Molecular dissection of the mouse interleukin-4 promoter.
Understanding the molecular mechanisms regulating the expression of interleukin 4 (IL-4) may shed light on the differentiation of lymphokine-producing phenotypes of CD4+ T cells. We have identified two DNA segments that are necessary for full phorbol 12-myristate 13-acetate (PMA)-induced activity of the IL-4 promoter region in the thymoma cell line EL4. Through deletion and mutation analyses, one of these segments (-57 through -47) was shown to be indispensable for promoter function. We designated this sequence consensus sequence 1 (CS1), as it shares homology with a sequence (ATTTTCCNNTG) that appears five times in the proximal 302-base-pair (bp) region 5' of the gene. We examined CS1 in further detail, as well as a second consensus sequence, CS2, located at nucleotides -75 through -65; both are within a minimal 83-bp construct that expresses full promoter activity. CS1- and CS2-spanning oligonucleotides bound apparently distinct PMA-inducible, sequence-specific factors in mobility-shift assays. Multimer constructs linking CS1- or CS2-spanning oligonucleotides to a heterologous promotor revealed that the CS1 construct had the greater enhancer activity in EL4 cells. Mutating the CS1 sequence within the context of the 302-bp promoter abolished all activity of the promoter, while mutating the CS2 sequence alone had little effect. Furthermore, a CS1 multimer could drive a heterologous promoter in an IL-4-producing [helper T-cell type 2 (TH2-type)] T-cell clone but not in a non-IL-4-producing (TH1-type) clone, suggesting a mechanism by which IL-4 production could be differentially regulated in TH subsets.
Bruhn
K W
KW
Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892.
Nelms
K
K
Boulay
J L
JL
Paul
W E
WE
Lenardo
M J
MJ
D000818:Animals; D001483:Base Sequence; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D007376:Interleukin-2 / Q000235:genetics; D015847:Interleukin-4 / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D015345:Oligonucleotide Probes / Q000737:chemistry; D011401:Promoter Regions, Genetic; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D016176:T-Lymphocyte Subsets / Q000502:physiology*; D006377:T-Lymphocytes, Helper-Inducer / Q000502:physiology*
D004268:DNA-Binding Proteins; D007376:Interleukin-2; D009687:Nuclear Proteins; D015345:Oligonucleotide Probes; D015847:Interleukin-4