IMMU - Chapter 6 The Development of B Lymphocytes

Outcome of B cell development

1. Generation of B cells with diverse array of BCR

2. B cells capable of recognizing foreign antigen

3. Selection of B Cells with no reactivity to self-antigen (Positive selection)

4. Removal of B cells with strong reactivity to self-antigens (Negative selection)

Lymphoid tissues

Organized structure that support immune responses

Central/primary

Sites of lymphocyte development and maturation

• Bone marrow - B cells

• Thymus - T cells

Peripheral/Secondary

Sites of lymphocyte activation and differentiation into effector cell types

• spleen, lymph nodes, adenoids, tonsils, mucosal tisssue

Tertiary lymphoid tissue

Chronic inflammation can lead to the development of tertiary lymphoid tissue

Bone niche

B cells develop in the bone marrow and migrate to peripheral lymphoid organs, where they can be activated by antigens

B cell development occurs in..

Bone marrow

B cell activation starts when

The B cell meets an antigen in the lymph node

Bone marrow microenvironment is composed of

• a cellular compartment

• An extracellular matrix

• A liquid compartment

A liquid compartment

A mixture of growth factors, cytokines

Extracellular matrix

A complex network of matrix proteins

Cellular compartment

Cells of hematopoietic and non-hematopoietic origin and effector immune cells

Bone marrow stromal cells

A specialized network of non-lymphoid connective tissue that provide structural and physiological support to hematopoietic cells

Role of bone marrow enviornment

• provide cell-cell contact (structural support) for developing B cells and other immune cells

• Provide signals through cytokines for growth, survival and maintenance → IL-7 and CXCL12

What is notch?

• a cell surface receptor that interacts with a transmembrane

• Four different receptors

• Notch1 signaling - promotes T-cell differentiation from common lymphoid progenitor

IL-7 promotes the

• survival

• Proliferation

• And maturation of developing lymphocytes

The early stages of B-cell development are dependent on..

Bone marrow stromal cells

B-cell lineage

Notch 1 signal absent

Early IL-7R expression

Early T cell Progenitor

Notch 1 signal present

No early IL-7R expression

ILCs

Notch 1 signal present

IL-7R not required

Different transcription factor expression

Cells at different stages of development are identified by

Different combinations of CD proteins on their surface

CD34 are present on all HSC

• CD127 is IL7Ra

• CD19 is Pan B cell marker - all B cells have it

Checkpoint

• molecular mechanism by with the B cell determines if the preceding step has been successfully completed

1st checkpoint - did developing B cell successfully express new IgH as pre-BCR on cell surface

2nd checkpoint - is the B-cell self reactive?

Step 1 - Heavy Chain rearrangement and

1st checkpoint

Step 2- Light chain rearrangement

2nd checkpoint

Stages of B cell development

1. Early pro-B cell

2. Late pro-B cell

3. Large pre-B cell

4. Small pre-B cell

5. Immature B cell

6. Mature B cell

Cell proliferation

Increasing the number of cells

Cell differentiation

Differentiating into another type of cell

Early pro-B cell and Large pre-B cell

Does cell proliferation

(Increasing the number of cells)

What determines this sequence?

1. RSS sequence dictates the order of rearrangements

2. Enhancers and promoters initiate transcription in a sequence and linked manner

Rearrangement and transcription of immunoglobulin genes are linked processes

• In HSCs, Ig Heavy-chain locus is closes

• In pro-B cells, B cell-specific transcription factors bind Ig enhancers and promoters

• In late pro-B cell, DJ rearrangement occurs

• In large B-cell, V-DJ rearrangement occurs

B cell lineage occurs when:

• Notch 1 signal is absent

• Early IL-7R expression

Early T cell progenitor occurs when:

• Notch 1 signal present

• NO Early IL-7R expression

ILCs (innate-like cells) occurs when:

• Notch 1 signal present

• IL-7R not required

• Different transcription factor expression

Does the rearrangement give rise to a fully function heavy chain?

If no:

The other chromosome rearranges

Does the rearrangement give rise to a fully function heavy chain?

The rearrangement is then tested

If it fails, to produce a productive rearrangement

• The developing B cell dies by apoptosis

Option 1: B cell maturation is coupled to immunoglobulin rearrangement

Option 1: B cell maturation is coupled to immunoglobulin rearrangement

Only one productive rearrangement of the heavy chain is allowed, which is called

Allelic exclusion

Allelic exclusion at the immunoglobulin loci gives rise to B cells have antigen receptors of:

Monospecifity (single specifity)

Apoptosis

Used by immune system for deletion of immune cells

• does not induce any inflammatory reaction

How do you test if the new immunoglobulin heavy chain (IgH) is functional?

Large pre- cell express

• a newly made IgH

• CD79a/b

• A surrogate light chain

What would happen if developing B cells could not express Lambda 5 or VpreB

B cell development would stop at the pre B-cell stage because the first checkpoint signal was not received

• without checkpoint line, it cannot rearrange light chain

The surrogate light chain (SLC) is made up of two proteins:

VpreB and lambda 5

Pre B-cell receptor complex:

1. Immunoglobulin heavy u chain

2. Surrogate light chains

3. Signaling component: IgA (Cd79A) and IgB (CD79B)

4. Other signaling molecules: BTK, BLNK involved

Checkpoint 1 test

Successful signaling involves this protein:

BTK (bruton’s tyrosine kinase)

Importance of pre-BCR signaling

Crucial for Allelic exclusion

• inhibits further heavy-chain locus rearrangement

• Enforces Allelic exclusion

• Induces proliferation of pre-B cells

• Commences light chain gene rearrangement

Pre-BCR signaling

there are no foreign (non-self) antigen in bone marrow

• it can signal independent of ligand engagement (tonic signaling) or self-crosslinking by binding to self-antigens (found in the bone marrow)

After checkpoint 1:

Large pre-B cells undergo proliferation, this results in making

• 100 small B-cells with the same heavy chains

• SLC is no longer made

• Light chain rearrangement begins

Heavy chain same + different light chain = BCR with different antigen specificites

These cells divide 5-6 times (cell proliferation) before starting light chain arrangement

Light chain rearrangement begins

• Pre-B stops dividing and becomes small Pre-B

• Rag genes turn on again - RAG ½ are made

• K-light chain rearrangement begins

• Several chances for productive rearrangement

• Isotopic exclusion (k or lambda light chain)

Light chain rearrangement begins with k light chain

Rearrangement of light chain loci by pre-B cells is relatively efficient

• k locus is first to rearrange

Heavy chains get

One chance per locus

Light chains get

Multiple chances per locus

Isotopic exclusion

You can make a B cell receptor with a heavy chain and one type of light chain (k or lambda)

Check point 2 is the self-reactivity check

If checkpoint 2 is pass → next step is B cell maturation

Before checkpoint 2 diagram

Before checkpoint 2 diagram

Why is recognition of self-antigen bad?

• it could potentially cause autoimmune disease

• Examples of self-antigen: glycoproteins, proteoglycans, glycolipids

Central tolerance

Occurs in central lymphoid organs (bone marrow and thymus)

• Elimination of any developing T or B lymphocytes that are relative to self in the cLO

Peripheral tolerance

Occurs in peripheral lymphoid organs

• B cells that escape the test in bone marrow may still be removed form the repertoire after leaving bone marrow

• Elimination of any mature or T or B lymphocytes that are reactive to self in the PLO

Self-tolerance depends on the:

Concerted action of a variety of mechanisms that operate at different sites and stages of development

Immunologically privileged sites represents:

A special microenvironment where the systemic immune response to allo- (foriegn) and autoantigens (self) are remarkably reduced

Immune tolerance by privilege

Immune system will ignore antigens expressed or present in immunologically privileged sites

• these sites are NOT under immune surveillance

Positive selection - selection of non-self reactive B cells

Immature → Mature (through B cell maturation)

Co express IgM and IgD

Leave bone marrows

What happens to self-reactive immature B cells in the bone marrow?

1. Death by apoptosis (programmed cell death)

2. Receptor editing (attempt at losing self-reactivity)

3. Low affinity binding

Negative selection - deletion of self-reactive B cells

Attempt at rescue

Immature B cells are retained in the bone marrow for receptor editing

Option 1 death by apoptosis

Option 1 death by apoptosis

Option 2 receptor editing (to attempt losing self-reactivity)

Option 2 receptor editing

Rescue attempt for self-reactive B cells

Rescue attempt for self-reactive B cells

Option 3 Low affinity binding enter state of anergy

• IgD is still expressed

• State of Anergy: B cell and no longer be activated

Clonally ignorant B cells

Weakly self-reactive B cells that fail to induce receptor editing, deletion or anergy, mature and differentiate into follicular or marginal zone B cells

Anergic B cells

B cells that can no longer be activated are in a state of developmental arrest

• these cells cannot be activated in the periphery

B cell maturation

Immature B cells that only express IgM, then mature and co-express IgD and IgM

There was 2 ways to exit the bone marrows

1. As mature B Cells (naive B cells) that co-express IgM and IgD after passing checkpoint 2

2. As transitional B cells which are still immature B cells that have passed checkpoint 2, express only IgM on their cell surface

Timing of proteins involved in immunoglobulin gene rearrangement and expression

• IL-7R expressed early on

• RAG ½ ar not expressed when heavy chain rearrangement is done

• Surrogate LC are not expressed once light chain rearrangement begins

Successful signaling via pre-BCR and BCR involves these proteins

CD79A/B and Btk

Mutations in Btk cause

X linked agammaglobulinemia

• they fail to signal through the pre-B cell receptor

B2 B cell

Conventional B cells

B1 B cell

Made during fetal development

• limited repertoire

• Make natural antibodies

• Primarily exhibit T-cell independent activation

Follicular (FO) B cell

B cells mainly in the lymphoid follicles of secondary lymphoid organs (SLOs)

Marginal zone (MZ) B cell

Found mainly in the marginal zone of the spleen and serves as first line of defense against blood-borne

Regulatory B (Breg) cell

An immunosuppressive B cell

Plasma cell

Activated B cells

Memory B cell

Arising from B cell activation (post-infection or post-vaccination)

B-1 B cells

• primarily made during fetal development

• Primarily exhibits T cell-independent activation

Differences between B1 and B2 B cells

Difference between B1 and B2 B cells

Multitalented antigen

Carry multiple epitopes (crosslinking)

Soluble self-molecule

Crosslinking

Univalent antigen

Carry only one copy of an epitope (antigenic determinant)

• non-crosslinking

Transitional B cells that recognize self-antigens undergo peripheral tolerance

These B cells are still subject to tolerance in the spleen after engagement of their sIgM receptor by a self-antigen

How do naive (mature) B cells know when to leave the bone marrow?

• B cells express S1PR1 and exit the bone marrow into the blood

• Extracellular S1P levels are high relative to cellular S1P

Journey of naive B cells

• circulate and recirculate throughout the body until they encounter their specific antigen

• Varied half-life of 1-2 months, some may survive to 3 months

naive B cell in lymph node

They encounter their specific antigen/pathogen then they stop recirculating

• ones that DO NOT encounter their specific antigen → obtain survival signals

Know the protein names with receptor genes

Know the protein names with the receptor genes

Chapter 6 summary

Chapter 6 summary