Deletional switch recombination occurs in interleukin-4-induced isotype switching to IgE expression by human B cells.

Abstract

There is controversy as to whether deletional rearrangement occurs between the IgM and IgE switch regions (S, and S,, respectively) during switching to the IgE isotype.We have addressed the issue by stimulating normal human B cells, sorted for lack of expression of surface IgE, to produce IgE by infection with Epstein-Barr virus (EBV) in the presence of interleukin 4 (IL-4). Genomic DNA was amplified for Sp/Se switch junction fragments by utilizing the nested-primer polymerase chain reaction. Switch junction fragments were amplified from B cells infected with EBV in the presence of IL-4 but not from B cells infected with EBV alone. The DNA sequence of these "switch fragments" revealed direct joining of Si to Se in each case. The recombination sites within SP were clustered within 900 base pairs at the 5' end of the switch region, suggesting that there are "hot spots" for recombination within S.,. The S, recombination sites were scattered throughout the Se region. These findings indicate that IL4-induced isotype switching to IgE production in human B cells is accompanied by DNA rearrangements with joining of S., to Se.

Antibodies specific for a segment of human membrane IgE deplete IgE-producing B cells in humanized mice

“…The regulation and existence of IgE memory B cells is controversial and poorly understood (50)(51)(52)(53)(54)(55)(56). It has been suggested that a significant fraction of IgE memory resides in the IgG1 compartment, with generation of a memory IgE response resulting from sequential switching of IgG1 memory B cells to the IgE B cell lineage (50,54,(57)(58)(59)(60)(61)(62)(63)(64). While our studies have not directly addressed this issue, it is interesting that we and others have observed that both primary and memory IgE responses occur prior to IgG1 responses in our immunization and asthma models, perhaps indicating that the vast majority of IgE responses do not develop by a sequential switching mechanism through an IgG1 B cell intermediate.…”

Molecular analysis of the induction of immunoglobulin E synthesis in human B cells by interleukin 4 and engagement of CD40 antigen.

“…We have previously shown that nested primer PCR amplifies switch fragments from as little as 10 pg (one diploid genome) of DNA from the IgE plasmacytoma U266 (3). If all the B cells containing cytoplasmic IgE in the CD40/IL,4 system (=10% of the total B cells) had undergone deletional switch recombination, switch fragments should have been detected in 100 pg DNA samples.…”

“…Highly purified human peripheral blood B cells from nonatopic subjects were isolated and cultured as previously described (1). slgE-B cells were obtained by cell sorting (3). Resting B cells were collected at the 50-60% interface after Percoll gradient centrifugation (4).…”

“…Nested PCK runs for switch (S)~/Se switch fragments were performed on high molecular weight DNA isolated from B cells as described (3). Four PCK primers were used (see rig.…”

“…We show herein that engagement of CD40 significantly enhanced the level of IL-4-induced e germline transcripts (GLT) 1 and resulted in the production of mature Ce transcripts. A recendy established nested primer PCR approach (3) showed that ddetional switch recombination underlies IgE class switching in the CD40/11,4 system.…”