Gene conversion as a part of BCR diversification

ematsen · February 21, 2016, 5:11pm

A new paper shows that gene conversion from a different chromosome is another means of antibody diversification.

This could either happen through RAG or through double-stranded breaks made by AID at a later stage of development.

Here’s the paper:

ncbi.nlm.nih.gov

A LAIR1 insertion generates broadly reactive antibodies against malaria variant antigens.

J Tan, K Pieper, L Piccoli, A Abdi, MF Perez, R Geiger, CM Tully, D Jarrossay, F Maina Ndungu, J Wambua, P Bejon, CS Fregni, B Fernandez-Rodriguez, S Barbieri, S Bianchi, K Marsh, V Thathy, D Corti, F Sallusto, P Bull and A Lanzavecchia, Nature, Jan 2016 07

Plasmodium falciparum antigens expressed on the surface of infected erythrocytes are important targets of naturally acquired immunity against malaria, but their high number and variability provide the pathogen with a powerful means of escape from host antibodies. Although broadly reactive antibodies against these antigens could be useful as therapeutics and in vaccine design, their identification has proven elusive. Here we report the isolation of human monoclonal antibodies that recognize erythrocytes infected by different P. falciparum isolates and opsonize these cells by binding to members of the RIFIN family. These antibodies acquired broad reactivity through a novel mechanism of insertion of a large DNA fragment between the V and DJ segments. The insert, which is both necessary and sufficient for binding to RIFINs, encodes the entire 98 amino acid collagen-binding domain of LAIR1, an immunoglobulin superfamily inhibitory receptor encoded on chromosome 19. In each of the two donors studied, the antibodies are produced by a single expanded B-cell clone and carry distinct somatic mutations in the LAIR1 domain that abolish binding to collagen and increase binding to infected erythrocytes. These findings illustrate, with a biologically relevant example, a novel mechanism of antibody diversification by interchromosomal DNA transposition and demonstrate the existence of conserved epitopes that may be suitable candidates for the development of a malaria vaccine.

Here’s a commentary:

Apparently AID-mediated gene conversion is observed in chickens and fish, as reviewed in this paper:

ncbi.nlm.nih.gov

Molecular mechanisms of antibody somatic hypermutation.

JM Di Noia and MS Neuberger, Annual review of biochemistry, 2007

Functional antibody genes are assembled by V-D-J joining and then diversified by somatic hypermutation. This hypermutation results from stepwise incorporation of single nucleotide substitutions into the V gene, underpinning much of antibody diversity and affinity maturation. Hypermutation is triggered by activation-induced deaminase (AID), an enzyme which catalyzes targeted deamination of deoxycytidine residues in DNA. The pathways used for processing the AID-generated U:G lesions determine the variety of base substitutions observed during somatic hypermutation. Thus, DNA replication across the uracil yields transition mutations at C:G pairs, whereas uracil excision by UNG uracil-DNA glycosylase creates abasic sites that can also yield transversions. Recognition of the U:G mismatch by MSH2/MSH6 triggers a mutagenic patch repair in which polymerase eta plays a major role and leads to mutations at A:T pairs. AID-triggered DNA deamination also underpins immunoglobulin variable (IgV) gene conversion, isotype class switching, and some oncogenic translocations in B cell tumors.

[As if repertoire analysis wasn’t complex enough already… sigh.]

bussec · March 7, 2016, 11:30pm

I remember that I occasionally encountered joints of V segments to genomic regions on other chromosomes when doing manually sequence analysis. However, there never seemed to be a consistent target, which was very likely caused by absence of selective pressure in the experimental setup.
In addition, I now realized that IgBLAST (at least versions 1.3.0 and 1.4.0) has a hard-coded maximum number of base pairs (bp) that it will accept in between V_end and J_start. The limit is set to 90 bp, everything above will remove the identified J segments from the hit list. There is also a limit for the distance between V_end and D_start, however it seems to be different between the stand-alone version (40 bp) and the online tool (55 bp).
For our processing pipeline, sequences without any identified J segments will be excluded from subsequent analysis, thus we would effectively filter out events with long CDR3s as described in this paper. Anyone encountered similar issues?

caschramm · March 14, 2016, 9:16pm

Our pipeline also typically excludes reads with good V but no J -which is what these antibodies would look like in NGS, unless you used PacBio. Could be interesting to go back to old datasets and look…

shishi.luo · April 1, 2016, 7:09pm

In chickens and rabbits, gene conversion seems to be the primary manner in which initial BCR diversity is generated. In both these species, the IGHV locus contains only a single functional V gene and many pseudogenes. Gene conversion between the functional and pseudogenes (post VDJ rearrangement), then produces receptor diversity. The following is a nice review article about V genes in chickens and rabbits:

http://www.nature.com/nri/journal/v2/n9/full/nri889.html