Molecular cloning and characterization of NF-IL3A, a transcriptional activator of the human interleukin-3 promoter Journal Article
| Authors: | Zhang, W.; Zhang, J.; Kornuc, M.; Kwan, K.; Frank, R.; Nimer, S. D. |
| Article Title: | Molecular cloning and characterization of NF-IL3A, a transcriptional activator of the human interleukin-3 promoter |
| Abstract: | To isolate transcription factors important in the regulation of the human interleukin-3 (IL-3) gene, we screened a λgt11 cDNA library, constructed from phytohemagglutinin-stimulated human T-cell RNA, with a probe containing regulatory sequences in the upstream region of the IL-3 gene (located from bp -165 to -128 and referred to as the DNase I footprint A region). We isolated a 0.96-kb cDNA that encoded a basic amino acid domain and a leucine zipper domain and used the 'rapid amplification and cloning of 3' ends' technique to isolate the 3' half of the cDNA clone, generating a 1.9-kb full-length cDNA clone. Using in vitro-translated protein, which we call NF-IL3A, we defined the IL-3 promoter sequences bound by NF-IL3A in DNase 1 footprinting assays as TAATTACGTCTG and, using gel shift assays, defined ATTACG as the minimal sequence required for binding of NF-IL3A in vitro. Proteins that bind to the NF-IL3A binding site are found in both unstimulated and stimulated T-cell lines in similar amounts, although the level of NF-IL3A mRNA increases after T-cell activation in several mature T-cell lines. The NF-IL3A protein is nearly identical to a recently identified transcriptional repressor protein, E4BP4, and NF-IL3A binds specifically to regulatory sequences in both the adenovirus E4 promoter and the human gamma interferon promoter. Cotransfection experiments demonstrate that introduction of an expression vector containing the NF-IL3A cDNA into resting T cells transactivates IL-3 promoter-chloramphenicol acetyltransferase gene plasmids that contain the A region; this effect requires the presence of an intact NF-IL3A binding site. One or more copies of the A region also confer NF-IL3A responsiveness on a heterologous promoter in T cells. NF-IL3A appears to play an important role in the expression of IL-3 by T cells. |
| Keywords: | controlled study; human cell; promoter region; transcription initiation; transcription factor; molecular cloning; amino acid sequence; gamma interferon; nucleotide sequence; repressor protein; t lymphocyte activation; electrophoretic mobility; complementary dna; adenoviridae; phorbol 13 acetate 12 myristate; dna footprinting; phytohemagglutinin; deoxyribonuclease i; dna transfection; human; priority journal; article; chloramphenicol acetyltransferase |
| Journal Title: | Molecular and Cellular Biology |
| Volume: | 15 |
| Issue: | 11 |
| ISSN: | 0270-7306 |
| Publisher: | American Society for Microbiology |
| Date Published: | 1995-11-01 |
| Start Page: | 6055 |
| End Page: | 6063 |
| Language: | English |
| DOI: | 10.1128/mcb.15.11.6055 |
| PUBMED: | 7565758 |
| PROVIDER: | scopus |
| PMCID: | PMC230857 |
| DOI/URL: | |
| Notes: | Article -- Export Date: 28 August 2018 -- Source: Scopus |
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