Immunology #6 - B Cell Development

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Hematopoiesis

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- B cell arise in the bone marrow from progenitor lymphoid cells
- During development, the progenitor B cell acquires its antigen specific receptor (immunoglobulin)

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Antigen Specific Receptors

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B lymphocytes (B cells) express antigen recognizing receptors (membrane immunoglublin/ antibodies) on their cell surface. These receptors mediate interaction between the B cell and the intact antigen.

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mIg

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- Membrane immunoglobulin
- Expressed in association with a heterodimer, CD79a/CD79b (Igα-Igβ)

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Structure of immunoglobulins

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- Composed of 2 identical heavy chains, and 2 identical light chains linked by disulfide bonds
- Each chain is made up of a variable region and a constant region
- In naïve mature B cells, the heavy chain constant regions are mu (μ) and delta (δ), while the light chain constant regions are either kappa (κ) or lambda (λ)

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Light chain variable region

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Composed of segments designated "V" and "J"

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Heavy chain variable region

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"V", "D", and "J" segments, where the letters "V", "D", and "J" refer to the Variable, Diversity, and Joining segments

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Paratope

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The antigen binding site is contained within the combined variable regions of the light and heavy chains

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Interactive forces

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- Non covalent - antigens can come on and off
- Attraction between the antigen binding site and an epitope determines the affinity = AVIDITY

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Somatic recombination

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- DNA in the loci that encode the variable regions is cut and recombined to make an intact gene for the variable regions of the light and heavy chains
- The variable region genes and constant region genes are transcribed to hnRNA which is spliced to mRNA which is in turn translated to a light chain and a heavy chain
- This occurs in the bone marrow during B cell development

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RAG-1 and RAG-2 genes

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- Recombination activating genes which trigger somatic recombination

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Light Chain Rearrangement

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- Selection and ligation of 1 "J", and 1 "V" to form a VJ segment

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Heavy Chain Rearrangement

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- 1 "D" and 1 "J" are randomly selected, to form a DJ gene segment, which then combines with a randomly selected "V" gene segment to form the variable region (VDJ) of that particular heavy chain

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Combinatorial diversity

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- During the recombination process of either the light or heavy chain variable region, intervening unselected "Vs", "Ds" or "Js" are deleted

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Terminal deoxynucleotidyl transferase

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Incorporation of nucleotides at junctions is mediated by a template independent DNA polymerase

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Allelic Exclusion

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- When there is successful rearrangement of a heavy chain variable region from one chromosome - it inhibits the somatic recombination of the heavy chain variable region on the other member of the chromosome pair, a process known as allelic exclusion
- All mIg present on the surface of any one B cell will have the same heavy chain variable region

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Stages in the Bone Marrow

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- B cell differentiation in the bone marrow occurs prior to any exposure to foreign antigen
- It is characterized both by the expression and silencing of distinct sets of genes

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Pro-B cell Stage (Transcription)

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- Transcription of multiple genes, including RAG-1 and RAG –2, CD19, Tdt and CD79, required for differentiation of the B cell in the next developmental stage

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Pre-B cell Stage (Expression of Pre-BCR)

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- Pre-B cell receptor (pre-BCR) in association with the CD79a/CD79b heterodimer
- Signaling via the pre-BCR complex directs proliferation and further differentiation of pre-B cells
- CD19 and CD20 is expressed at this stage of the developing B cell

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Immature B cell Stage (Tolerance Induction)

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- Pre-BCR to BCR
- Assembly of rearranged light chains and the heavy chains with CD79 heterdomers to form the BCR
- Pre-BCR is downregulated
- Apoptosis or anergy of auto-reactive mIg (tolerance induction)
- BCR on surface

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Mature B cell Stage (Alternative Splicing)

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- Co-expression of cell surface IgM and IgD, which is the consequence of alternative splicing
- Mature naive B cells leave the bone marrow, enter the blood stream, migrate to peripheral lymphoid tissues, and re-circulate if they do not encounter antigen in a secondary lymphoid tissue

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Summary

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Exit from blood

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- Mediated by L-selectin on B cells on high endothelial venules (HEV)
- Entry into the spleen occurs at terminal branches of the central arterioles
- Failure to encounter antigen while transiting through secondary lymphoid tissues results in naive B cells entering the re-circulating pool via efferent lymphatics, en route to the thoracic duct or right lymphatic duct

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Bruton’s

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- Mutations in the Btk kinase are associated with X-linked agammaglobulinemia
- Plays a critical role in B cell activation, differentiation, and proliferation
- Btk defect occurs at the pro-B to pre-B cell transition
- Patients with XLA have (DEFECTIVE) pre-B cell populations in the bone marrow such that these cells fail to mature and enter the circulation
XLA
- Profoundly reduced numbers of B cells in the peripheral blood, and serum Ig levels of all classes are low