Adaptive Immunity Central objective: Protect against foreign invaders memoryCreate memory of invasion to prevent recurrent infection specificResponse must be highly specific to the invader

Ig Isotypes: Classes and Subclasses  Ig isotypes are defined by the C- terminal portion of the molecule  There are 5 classes of Ig and there are 4 sub-classes of IgG.  Isotype class determines effector function.

Ig Isotypes

Ig Isotypes Localize to Different Anatomical Sites in the Body

Ig Isotypes Have Different Effector Functions

When Do Isotypes Matter? The primary -> secondary immune response: Increase Ig titer Switch from IgM to IgG Increase Ag binding affinity

When Do Isotypes Matter? Slide 2 Primary ->Secondary Immune Response Transition

Primary/Secondary Immune Response Slide 2 Events associated with an immune response over time Events associated with an immune response over time:  Increase of IgM titer specific  Increase of IgM titer is specific to the immunizing Ag

Isotypes Switch: IgM->IgX Three Types of Switch: RNA level IgM->IgD Membrane->Secreted DNA level Class Switch Recombination

IgM and IgD Are Coexpressed in Mature B Cells Differentiation to the mature B cell stage is marked by the co- expression of IgM and IgD on the plasma membrane

A Shift of IgM to IgM/IgD Expression in Mature B cells Mechanism: Alternative RNA polyadenylation and splicing

In Mature B Cells Ig is Located on the Mb and Functions as the BCR Location of Ig on the cell surface is transmb and requires the presence of the mb exon

In Plasma Cells Ig is Secreted Secretion of Ig from the cell requires the presence of the secretion exon

 In mature B cells Ig is located on the plasma mb.  Ig is secreted from terminally differentiated plasma cells Secreted Ig Promotes Critical Functions

 In mature B cells Ig is located on the plasma mb.  Ig is secreted from terminally differentiated plasma cells Secreted IgM and IgA are Multimerized by J Chain

Isotypes Switch: IgM->IgX Three Types: RNA level IgM->IgD Membrane->Secreted DNA level Class Switch Recombination

C Regions Are Arrayed in the IgH Locus

Looping Out and Deletion Model of SR

Germline Transcripts are Required for SR

When Do Isotypes Matter? The primary -> secondary immune response: Increase Ig titer Switch from IgM to IgG Increase Ag binding affinityIncrease Ag binding affinity

Affinity Maturation and Ig Isotype Switch In a primary immune response, affinity of Ig for its Ag is usually not high enough to immediately clear the pathogen from the system. However, affinity is increased by somatic hypermutation (SHM). DNA level SHM alters V genes at the DNA level

Primary/Secondary Immune Response Slide 2 Acquisition of mutations in the V region of the VH and VL proteins nonrandom distribution Acquisition of mutations in the V region of the VH and VL proteins: nonrandom distribution Protein level Protein level

Primary/Secondary Immune Response Slide 2 The position of the newly acquired mutations is in the antigen binding pocket. Protein level Protein level

Somatic Hypermutation and Affinity Maturation In SHM, individual nucleotides in VJ or VDJ units are mutated, thus adjusting the specificity of and potentially increasing the affinity for Ag.

Somatic Hypermutation and Affinity Maturation cont... Mutations translate into a change in the Ag binding pocket of the Ab. Some mutations are deleterious while others are neutral or advantageous for Ab:Ag fit. Some deleterious mutations generate autoreactive Ig. Question: are mutations random or directed at the level of the genes?

Somatic Hypermutation and Affinity Maturation DNA level

Somatic Hypermutation and Affinity Maturation cont... Selection of high affinity B cell clones leads to affinity maturation of the immune response. B cells expressing high affinity BCR are favored by clonal selection. This occurs when Ab:Ag complexes are highly compatible and possess a “good fit”. The high affinity Ab:Ag interaction transduces a strong signal to the B cell to proliferate.

Clonal Selection: Type I Clonal selection occurs following VDJ in the BM to delete B cells expressing autoreactive BCR.

Mutations which increase affinity of Ab for Ag will create a BCR which promotes strong proliferation. Selection also occurs in the periphery following SHM on the GC to delete newly autoreactive BCR and to preclude the escape of self-reactive B cells. Clonal Selection: Type II

SHM Occurs in the Germinal Center B Cells

The Surprising Link between CSR, SHM and Gene Conversion AID  Activation induced deaminase (AID) is responsible for CSR and SHM.  It is the only B cell specific gene product required for both these genetic alterations.

The Surprising Link Between CSR, SHM and Gene Conversion How does AID work?  AID has homology to the APOBEC family of proteins which are involved in RNA editing  AID is also a cytidine deaminase and converts dC ->uracil  Uracil is mutagenic and triggers the BER pathway

AID Deaminates dC and Triggers the BER Pathway

What happens when AID is mutated? What happens when UNG is mutated? What happens when DSB repair proteins are mutated?

Hyper-IgM Syndrome: No Secondary Isotypes Four Types: Cause???? Mutations in: CD40 (T cell); CD40L (B cell) AID UNG Subgroup with unknown cause

CSR Requires DNA Repair Proteins DNA Repair Proteins Involved in CSR: DNA-PKcs, Ku70, Ku80, H2AX, NBSl, ATM MMR (Msh2, Msh6, Pms2, Mlhl)

The End

U=T During Replication

BCR and TCR Break the Rules of Mendelian Genetics Monospecific Ag receptors: Only one H chain and one L chain is expressed per B/T cell whereas each cell has 2 alleles for H and L chains