Regulation of somatic hypermutation in germinal centers

Abstract

To determine if Ig hypermutation was regulated in GCs, the genetically restricted response to phosphorylcholine (PC) was chosen as a model system. The response to PC is unusual in that different bacterial strains containing PC on their surface elicit antibodies with greatly different frequencies of mutation in their Ig genes. Immunization of BALB/c or C57BL/6 mice with PC-protein produced a normal response in terms of the number and overall appearance of GC. However, microdissection of PC-specific GC B cells and sequencing of their V{dollar}\rm \sb{lcub}H{rcub}{dollar} genes revealed a delay in the onset of somatic hypermutation, with mutations being introduced at significant frequencies only by day 16 of the primary immune response. This delay was not associated with impaired formation of GCs or serum antibody production. Indeed, PC-induced GCs form earlier than those elicited by NP-CG. Several studies were undertaken to determine if the delay in the onset of somatic hypermutation was an intrinsic factor of PC-specific B cells or was due to an altered GC microenvironment. Transfer studies into nude mice identified the CD4{dollar}\sp{lcub}+{rcub}{dollar} subset of T cells as a dose-dependent factor in the initiation of somatic hypermutation. Co-immunization with PC and NP demonstrated that delayed hypermutation is a dominant effect that acts in trans; mutation in V{dollar}\rm \sb{lcub}H{rcub}{dollar} genes activated by NP was also suppressed/delayed. Thus, B cells responding to PC alter the GC microenvironment to effect a dominance of the PC phenotype in the B cells responding to either hapten. The regulation of somatic hypermutation in vivo may be an important factor in the altered antibody responses seen in both immunodeficient and autoimmune states. This was demonstrated by using the aged mouse as a model, as immune defects in both T and B cells have been well documented in these animals. The GC B cell response is diminished in aged mice, and somatic hypermutation is virtually absent in the response to NP-protein. Taken together, these studies provided evidence for a dynamic microenvironment within GC that contributes to the regulation of Ig hypermutation. Dysregulation of the environmental cues that drive the GC reaction may play an important role in altered immune states.