Reduced isotype switching in splenic B cells from mice deficient in mismatch repair enzymes.
Mice deficient in various mismatch repair (MMR) enzymes were examined to determine whether this repair pathway is involved in antibody class switch recombination. Splenic B cells from mice deficient in Msh2, Mlh1, Pms2, or Mlh1 and Pms2 were stimulated in culture with lipopolysaccharide (LPS) to induce immunoglobulin (Ig)G2b and IgG3, LPS and interleukin (IL)-4 to induce IgG1, or LPS, anti-delta-dextran, IL-4, IL-5, and transforming growth factor (TGF)-beta1 to induce IgA. After 4 d in culture, cells were surface stained for IgM and non-IgM isotypes and analyzed by FACS((R)). B cells from MMR-deficient mice show a 35-75% reduction in isotype switching, depending on the isotype and on the particular MMR enzyme missing. IgG2b is the most affected, reduced by 75% in Mlh1-deficient animals. The switching defect is not due to a lack of maturation of the B cells, as purified IgM(+)IgD(+) B cells show the same reduction. MMR deficiency had no effect on cell proliferation, viability, or apoptosis, as detected by [(3)H]thymidine incorporation and by propidium iodide staining. The reduction in isotype switching was demonstrated to be at the level of DNA recombination by digestion-circularization polymerase chain reaction (DC-PCR). A model of the potential role for MMR enzymes in class switch recombination is presented.
Schrader
C E
CE
Department of Molecular Genetics and Microbiology, Program in Immunology and Virology, University of Massachusetts Medical School, Worcester 01655-0122, USA. carol.schrader@banyan.ummed.edu
Edelmann
W
W
Kucherlapati
R
R
Stavnezer
J
J
D048868:Adaptor Proteins, Signal Transducing; D000251:Adenosine Triphosphatases; D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism*; D020137:Base Pair Mismatch / Q000276:immunology*; D002352:Carrier Proteins; D002453:Cell Cycle / Q000235:genetics / Q000276:immunology; D002455:Cell Division / Q000235:genetics / Q000276:immunology; D002470:Cell Survival / Q000235:genetics / Q000276:immunology; D002478:Cells, Cultured; D004260:DNA Repair / Q000276:immunology*; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins; D005434:Flow Cytometry; D017578:Immunoglobulin Class Switching / Q000235:genetics*; D007132:Immunoglobulin Isotypes / Q000096:biosynthesis* / Q000235:genetics*; D051379:Mice; D000070976:Mismatch Repair Endonuclease PMS2; D000070957:MutL Protein Homolog 1; D009363:Neoplasm Proteins / Q000235:genetics / Q000276:immunology; D009687:Nuclear Proteins; D011506:Proteins / Q000235:genetics / Q000276:immunology; D013154:Spleen
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D007132:Immunoglobulin Isotypes; C501486:MLH1 protein, human; C501485:Mlh1 protein, mouse; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D011506:Proteins; D000251:Adenosine Triphosphatases; C499533:Pms2 protein, mouse; D000070976:Mismatch Repair Endonuclease PMS2; D000070957:MutL Protein Homolog 1; D045643:DNA Repair Enzymes