Immunoglobulin Genetics

Problem…the immune system makes over one billion different antibody proteins In 1950’s: central dogma stated DNA—to RNA—to protein One gene for each protein Required millions of genes just for the immune system Does not seem possible, but most scientists thought it might be Today we know the human genome is less than 30,000 genes So, what is really going on???

B lymphocyte development

B lymphocyte development (2)

Immunoglobulin Genetics History Same C region could associate with many V regions –IgG Ab with different specificities Same V region could associate with many C regions –Same idiotype on IgG and IgM Ab V and C regions coded for by separate genes

Genes for immunoglobulin proteins are found on different chromosomes

Light Chain Gene Families Germ line gene organization J1J1 C1C1 E J2J2 C2C2 E J3J3 C3C3 E J4J4 C4C4 E P L V1V1 P L VnVn P L V2V2 Lambda light chain genes; n=30 P L V  1 P L V  n P L V  2 J2J2J3J3J5J5 CC E J1J1J4J4 Kappa light chain genes; n=300

Light Chain Gene Families Gene rearrangement and expression L V 1V 1 L V nV n L V 2V 2 J2J2J3J3J5J5 CC J1J1J4J4 DNA PPP E P P E L V 1V 1 L J5J5 CC V 2V 2J4J4 DNA Rearrangement VC J V C Protein Transport to ER Protein Translation C L V J E L V CJ RNA Transcription Primary transcript C L V J RNA Processing RNA mRNA

Heavy Chain Gene Family Germ line gene organization

Heavy Chain Gene Family Gene rearrangement and expression P L V1 P L Vn P L V2 D2D2 D3D3 D1D1DnDn CC CC J2J2J3J3J5J5J1J1J4J4 E DJ rearrangement P L Vn P L V1 P L V2 D2D2 D1D1 CC CC J5J5 J4J4 E DNA VDJ rearrangement D2D2 CC CC J5J5 J4J4 E P L V1V1 P L V2V2 DNA Transcription D2D2 CC CC J5J5 J4J4 E L V2V2 RNA Primary transcript

Heavy Chain Gene Family Gene rearrangement and expression E Primary transcript E J4J4D2D2 V2V2 CC CC J5J5 L CC CC J5J5 L D2D2J4J4 V2V2 RNA Processing AnAn mRNA for C  CC L DJV CC L DJV AnAn mRNA for C  Translation C  heavy chain CC L DJVCC L DJV C  heavy chain Transport to ER C  heavy chain V C CC DJVCC DJV C  heavy chain V C

Mechanism of DNA Rearrangements Recombination signal sequences (RSS) –Nonmer –Heptamer –1 or 2 turn signals Rag-1 and Rag-2 L V 2 J C 1 Lambda light chains L V 1 J C 2 Kappa light chains Heavy chains L V 2 J C D Heptamer CACAGTG GTGTCAC Nonamer ACAAAAACC TGTTTTTGG 23 bp Two turn RSS 2 Nonamer GGTTTTTGT CCAAAAACA Heptamer CACTGTG GTGACAC 12 bp One turn RSS 1

Order of Ig Gene Expression + - Heavy chain gene VDJ recombination Productive rearrangement ? Yes No Transcribe/translate 1st 2nd or Apoptosis Progenitor B cell Pre B cell    (  ) Heavy chain Paternal Maternal

Order of Ig Gene Expression + - Light chain gene VJ recombination Productive rearrangement ? Yes No Transcribe/translate Light chain 4 th Apoptosis 1st -  2nd -  3rd -  4th -  or Pre B cell Mature B cell     + 

Order of Ig Gene Expression Explains: Why an individual B cell only produces one kind of Ig with one kind of heavy and one kind of light chain Why an individual B cell makes only makes antibodies of one specificity Why there is allelic exclusion

Origin of Antibody Diversity History Definition – Diversity (repitoire) is the total of all the Ab specificities that an organism is capable of expressing Theories –Germ line theory –Somatic mutation theory

Origin of Antibody Diversity Current concepts Multiple V genes V-J and V-D-J joining Junctional diversity N region insertions – Amino acid sequences not encoded in the germ line D region TGGCCC Pro Trp CCCCCC V gene TGG D region CCCCCC V gene TGG CGGCCC Pro Arg CCCCCC V gene TGG D region CGCCCC Pro

- GTCAATG - GTCAATG -+ Break DNA TdT adds nucleotides Ligate N region insertionIntron JD J J J D D D Generation of N region insertions

Origin of Antibody Diversity Current concepts Multiple V genes V-J and V-D-J joining Junctional diversity N region insertions Somatic mutation Combinatorial association Multispecificity

Generation of Diversity Ag independent process Clonal selection

Copyright © 2007 by W. H. Freeman and Company ANTIBODY GENE REARRANGEMENTS CLASS SWITCHING : Selection of CH Genes Mechanism Unclear, but Involves Switch Region Switch Recombinase Isotype Specific Cytokine Signals

Class Switching among Constant-Region Genes