S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process

Andrea Bottaro, Rusty Lansford, Lixing Xu, Jue Zhang, Paul Rothman, Frederick W. Alt

Research output: Contribution to journalArticle

180 Citations (Scopus)

Abstract

Stimulation of B lymphocytes with a combination of lipopolysaccharide (LPS) and interleukin-4 ((IL-4)) induces germline transcription of and subsequent switching to the ε heavy chain constant region (Cε) gene. Mature germline Cε transcripts contain a non-coding exon (Iε exon) spliced to the Cε exons. To distinguish between the potential roles of germline transcription and those of germline transcripts in regulating the class switch process, we replaced the LPS- and IL-4-inducible Iε promoter and exon in ES cells with an LPS-inducible Eμ enhancer/V(H) promoter expression cassette. Wildtype, heterozygous or homozygous mutant ES cells were injected into RAG-2 deficient blastocysts to generate somatic chimeras in which all B cells derived from ES cells. In contrast to normal B cells, heterozygous and homozygous mutant B cells had substantial transcription through the ε switch recombination region (Sε) following treatment with LPS alone and, under these conditions, both underwent low level switching (10- to 100-fold less than wildtype cells stimulated with LPS+IL-4) to IgE production. Heterozygous mutant cells underwent switching to IgE at essentially wildtype levels when stimulated with LPS and IL-4. However, homozyous mutant cells still showed extremely low levels of switching to IgE upon LPS and IL-4 stimulation. Analyses of hybridomas from heterozygous mutants indicated that the mutation is cis-acting and normal switching to other isotypes indicated that it is specific for IgE. Thus transcription per se generates low levels of class switch recombination in the absence of I region sequences. However, we demonstrate for the first time that, for optimal efficiency, the process requires the presence of the intact I region and/or I region promoter in cis, implicating factors beyond transcription through the S region in the regulation of class switching.

Original languageEnglish (US)
Pages (from-to)665-674
Number of pages10
JournalEMBO Journal
Volume13
Issue number3
StatePublished - Feb 1 1994
Externally publishedYes

Fingerprint

Transcription
Immunoglobulin E
Genetic Recombination
Lipopolysaccharides
Switches
Interleukin-4
Exons
B-Lymphocytes
Cells
Immunoglobulin Class Switching
Lymphocytes
Hybridomas
Blastocyst
Genetic Promoter Regions
Transcription Factors
Genes
Mutation

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Bottaro, Andrea ; Lansford, Rusty ; Xu, Lixing ; Zhang, Jue ; Rothman, Paul ; Alt, Frederick W. / S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process. In: EMBO Journal. 1994 ; Vol. 13, No. 3. pp. 665-674.
@article{ef556400278348279d89c7ce17d71f9c,
title = "S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process",
abstract = "Stimulation of B lymphocytes with a combination of lipopolysaccharide (LPS) and interleukin-4 ((IL-4)) induces germline transcription of and subsequent switching to the ε heavy chain constant region (Cε) gene. Mature germline Cε transcripts contain a non-coding exon (Iε exon) spliced to the Cε exons. To distinguish between the potential roles of germline transcription and those of germline transcripts in regulating the class switch process, we replaced the LPS- and IL-4-inducible Iε promoter and exon in ES cells with an LPS-inducible Eμ enhancer/V(H) promoter expression cassette. Wildtype, heterozygous or homozygous mutant ES cells were injected into RAG-2 deficient blastocysts to generate somatic chimeras in which all B cells derived from ES cells. In contrast to normal B cells, heterozygous and homozygous mutant B cells had substantial transcription through the ε switch recombination region (Sε) following treatment with LPS alone and, under these conditions, both underwent low level switching (10- to 100-fold less than wildtype cells stimulated with LPS+IL-4) to IgE production. Heterozygous mutant cells underwent switching to IgE at essentially wildtype levels when stimulated with LPS and IL-4. However, homozyous mutant cells still showed extremely low levels of switching to IgE upon LPS and IL-4 stimulation. Analyses of hybridomas from heterozygous mutants indicated that the mutation is cis-acting and normal switching to other isotypes indicated that it is specific for IgE. Thus transcription per se generates low levels of class switch recombination in the absence of I region sequences. However, we demonstrate for the first time that, for optimal efficiency, the process requires the presence of the intact I region and/or I region promoter in cis, implicating factors beyond transcription through the S region in the regulation of class switching.",
author = "Andrea Bottaro and Rusty Lansford and Lixing Xu and Jue Zhang and Paul Rothman and Alt, {Frederick W.}",
year = "1994",
month = "2",
day = "1",
language = "English (US)",
volume = "13",
pages = "665--674",
journal = "EMBO Journal",
issn = "0261-4189",
publisher = "Nature Publishing Group",
number = "3",

}

S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process. / Bottaro, Andrea; Lansford, Rusty; Xu, Lixing; Zhang, Jue; Rothman, Paul; Alt, Frederick W.

In: EMBO Journal, Vol. 13, No. 3, 01.02.1994, p. 665-674.

Research output: Contribution to journalArticle

TY - JOUR

T1 - S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process

AU - Bottaro, Andrea

AU - Lansford, Rusty

AU - Xu, Lixing

AU - Zhang, Jue

AU - Rothman, Paul

AU - Alt, Frederick W.

PY - 1994/2/1

Y1 - 1994/2/1

N2 - Stimulation of B lymphocytes with a combination of lipopolysaccharide (LPS) and interleukin-4 ((IL-4)) induces germline transcription of and subsequent switching to the ε heavy chain constant region (Cε) gene. Mature germline Cε transcripts contain a non-coding exon (Iε exon) spliced to the Cε exons. To distinguish between the potential roles of germline transcription and those of germline transcripts in regulating the class switch process, we replaced the LPS- and IL-4-inducible Iε promoter and exon in ES cells with an LPS-inducible Eμ enhancer/V(H) promoter expression cassette. Wildtype, heterozygous or homozygous mutant ES cells were injected into RAG-2 deficient blastocysts to generate somatic chimeras in which all B cells derived from ES cells. In contrast to normal B cells, heterozygous and homozygous mutant B cells had substantial transcription through the ε switch recombination region (Sε) following treatment with LPS alone and, under these conditions, both underwent low level switching (10- to 100-fold less than wildtype cells stimulated with LPS+IL-4) to IgE production. Heterozygous mutant cells underwent switching to IgE at essentially wildtype levels when stimulated with LPS and IL-4. However, homozyous mutant cells still showed extremely low levels of switching to IgE upon LPS and IL-4 stimulation. Analyses of hybridomas from heterozygous mutants indicated that the mutation is cis-acting and normal switching to other isotypes indicated that it is specific for IgE. Thus transcription per se generates low levels of class switch recombination in the absence of I region sequences. However, we demonstrate for the first time that, for optimal efficiency, the process requires the presence of the intact I region and/or I region promoter in cis, implicating factors beyond transcription through the S region in the regulation of class switching.

AB - Stimulation of B lymphocytes with a combination of lipopolysaccharide (LPS) and interleukin-4 ((IL-4)) induces germline transcription of and subsequent switching to the ε heavy chain constant region (Cε) gene. Mature germline Cε transcripts contain a non-coding exon (Iε exon) spliced to the Cε exons. To distinguish between the potential roles of germline transcription and those of germline transcripts in regulating the class switch process, we replaced the LPS- and IL-4-inducible Iε promoter and exon in ES cells with an LPS-inducible Eμ enhancer/V(H) promoter expression cassette. Wildtype, heterozygous or homozygous mutant ES cells were injected into RAG-2 deficient blastocysts to generate somatic chimeras in which all B cells derived from ES cells. In contrast to normal B cells, heterozygous and homozygous mutant B cells had substantial transcription through the ε switch recombination region (Sε) following treatment with LPS alone and, under these conditions, both underwent low level switching (10- to 100-fold less than wildtype cells stimulated with LPS+IL-4) to IgE production. Heterozygous mutant cells underwent switching to IgE at essentially wildtype levels when stimulated with LPS and IL-4. However, homozyous mutant cells still showed extremely low levels of switching to IgE upon LPS and IL-4 stimulation. Analyses of hybridomas from heterozygous mutants indicated that the mutation is cis-acting and normal switching to other isotypes indicated that it is specific for IgE. Thus transcription per se generates low levels of class switch recombination in the absence of I region sequences. However, we demonstrate for the first time that, for optimal efficiency, the process requires the presence of the intact I region and/or I region promoter in cis, implicating factors beyond transcription through the S region in the regulation of class switching.

UR - http://www.scopus.com/inward/record.url?scp=0027952630&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027952630&partnerID=8YFLogxK

M3 - Article

C2 - 8313911

AN - SCOPUS:0027952630

VL - 13

SP - 665

EP - 674

JO - EMBO Journal

JF - EMBO Journal

SN - 0261-4189

IS - 3

ER -