Germline transcription and switch recombination of a transgene containing the entire H chain constant region locus: effect of a mutation in a STAT6 binding site in the gamma 1 promoter

J Immunol. 2004 Nov 1;173(9):5531-9. doi: 10.4049/jimmunol.173.9.5531.

Abstract

The switch (S) in H chain class is preceded by germline transcription and then mediated by a DNA recombination event. One of the impediments toward understanding the mechanism is the lack of a system in which a recombinant DNA molecule undergoes cytokine-regulated class S recombination. To study class S recombination, we used transgenic mice with a 230-kb bacterial artificial chromosome that included a rearranged VDJ gene and the entire murine H chain constant region locus. We found that both germline transcription and S recombination to the transgenic gamma1 H chain gene were regulated by IL-4 like that of the endogenous genes. In mice with two or more copies of the H chain locus transgene, both germline transcripts and S recombination took place at levels comparable to those from the endogenous loci. We also prepared a version of the transgene with a 4-bp mutation in a STAT6 binding site in the gamma1 promoter region. On the average, this mutation reduced germline transcription by 80%, but did not change the amount of S recombination in vitro. Among both the wild-type and mutant transgenes, we found no significant correlation between the amount of germline transcripts and the amount of S recombination. We infer that the physiologic level of germline transcription of the gamma1 gene is in excess over the amount required for efficient S recombination.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Arsenates / immunology
  • B-Lymphocytes / immunology
  • B-Lymphocytes / metabolism
  • Binding Sites / genetics
  • Binding Sites / immunology
  • Cells, Cultured
  • Gene Rearrangement, B-Lymphocyte, Heavy Chain
  • Genetic Markers / immunology
  • Germ-Line Mutation*
  • Immunoglobulin Class Switching* / genetics*
  • Immunoglobulin Constant Regions / genetics*
  • Immunoglobulin Constant Regions / metabolism
  • Immunoglobulin Heavy Chains / genetics*
  • Immunoglobulin Heavy Chains / metabolism
  • Immunoglobulin gamma-Chains / genetics*
  • Immunoglobulin gamma-Chains / metabolism
  • Mice
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Promoter Regions, Genetic / immunology*
  • STAT6 Transcription Factor
  • Trans-Activators / genetics*
  • Trans-Activators / metabolism
  • Transcription, Genetic*
  • Transgenes* / immunology

Substances

  • Arsenates
  • Genetic Markers
  • Immunoglobulin Constant Regions
  • Immunoglobulin Heavy Chains
  • Immunoglobulin gamma-Chains
  • STAT6 Transcription Factor
  • Stat6 protein, mouse
  • Trans-Activators
  • arsenic acid