Chapter 14 B Lymphocytes

Contents  B cell receptor and B cell complex  B cell accessory molecules  B cell subpopulations  Functions of B cells  B cell maturation  BCR diversity and TCR diversity

I. B cell antigen receptor and B cell receptor complex  BCR  BCR complex  Function: Recognize antigen

1. B cell receptor----BCR Membrane immunoglobulin (mIg) on B cells: mIgM, mIgD Recognize and bind antigen specifically

2. BCR complex BCR and Ig  - Ig  (heterodimer) Ig  (CD79a), Ig  (CD79b) Participate in BCR formation ITAM------bind to tyrosine kinase Transmit activating signal

II. Accessory membrane molecules on B cells 1.CD19, CD21(CR2), CD81(TAPA-1) ---- B cell co-receptor complex  CD21(CR2): receptor of C3dg,C3d and iC3b ----Enhance the binding of BCR and antigen ----Pass activating signal to CD EB virus receptor  CD19: Transmit activating signal into B cell

CD40 on B cell binds to CD40L on activated T cell Transmit an important co-stimulatoryco signal to B cells Upregulate expression of B7 on B cells Participate in class switching of antibody 2. CD40----co-stimulatory receptor

3. Co-stimulatory molecule: B7 B7-1 (CD80) and B7-2 (CD86) Expressed on B cells or other APC B7-CD28: activation signal B7-CTLA-4: inhibitory signal 4. MHC molecules Class Ⅰ, Ⅱ MHC molecules

5. Mitogen receptor SPA, LPS PWM 6. Cytokine receptor IL-4R, IL-5R, IL-6R 7.Fc receptor (CD32---Fc  RII-b) Related to immunological regulation

III. B cell subpopulations According to expression of CD5 or not  B1 cell (CD5 + )  B2 cell (CD5 - )

Comparison of B1 and B2 cells B1 B2 Development early late BCR mIgM mIgM and mIgD CD5 + - Reproduction self-renewing from pre-B cell in BM Recognized Ag TI-Ag and auto-Ag TD-Ag Ab type IgM >IgG IgG >IgM Ab avidity low high Second IR - + Function innate immunity adaptive immunity

IV. Functions of B cells  Produce the antibody----HI  Present antigen----APC  Participate in immunological regulation: secrete various cytokines, Fc  RII-b

T lymphocyteB lymphocytefunctions Antigen receptorTCRBCRBind to Ag, First signal Ag receptor complex CD3 and ξ 链 Igαand IgβTransmit first signal Co-receptorCD4,CD8CD19-CD21 complex Help to bind Ag ， Transmit first signal Co-stimulatory receptor CD28CD40Second signal, Transmit second signal Co-stimulatory molecules CD40LB7(CD80/CD86)Offer second signal MHC Iexpression Antigen processing MHC IIActivated expression expressionAntigen processing The comparison of main membrane molecules between T and B cells

Section Ⅱ Development and differentiation of B cells Differentiation of B cells in Bone marrow Differentiation of B cells in peripheral lymphoid tissue (B cell mediate immune response, HI)

μ

1.Differentiation of B cells in Bone marrow----Ag independent Hematopoietic stem cells Lymphoid progenitor Pro-B cells(  chain rearrangement) Pre-B cell(  chain + surrogate light chain ) Immature B(mIgM,  chain +κchain orλchain)Immature B Mature B(mIgM, mIgD) Functional B repertoire

Negative selection of B cells in bone marrow

2. Differentiation of B cells in peripheral lymphoid tissue----Ag dependant Virgin B/naïve B cell most die Plasma cell Ab Memory B cell secondary immune response

3.Events in the differentiation of B cells: 3. Events in the differentiation of B cells: Gene rearrangement of Ig Negative selection Immature B cells : mIgM--self antigen mIgM -- self antigen apoptosis or anergy surviving to develop mature B cells

Questions? Why can TCR or BCR recognize so many Ag in nature? Why does IgM produce earlier than others? How does Ig produce BCR and Ab? How can B produce different type of Igs? ?

Part Ⅲ BCR diversity and TCR diversity  BCR diversity  TCR diversity

Gene structure of Ig Gene rearrangement of Ig Characteristics of Ig gene expression Mechanism of Ig diversity Section Ⅰ BCR diversity Section Ⅰ BCR diversity

1. Germ line gene structure of Ig (human) H chain:14 chromosome V region encoding genes: V H (variable gene segments) – 65 D H (diversity gene segments) – 27 J H (joining gene segments) – 6 Leader sequence—signal peptide C region encoding genes: C H (constant gene segments): Cμ, Cδ, Cγ et al. (11)

L chain(  --2 chromosome, --22 chromosome) V region encoding genes:  --V , J  – 40, 5 -- V, J – 30, 4 Leader sequence—signal peptide C region encoding genes: C  (1); C (4)

In heavy chains, the V, D and J segments encode the variable domain while the C segment encodes the constant domain. In light chains, the V and J segments encode the variable domain while the C segment encodes the constant domain.

V J C J C J C J C (a) Chain (22 chromosome)) (2 chromosome)

2. Gene rearrangement of Ig 2. Gene rearrangement of Ig V-D-J rearrangement of H chain pro-B cells: D-J V-DJ VDJ DNA pre-B cells: VDJCμ VDJ- Cμ RNA mRNA V-J rearrangement of L chain pre-B cells: V  -J  V  J  DNA immature B cells: V  J  C  V  J  -C  RNA mRNA transcription splicing

C  C  C  3 C  1 C  1 C  2 C  4 C  C  2 CC CC CC CC 

The expression of BCR The expression of BCR Intranuclear: DNA rearrangement: V region encoding gene (VDJ or VJ) Transcription and splicing leader sequence + V region encoding gene + C region encoding gene (L gene-V gene –C gene) Extranuclear: Translation nascent peptide L-V-C Endoplasmic reticulum: assembly H chain and L chain (IgM or IgD) transportation------BCR (membrane Ig, mIg)

3. Characteristics of Ig gene expression ① recombination enzyme: RAG (recombination activating gene) TdT (terminal deoxynucleotidyl transferase) other DNA enzymes

② Allelic exclusion and isotype exclusion ② Allelic exclusion and isotype exclusion Allelic exclusion: only one of the two alleles in homologous chromosomes can be expressed. Isotype exclusion: only one of the two types of light chain genes can be expressed(  : =65:35).

Kuby Figure 5-10 Read Kuby pages : Allelic Exclusion Ensures a Single Antigenic Specificity

③ Isotype switching ③ Isotype switching ( class switching ) Ag activated B cells proliferate VDJ is switched to recombine with another C region encoding gene IgM IgD, IgG, IgA, IgE Switching region

④ Membrane type (BCR) and Secretory type Ig (Ab)

4. Mechanism of Ig diversity ① Combinatorial diversity human Ig: 65V H ×27D H ×6J H =10530V 40V  ×5J  =200V 30V ×4J =120V

C  C  C  3 C  1 C  1 C  2 C  4 C  C  2 CC CC CC CC 

② Junctional diversity  CDR3 lies in V-DJ or D-J junctions  Lose or insert of several nucleotides will increase the diversity of CDR3.  N-nucleotides insert by TdT without template  There is no N-nucleotides insert in L chain

③ Somatic hypermutation ③ Somatic hypermutation Ag activated B cells proliferate gene mutation in V region encoding genes affinity maturation mature B cells which finished V gene rearrangement

Section Ⅱ Gene structure and rearrangement of TCR 1.Gene structure of TCRαβ α chain (14 chromosome): V, J, C β chain (7 chromosome): V, D, J, C

(14 chromosome) (7 chromosome)

2. Gene rearrangement of TCRαβ TCR β chain rearrange first Inactivate δ gene within α gene

3. Gene structure of TCRγδ γ chain (7 chromosome): V, J, C δ chain (14 chromosome): V, D, J, C

(14 chromosome) (7 chromosome) 4. Gene rearrangement of TCR γδ No junctional diversity in TCR γδ No junctional diversity in TCR γδ

5. Characteristics of TCR gene expression Without somatic hypermutation More N- nucleotides insert than BCR More valid rearrangement in V region of TCR BCR: TCR: 10 16

Comparison of BCR and TCR functional genes NO. of chain Chromosome V D J C BCR H  TCR    

What you should know of this lecture  Definition of BCR/BCR complex  Major surface membrane molecules on B cells (comparison)  Comparison of B1 cell and B2 cell  Functions of B cells  B cell maturation