Chapter 13Lymphocyte Maturation and Antigen Receptor Expression

Contents

Part Introduction of hematopoietic stem cell

Part T cell maturation and B cell maturation

Part BCR diversity and TCR diversity

Chapter 13Lymphocyte Maturation and Antigen Receptor Expression

Part Hematopoietic stem cellPluripotent hematopoietic stem cell--bone marrowMarker: CD34, CD117(c-kit)Myeloid progenitor + lymphoid progenitor Myeloid progenitor EPO, TPO, CSF, ILLymphoid progenitor NK, DC, T, B NK marker: CD56+CD16+T cell: TCR, CD3, CD4/CD8, CD28, CD2, LFA-1B cell: BCR, IgIg, CD19/CD21/CD81,CD40 B7, MHC, CKR

Part T cells maturation and B cells maturation Development and differentiation of T cells

Development and differentiation of B cells

Section Differentiation of T cells in thymusThymic microenviroment

Differentiation course of T cells

Selection of T cells in thymus---- positive selection and negative selection

1. Thymic microenvironment

2. Differentiation course of T cells 1) pro T cells CD3- TCR- CD4- CD8- TCR chain starts to rearrange DN

2) pre T cellsCD3+ TCRpT: CD4+ CD8+

3) immature T cells DP CD3+ TCR+ CD4+ CD8+ the rearrangement of TCR chain

4) mature T cells CD3+ TCR+ CD4+ or CD3+ TCR+ CD8+ SPTCR rearrangementThymus selection

Differentiation of T cells in thymusChanges in thymusTCR rearrangement ----functional TCRPositive selection and negative selection T cells acquire MHC restriction and Self tolerance

3. Selection of T cells in thymusDepend on TCR , MHC and Ag peptide TCRMHC: positive selection TCR---self antigen peptide : negative selectionDuring the course from DP(double positive) cells to SP(single positive) cells

Positive selection DP cells whose TCRs recognize and combine with MHC molecules can differentiate and develop continuously----SP DP cells whose TCRs cant recognize with MHC molecules or bind with high affinity go apoptosis Get self MHC restrictionMHC molecules play an important role in positive selection: MHC-------CD8+ expression MHC-------CD4+ expression

Negative seletioncells whose TCRs cant recognize with self antigen peptide develop and differentiate continuouslySP cells whose TCRs recognize and combine with self antigen peptide tightly go apoptosis or become clonal anergyAcquired self tolerance

Section Development and differentiation of B cellsDifferentiation of B cells in Bone marrow

Differentiation of B cells in peripheral lymphoid tissue

Differentiation of B cells in Bone marrow----Ag independentHematopoietic stem cells Lymphoid progenitor Pro-B cells( chain rearrangement) Pre-B cell( chain + surrogate light chain ) Immature B(mIgM, chain +chain orchain) Mature B(mIgM, mIgD) Functional B repertoire

Negative selection of B cells inbone marrow

2. Differentiation of B cells in peripheral lymphoid tissue----Ag dependantVirgin B/nave B cell most diePlasma cell AbMemory B cell secondary immune response

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 BCR diversity and TCR diversityBCR diversity

TCR diversity

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

1. Gene structure of Ig (human)H chain:14 chromosome V region encoding genes: VH (variable gene segments) 65 DH (diversity gene segments) 27 JH (joining gene segments) 6 Leader sequencesignal peptide C region encoding genes: CH (constant gene segments): C, C, C et al. (11)

L chain(--2 chromosome, --22 chromosome)

V region encoding genes: --V, J 40, 5 -- V, J 30, 4 Leader sequencesignal 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 whilethe C segment encodes the constant domain.

VJC JC JC JC(a) Chain (22 chromosome))(2 chromosome)

2. Gene rearrangement of Ig

V-D-J rearrangement of H chain pro-B cells: D-J V-DJ VDJ DNA

pre-B cells: VDJC VDJ- C 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 mRNAtranscriptionsplicing

C C C3 C 1 C1 C2 C4 C C2C C C3 C1 C1 C2 C4 C C2C C C3 C1 C1 C2 C4 C C2CCCC

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-CEndoplasmic reticulum:assembly--------H chain and L chain (IgM or IgD)

transportation------BCR (membrane Ig, mIg)

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

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 5-10Read Kuby pages 115-117: Allelic Exclusion Ensures a Single Antigenic Specificity

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

Thymus dependent antigenAPC

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

4. Mechanism of Ig diversity Combinatorial diversity human Ig: 65VH27DH 6JH=10530V 40V 5J =200V 30V 4J =120V

C C C3 C 1 C1 C2 C4 C C2C C C3 C1 C1 C2 C4 C C2C C C3 C1 C1 C2 C4 C C2CCCC

Junctional diversity

CDR3 lie in V-DJ or D-J junctionsLose or insert of several nucleotides will increase the diversity of CDR3.N-nucleotides insert by TdT without templateThere is no N-nucleotides insert in L chain

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 Gene structure and rearrangement of TCRGene structure of TCR chain(14 chromosome): V, J, C chain(7 chromosome): V, D, J, C

(14 chromosome)(7 chromosome)

2. Gene rearrangement of TCR TCR chain rearrange first inactivate gene within gene

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

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

5. Characteristics of TCR gene expressionWithout somatic hypermutationMore N- nucleotides insert than BCRMore valid rearrangement in V region of TCR BCR: 1014 TCR: 1016

Comparison of BCR and TCR functional genes

NO. of chain Chromosome V D J C----------------------------------------------------------------------- BCR H 14 65 27 6 9 2 40 5 1 22 30 4 4 TCR 14 70-80 61 1 7 52 2 13 2 7 12 5 2 14 4 3 3 1-------------------------------------------------------------------------

Clonal deletion: Functionally immature cells of a clone encountering antigen undergo a programmed cell death. For example, auto-reactive T-cell are eliminated in the thymus following interaction with self antigen during their differentiation (negative selection). Clonal deletion has been shown to occur also in the periphery. B cells expressing only IgM (no IgD) on their surface when exposed to antigen are eliminated.