IgG2a and IgA Co-Expression by the Natural Autoantibody-producing Murine B Lymphoma T560

Abstract

The T560 B lymphoma produces polyreactive IgG2a with the features of natural autoantibody. All T560 cells bear and secrete IgG2a but a small fraction spontaneously co-express IgA. Cells secreting IgA alone cannot be detected. IgA secretion is enhanced by interaction of T560 cells either with activated T cells and cognate antigen, or with LPS, but not with cytokines, including 1L-5 and TGF-fl IgA and IgG2a mRNAs have identical VI86.2, DFL 16 and JH1 sequences from framework 2 through JH1. PCR analysis reveals that previous recombination events have led to deletion of the μ., γ3, γl, γ2b constant region genes from both the productive and the unproductive chromosome but the former has retained γ2a, and α, the latter only α. Digestion-circularization (DC)-PCR experiments provide formal proof of DNA recombination between Ca and the intron upstream of Cα. Evidently, the productive chromosome has switched only as far as γ2a, the unproductive all the way to the a constant region gene. The unproductive allele is transcriptionally active as evidenced by the presence of mRNA encoding Cal inappropriately spliced to a cryptic splice site in the downstream intron of DQ52 (eliminated from the productive chromosome). A specific RT-PCR using oligonucleotide primers derived from the upstream initiation site of the la exon and from Cal discloses that T560 cells contain a-germ line mRNA, presumably transcribed from the la-region of the productive chromosome, spliced to Cα. Treatment with LPS stops production of these spliced transcripts suggesting that it may promote either DNA recombination in cells spontaneously transcribing la or a change in splicing such that la sequence is no longer joined to Ca. Verification of the DC-PCR product by sequencing reveals that the T560 and B10.A IgA (Ig2^b allotype) hinge is different from the BALB/c IgA (Ig2^a allotype) hinge: it has two extra Cys and has eliminated the first Thr, a potential glycosylation site in BALB/c IgA.

Key Words:

• GVH
• isotype switching
• IgA
• B lymphoma