Spt5 accumulation at variable genes distinguishes somatic hypermutation in germinal center B cells from ex vivo-activated cells

• Published in: The Journal of experimental medicine Vol. 211; no. 11; pp. 2297 - 2306
• Main Authors: Maul, Robert W, Cao, Zheng, Venkataraman, Lakshmi, Giorgetti, Carol A, Press, Joan L, Denizot, Yves, Du, Hansen, Sen, Ranjan, Gearhart, Patricia J
• Format: Journal Article
• Language: English
• Published: United States The Rockefeller University Press 20.10.2014
• Subjects: 3' Untranslated Regions; Animals; B-Lymphocytes - immunology; B-Lymphocytes - metabolism; Chromosomal Proteins, Non-Histone - metabolism; Cytidine Deaminase - genetics; DNA Polymerase II - metabolism; DNA, Single-Stranded - metabolism; Female; Gene Order; Genetic Loci; Germinal Center - immunology; Germinal Center - metabolism; Immunoglobulin Variable Region; Lymphocyte Activation - genetics; Lymphocyte Activation - immunology; Male; Mice; Models, Biological; Sequence Deletion; Somatic Hypermutation, Immunoglobulin; Transcriptional Elongation Factors - metabolism
• ISSN: 0022-1007; 1540-9538
• DOI: 10.1084/jem.20131512

Variable (V) genes of immunoglobulins undergo somatic hypermutation by activation-induced deaminase (AID) to generate amino acid substitutions that encode antibodies with increased affinity for antigen. Hypermutation is restricted to germinal center B cells and cannot be recapitulated in ex vivo-activated splenic cells, even though the latter express high levels of AID. This suggests that there is a specific feature of antigen activation in germinal centers that recruits AID to V genes which is absent in mitogen-activated cultured cells. Using two Igh knock-in mouse models, we found that RNA polymerase II accumulates in V regions in B cells after both types of stimulation for an extended distance of 1.2 kb from the TATA box. The paused polymerases generate abundant single-strand DNA targets for AID. However, there is a distinct accumulation of the initiating form of polymerase, along with the transcription cofactor Spt5 and AID, in the V region from germinal center cells, which is totally absent in cultured cells. These data support a model where mutations are prevalent in germinal center cells, but not in ex vivo cells, because the initiating form of polymerase is retained, which affects Spt5 and AID recruitment.