Abstract:
:Alpha-defensins are antimicrobial peptides that contribute to innate-immune functions of neutrophils and intestinal Paneth cells. Transcription of alpha-defensin genes occurs early in neutrophilic myelopoeisis. To examine the mechanisms that regulate alpha-defensin gene expression, we analyzed transcription of rat neutrophil alpha-defensin NP-3 in D4 cells, a subclone of the promyelocytic cell line IPC-81. Northern blot analysis showed that D4 cells express fivefold higher levels of alpha-defensin mRNA than the parental cell line in a manner relatively independent of passage number. Increased levels of steady-state mRNA in D4 cells correlated with markedly elevated peptide levels detected by immunocytochemical staining. To identify the cis-acting DNA elements involved in tissue-specific expression, D4 cells were transfected with luciferase reporter constructs containing NP-3 gene 5'-flanking sequences. Analyses of transfected D4 cells demonstrated that the proximal 87 base pair (bp) sequence contained cis-acting DNA elements necessary for optimal promoter activity. Mutational analyses within the 87-bp region suggested the involvement of the CAAT box and a putative polyoma enhancer-binding protein 2/core-binding factor (PEBP2/CBF) site in defensin gene transcription. Transient transfection analyses using tandem repeats of oligonucleotides containing these sequences demonstrated that proximity of the CAAT box and PEBP2/CBF site was important for defensin promoter activity. Electrophoretic mobility shift assays indicated that PEBP2/CBF or a PEBP2/CBF-related protein was involved in a specific protein-DNA interaction occurring within a DNA fragment containing the CAAT and PEBP2/CBF sequences. These data identify functional trans- and cis-elements that regulate rat defensin gene expression in high defensin-expressing promyelocytic cells.
journal_name
J Leukoc Bioljournal_title
Journal of leukocyte biologyauthors
Yamamoto CM,Banaiee N,Yount NY,Patel B,Selsted MEdoi
10.1189/jlb.0803384subject
Has Abstractpub_date
2004-02-01 00:00:00pages
332-41issue
2eissn
0741-5400issn
1938-3673pii
jlb.0803384journal_volume
75pub_type
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