Mechanism of rsv induced il-8 gene expression in a549 cells prior
to viral replication.
Fiedler, Michael A., Kara Wernke-Dollries, and James M. Stark.
Division of Pulmonary Medicine, Department of Pediatrics,
Children's Hospital Research Foundation and University of Cincinnati,
College of Medicine, Cincinnati, Ohio, 45229-3039
APStracts 3:0140L, 1996.
Previous studies demonstrated that respiratory syncytial virus (RSV)
infection of A549 cells induced interleukin-8 (IL-8) gene expression
and protein release from the cells, as early as 2 hours after
treatment (12,16). Furthermore, the effects of RSV at the 2 hour time
point were not dependent on viral replication. The studies reported
here were designed to test the hypothesis that active and inactive
RSV induce IL-8 gene expression in A549 cells at the two hour time
point by a mechanism dependent on the activation of the nuclear
transcription factor NF-kappa B (NF-MB). Northern blot analysis
indicated that IL-8 gene expression occurred independent of protein
synthesis two hours after A549 cells were treated with RSV. Analysis
of nuclear extracts from RSV treated A549 cells by electrophoretic
mobility shift assays (EMSA) demonstrated that NF-MB was activated as
early as 15 minutes after RSV was added to the cells, and remained
activated for at least 90 minutes. In contrast, baseline levels of
NF-IL-6 and AP-1 did not change over this period of time. DNase
footprint analysis of a portion of the 5' flanking region of the IL-8
gene demonstrated two potential regions for transcription factor
binding, which corresponded to the potential AP-1 binding site, and
potential NF-IL-6 and NF-M-B binding sites. Mutational analysis of
the 200 base pair 5' untranslated region of the IL-8 gene
demonstrated that activation of NF-MB and NF-IL-6 were required for
RSV induced transcriptional activation of the IL-8 gene.
Received 10 October 1995; accepted in final form 15 August 1996.
APS Manuscript Number L300-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 29 August 1996