Recombination of antibody genes in B cells can involve distant genomic loci and contribute a foreign antigen-binding element to form hybrid antibodies with broad reactivity for Plasmodium falciparum. So far, antibodies containing the extracellular domain of the LAIR1 and LILRB1 receptors represent unique examples of cross-chromosomal antibody diversification. Here, we devise a technique to profile non-VDJ elements from distant genes in antibody transcripts. Independent of the preexposure of donors to malaria parasites, non-VDJ inserts were detected in 80% of individuals at frequencies of 1 in 104 to 105 B cells. We detected insertions in heavy, but not in light chain or T cell receptor transcripts. We classify the insertions into four types depending on the insert origin and destination: 1) mitochondrial and 2) nuclear DNA inserts integrated at VDJ junctions; 3) inserts originating from telomere proximal genes; and 4) fragile sites incorporated between J-to-constant junctions. The latter class of inserts was exclusively found in memory and in in vitro activated B cells, while all other classes were already detected in naïve B cells. More than 10% of inserts preserved the reading frame, including transcripts with signs of antigen-driven affinity maturation. Collectively, our study unravels a mechanism of antibody diversification that is layered on the classical V(D)J and switch recombination.

Berlin Institute of Health at Charité 10117 Berlin Germany

Central European Institute of Technology Masaryk University 601 77 Brno Czech Republic

Charité Universitätsmedizin Berlin corporate member of Freie Universität Berlin and Humboldt Universität zu Berlin Charitéplatz 1 10117 Berlin Germany

Department of Biology Chemistry and Pharmacy Free University of Berlin 14195 Berlin Germany

Department of Genomics of Adaptive Immunity Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry 117997 Moscow Russian Federation

Department of Medical Parasitology and Infection Biology Swiss Tropical and Public Health Institute 4123 Allschwil Switzerland

Department of Molecular Technologies Pirogov Russian National Research Medical University 117997 Moscow Russian Federation

Institute for Research in Biomedicine Università della Svizzera Italiana Via Francesco Chiesa 5 6500 Bellinzona Switzerland

Max Delbrück Center for Molecular Medicine in the Helmholtz Association 13125 Berlin Germany

Service of Immunology and Allergy Department of Medicine Lausanne University Hospital and University of Lausanne 1011 Lausanne Switzerland

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