36

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Affecting lymphoid tissue. Predominating cells are B lymphocytes and lymphoblasts. Oral lesions include swollen and hyperplastic gingivae, ulceronecrotic lesions, and marked tendency to gingival hemorrhage.

B Lymphoid leukemia maybe involved in V(D)J

recombination

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III.

Maturation of B Cellsin Primary Lymphoid

Organs

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IV.

Induction of BCell Central Tolerance

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B-Cell Clone Deletion [Concept] In bone marrow ， immature B cells binding self-Ag with mIgM can cause die by apoptosis, so as to remove the self-reactive B cell clones. [Results] Set up B cell central tolerance to self-Ags, so that no any mature B cell clone can react to self-Ag.

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The Fate of Immature B Cell Depends The Fate of Immature B Cell Depends on Whether Binding with Self Antigens on Whether Binding with Self Antigens

or Notor Not

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Sec.2Surface Molecules

BCR Complex

Co-Receptor

Accessory Molecule

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[Component]

-BCR: (B Cell Receptor) (mIg)-Ig/ Ig(CD79a/CD79b)

I. BCR Complex

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BCR -4 peptide chains -4 peptide chains

= mIg.= mIg.-C end longer than -C end longer than an Ab H chain and pass an Ab H chain and pass through membrane.through membrane.-Cytoplasmic region is-Cytoplasmic region isshort, and can not short, and can not directly transfer directly transfer Ag-signal into cell.Ag-signal into cell.-transfer Ag-signal to -transfer Ag-signal to IgIg/Ig/Ig..

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Ig/IgHeterodimer(Heterodimer( & & ))..Extramembrane domaiExtramembrane domain: belong to IgSF(CAM)n: belong to IgSF(CAM)Transmembrane domaiTransmembrane domain: receives Ag-signal fron: receives Ag-signal from m BCR by [-][+] charge.BCR by [-][+] charge.Cytoplasmic domainCytoplasmic domain ：：Longer, with ITAMs, traLonger, with ITAMs, transfer Ag-signal into the nsfer Ag-signal into the B cell.B cell.

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[Functions of BCR][Functions of BCR] ① ①specifically recognize Ag.specifically recognize Ag.

② ②transfer the Ag-signals to Igtransfer the Ag-signals to Ig/Ig/Ig..

[Functions of [Functions of IgIg/Ig/Ig]] Transfer Ag-signals from BCR into Transfer Ag-signals from BCR into

the B cell.the B cell.

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Function of BCR Complex

IgIg

Intracytoplasmicsignalling domains

Extracellular antigenrecognition domains

The cytoplasmic domains of the Ig and Ig contain Immunoreceptor Tyrosine -based Activation Motifs (ITAMs) - 2 tyrosine residues separated by 9-12 amino acids - YXX[L/V]X6-9YXX[L/V]

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II.Co-receptor (B Cell)[Component]

-CD19

-CD21

-CD81

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Effects of B Cell Co-receptor

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[Functions of Co-receptor][Functions of Co-receptor] (1) enhance BCR-Ag binding.(1) enhance BCR-Ag binding. (2) help BCR transduce Ag signals.(2) help BCR transduce Ag signals.

[CD21[CD21(CR2)(CR2)]] Bind C3bBind C3b to link to Ag to link to Ag (other C3 splits: i(other C3 splits: i

C3bC3b 、、 C3dC3d 、、 C3dg can bind also)C3dg can bind also)..[CD19][CD19] Transfer Ag-signals from CD21 into the Transfer Ag-signals from CD21 into the

B cell to promote cell activation.B cell to promote cell activation.

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Epstein-Barr virus (EBV) binds to CD21 to invade into a B cell

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The Epstein-Barr virus is a member of the herpes family. A person can develop chronic Epstein-Barr infection, infectious mononucleosis, Burkitt’s lymphoma, or Hodgkin’s disease.

EBV infiltrates the squamous epithelial cells within the tongue.

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chronic Epstein-Barr infection

Burkitt’s lymphoma Hodgkin’s disease

Mononucleosis

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[Important Members][Important Members] -CD40-CD40

-B7(CD80/CD86)-B7(CD80/CD86)

-LFA-1-LFA-1

[Important Members][Important Members] -CD40-CD40

-B7(CD80/CD86)-B7(CD80/CD86)

-LFA-1-LFA-1[Function][Function] Transduce accessory Transduce accessory (2nd)(2nd) signals signalsinto a B cell for its complete activation into a B cell for its complete activation (proliferation & differentiation)(proliferation & differentiation)..

[Function][Function] Transduce accessory Transduce accessory (2nd)(2nd) signals signalsinto a B cell for its complete activation into a B cell for its complete activation (proliferation & differentiation)(proliferation & differentiation)..

III. Accessory Molecules

III. Accessory Molecules

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(B(B Cell)Cell) (T Cell)(T Cell) EffectEffect

CD40CD40 CD40LCD40L B B Act.Act.

CD80/86CD80/86 CD28 CD28 CTLA-4CTLA-4

T Act.T Act. T Inh.T Inh.

LFA-1LFA-1 ICAM-1ICAM-1 B & TB & T Act. Act.

Molecule/Ligand/Function

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B Cell

Th Cell

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[Two B Subsets] -B1 cell ， CD5+ ， innate. -B2 cell ， CD5- ， adaptive.

Sec.3B Cell

Subpopulations

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FeaturFeatur

eeB-1B-1 B-2B-2

SpecificitySpecificity PoorPoor StrongStrong

RenewRenew SelfSelf Bone Bone marrowmarrow

Th AssistTh Assist (-)(-) (+)(+)

Memory BMemory B (-)(-) (+)(+)

Major Ig ClassMajor Ig Class IgMIgM IgGIgGAgAg CarbohydraCarbohydra

teteProteinProtein

Origin timeOrigin time Fetal Fetal Neonatal Neonatal

DistributionDistribution Lamina proprLamina propriaia

2nd 2nd Lymphoid Lymphoid

OrganOrgan

Comparison of B1 &Comparison of B1 & B2B2

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Sec.4Functions of B Cell

Sec.4Functions of B Cell

-Ab Production(Ab Functions)-Ag Presentation

-Ab Production(Ab Functions)-Ag Presentation

55 Double Signals & Accessory Molecules

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The lower left a B-lymphocyte also phagotyzing particles.

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1.Concepts: V(D)J Recombination, Junction Diversity, Somatic Hypermutation, B-Cell Clone Deletion.

2.Features of B1 & B2 cells.

1.Concepts: V(D)J Recombination, Junction Diversity, Somatic Hypermutation, B-Cell Clone Deletion.

2.Features of B1 & B2 cells.

[Key Points][Key Points]