Immunoglobulin Gene Structure

The cells of the B line synthesize immunoglobulins. They are either produced at a membrane (on the surface of the B-lymphocytes) or are secreted (by the plasmocytes)

1. Dr. Dinesh C. Sharma,
Associate Professor & Head
Deptof Zoology
Km. MayawatiGovt. Girls P.G. College, Badalpur, Gb nagar

2. Animmunoglobulin (Ig)consists
of
2identicallightchains(L)
and
2identicalheavychains(H)

3. ForexampleIgG-type;atthethree-dimensionallevel,anIgchainconsistsofoneN-
terminalvariabledomain,V,andone(foranLchain)orseveral(foranHchain)C-
terminalconstantdomain(s),C.
ThecellsoftheBlinesynthesizeimmunoglobulins.Theyareeitherproducedat
amembrane (onthesurfaceoftheB-lymphocytes)oraresecreted(bythe
plasmocytes).

4. 1-Light chains (Kappa)
1.1. Kappa chain: V-J rearrangements
•IGK (kappa) genes at 2p11on chromosome 2.
•Multiple IGKV genes for the variable region, V (76 genes, of which 31 to 35 are functional); 5 IGKJ genes for the
junctional region, J; 1 IGKC gene for the constant region, C; the V, J and C genes are separated in the DNA of the
genome ('germline' configuration of the Ig genes).
•These are multigene families
•First the DNA is rearranged: this makes it possible to join 1 V and 1 J; the intermediate sequences are then deleted,
•The pre-messenger RNA is copied (transcription); this includes introns,
•Then comes splicing: the elimination of the introns from the pre-messenger RNA , to yield mature, messenger RNA,
•This is followed by protein synthesis (known as 'translation').
Note-
•It is crucial not to confuse DNArearrangementswith RNAsplicing.
•Only the genes for the immunoglobulins and T-receptors undergo DNA rearrangement.
IGK
(kappa/Lambda)
V-Region
IGKV
76 Genes
J-Region
IGKJ
5 Genes
C-region
IGKC
1 Gene

5. V-J rearrangements occur at
the recombination signals (RS),
which include a heptameric
sequence (7nucleotides) and a
nonameric sequence (9
nucleotides), separated by a
spacer.
Each IGKVgene is followed
downstream (in the 3' position)
by an RS consisting of a
CACAGTG heptamer, and then
by a 12-bp spacer, and then an
ACAAAAACC nonamer.
Each IGKJgene is preceded
upstream (in the 5' position) by
an RS consisting, between 5'
and 3', of a GGTTTTTGT
nonamer, a 23-bp spacer and a
CACTGTG heptamer
Nonameric
sequence
ACAAAAACC
Heptameric
sequence
CACAGTAG
Spacer (12bp)
Nonameric
sequence
GGTTTTTGT
Heptameric
sequence
CACTGTG
Spacer (23bp)

6. 1-Light chains (Lambda)
1.2.Lambdachain:V-Jrearrangements
IGL(lambda)genesatthe22q11positionon
chromosome22;
theV-Jrearrangementmechanismisthe
sameasthatdescribedfortheIGKgenes:the
rearrangementstakeplacebetweenoneofthe
29to33functionalIGLVgenesandaJgene;
itshouldbenotedthatthereare4to5
functionalIGLCgenes,eachofwhichis
precededbyaIGLJgene.
1.3.Alleleexclusionandisotype
Alleleexclusioncanbeexplainedinpartby
thetimingofrearrangements,andpartlyby
thesurfaceexpressionofafunctional
immunoglobulin,whichinhibitsthe
rearrangementsandthereforetheexpression
ofasecondchain.Onlyone14chromosome
andone2(or22)chromosomearetherefore
productive

7. RecombinationsignalsequencesareconservedsequencesofnoncodingDNAthat
arerecognizedbytheRAG1/RAG2enzymecomplexduringV(D)Jrecombinationin
immatureBcellsandTcells.Recombinationsignalsequencesguidetheenzyme
complextotheV,D,andJgenesegmentsthatwillundergorecombinationduringthe
formationoftheheavyandlight-chainvariableregionsinT-cell
receptorsandimmunoglobulinmolecules
RSSsaremadeupofhighlyconservedheptamer sequences (7base
pairs),spacersequences,andconservednonamersequences(9basepairs)thatare
adjacenttotheV,DandJsequencesintheheavy-chainregionofDNAandtheVandJ
sequencesinthelight-chainDNAregion.Spacersequencesarelocatedbetweenheptamerand
nonamersequencesandexhibitbasepairvarietybutarealwayseither12basepairsor23base
pairslong.HeptamersequencesareusuallyCACAGTGandnonamersare
usuallyACAAAAACC.ThenucleotidesinREDaremorehighlyconserved.TheRAG1/RAG2
enzymecomplexfollowsthe12-23rulewhenjoiningV,D,andJsegments,pairing12-bp
spacerRSSsto23-bpspacerRSSs.Thispreventstwodifferentgenescodingforthesame
regionfromrecombining(ex.V-Vrecombination).RSSsarelocatedbetweenV,D,andJ
segmentsofthegerm-lineDNAofmaturingBandTlymphocytesandarepermanentlyspliced
outofthefinalIgmRNAproductafterV(D)Jrecombinationiscomplete.

8. TheRAG1/RAG2enzymecomplexrecognizestheheptamer
sequencesflankingtheVandJcodingregionsandnickstheir5'
end,releasingtheinterveningDNAbetweentheVandJcoding
regions.Intheheavy-chaincodingregionofDNA,theRAG1/RAG2
enzymecomplexrecognizestheRSSsflankingtheDandJ
segmentsandbringsthemtogether,formingaloopcontaining
interveningDNA.TheRAG1/RAG2complexthenintroducesanick
atthe5'endoftheRSSheptamersadjacenttothecodingregions
onboththeDandJsegments,permanentlyremovingtheloopof
interveningDNAandcreatingadouble-strandedbreakthatis
repairedbyVDJrecombinaseenzymes.Thisprocessisrepeatedfor
thejoiningofVtoDJ.Inlight-chainrearrangement,onlyVandJ
segmentsarebroughttogether.

9. •IGH('heavy')genesat14q32onchromosome14.
•Thereare11IGHCgenes,9ofwhicharefunctional(IGHM,IGHD,IGHG1,IGHG2,IGHG3,IGHG4,IGHA1,
IGHA2andIGHE)andcorrespondrespectivelyto9heavychainisotypesm,d,g1,g2,g3,g4,a1,a2ande.
•DNArearrangementsbetweenoneofthe38to46functionalvariableIGHVgenes,oneofthe23functional
diversityIGHDgenes,andoneofthe6functionaljunctionIGHJgenes:therearealsosomeRSs,whichare
locateddownstream(inposition3')oftheVgenes,eithersideoftheDgenesandupstream(at5')oftheJgenes.
DuringV-D-Jrearrangement,ajunctionisfirstformedbetween1Dand1J,andthenonebetween1Vandthe
D-Jcomplex.
2-Heavy chains
Note:therearealso2or3openreadingframes
fortheDgenes;eachofwhichcancodefor2or3
differentpeptidesequences.TheV-D-Jjunctions
arealsocharacterizedbynucleotidedeletions(by
anexonuclease)andbytherandomadditionof
nucleotides(bymeansofTdT,terminal
deoxynucleotidyltransferase);theVregions
whichresultarenot,therefore,codedinthe
genomeoftheindividualandconsiderably
increasethediversityoftheV-D-Jjunctionsof
thevariabledomainsoftheheavychainsofthe
immunoglobulins.

10. 2.2.Isotypeswitching
•Inthepre-Blymphocyte,amuchainisfirstsynthesized,becausetheconstantIGHMgene(C)islocatedneartotheV-D-Jrearrangement.
Thismuchainisassociatedwiththepseudo-lightchainandthecombinationconstitutesthepre-Breceptor.ThefirstcompleteIg
synthesizedbytheB-lymphocyteisanIgM,inwhichthemuchainiscombinedwithalightkappaorlambdachain.
•Duringitsdifferentiation,theBlymphocytecanexpresssomeotherisotypeorsub-isotypeofIg.Thisinvolvesthereplacementofan
IGHCgenebyanother,astheresultofDNArecombination(isotypeswitch),withtheexcisionoftheentireintermediatepartofadeletion
loop.Thisexcisionoccursattheswitchsequences(rolerelatedtothatoftheRSs).
•Theusualsequenceisthenasfollows:synthesisofpre-messengerRNA,splicingoftheintrons,resultinginmatureRNA,andthenprotein
synthesis.
•Thisexplainswhy1)aB-lymphocytecanatfirstsynthesizeanIgMandthen,duringitsdifferentiation,anIgG(IgG1,IgG2,IgG3or
IgG4),anIgA(IgA1orIgA2)oranIgE,and2)thatitretainsthesameV-D-Jrearrangementandthereforethesameantigenrecognition
site(idiotype)

11. •Alternativesplicingofthepre-messengerRNAoftheheavychaincanyieldeithera
membraneheavychain(membraneIgofBlymphocytes),orasecretedheavychain
(plasmocytesecretedIg),whichretainthesameV-D-Jrearrangement(idiotype)andthe
sameconstantregion(isotype).
•Note:thesamemechanism(alternativesplicingofapre-messenger)expressestheIgMsand
IgDsinthesameBcell(situationinmatureBcellsleavingthebonemarrowandreachthe
lymphnodesviathecirculation).
3. Membrane and secreted Igs

12. 1.Germlinediversity:multigenefamilies
•'Germline'diversitydependsonthenumberofgenesateachlocus.Thesearefamiliesofgenes,offeringthepossibility
ofachoicebetweensimilar?functionalsequences.Possibleintergenerecombinationspermitthelong-termevolutionof
thelocuswithduplicationordeletionofthegenes.
•Thesegenesundergointrageneconversionsandrecombinations,leadingtomixinganddiversity(polymorphism)
betweenindividuals.
•Thepresenceofseveralopenreadingframes,inthecaseofIGHDgenes,furtherincreasesthepossibilityofchoice
betweensimilarfunctionalsequences.
2.DiversityduetoDNArearrangements
•Combinationdiversity-inthemathematicalsenseoftheterm-permitsthepotentialsynthesisofamillion
immunoglobulins.TheIGHgenespermitthesynthesisofabout6000heavychains,theIGKorIGLgenesofabout160
lightchains,whichisequivalenttoaboutamillionpossiblecombinations6x10
3
x160).
Inadditiontothis,duringtherearrangementsof
theIGHoftheheavychains,theacquisitionof
theNregions,andusingoneorotherofthe
readingframesfortheDgenesattheV-D-J
junctions,andduringtheIGKorIGL
rearrangementsofthelightchains,flexibilityof
theV-Jjunctions.Thesemechanismscontribute
toincreasingthediversitybyafactor10
3
to
10
4
(potentialsynthesisof10
9
Igchains).
Conclusions

13. 3.Diversityasaresultofsomatichypermutations
•Finally,somaticmutationsareextremelynumerous(somatichypermutations)
andproduceverytargetedcharacterizationoftherearrangedV-JandV-D-J
genesoftheIg,buttheirmechanismofonsetisnotyetknown.AID
(activation-inducedcytidinedeaminase)maybeimplicatedbothinthe
occurrenceofthemutationsandtheswitchmechanism.Themutations
appearduringthedifferentiationoftheBlymphocyteinthelymphglands
andcontributetoincreasingthediversityoftheIgsbyafurtherfactorof10
3
,
whichmakesitpossibletoachieveapotentialdiversityof10
12
differentIgs.
•Thesedifferentmechanismsofdiversitymakeitpossibletoobtain
10
12
differentimmunoglobulins,capableofrespondingtotheseveralmillion
knownantigens.
•ThenumberofdifferentIgsisinfactlimitedbythenumberofBcellsina
givenspecies.

14. http://atlasgeneticsoncology.org/Educ/PolyIgEng.html