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Molecular analysis of double isotype expression in IgA switching
D Y Kunimoto1, M C Sneller, L Claflin
1Department of Medical Microbiology and Infectious Diseases, University of Alberta, Edmonton, Canada.
Journal of Immunology (Baltimore, Md. : 1950)
|February 15, 1993
Summary
B cells can simultaneously produce both membrane immunoglobulin M (mIgM) and membrane immunoglobulin A (mIgA) during IgA isotype switching. This occurs without deleting the C mu gene segment, suggesting alternative splicing mechanisms are involved in B cell differentiation.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- B cell differentiation involves sequential expression of immunoglobulin (Ig) isotypes.
- The CH12.LX B cell line undergoes IgM to IgA isotype switching.
- Dual expression of mIgM and mIgA on B cells has been observed during this process.
Purpose of the Study:
- To investigate the molecular mechanisms underlying the co-expression of mIgM and mIgA in B cells.
- To determine if C mu gene segment deletion occurs during IgA isotype switching.
- To elucidate the origin of both mu and alpha mRNA transcripts in dual-bearing B cells.
Main Methods:
- Cloning of dual mIgM/mIgA-bearing B cell subpopulations.
- Primer extension analysis to identify VHDJH gene segments in mu and alpha mRNA.
- DNA restriction fragment analysis to assess C mu and C alpha gene segment association with JH.
- Northern blot analysis using VH region-specific probes to detect mu and alpha mRNA.
Main Results:
- Identical VHDJH gene segments were found in both mu and alpha mRNA.
- The C mu gene segment, but not the C alpha gene segment, was associated with the JH gene segment.
- Both mu and alpha mRNA transcripts cohybridized to the same VH region-specific probe, indicating co-transcription or alternative splicing.
Conclusions:
- Dual mIgM/mIgA expression in CH12.LX cells occurs independently of C mu gene segment deletion.
- Simultaneous production of mu and alpha mRNA transcripts suggests alternative splicing or transplicing mechanisms.
- These findings support the hypothesis that B cells can produce both mu and alpha mRNA concurrently during IgA isotype switching.