Interleukin-8 gene regulation in intestinal epithelial cells infected with rotavirus: role of viral-induced IkappaB kinase activation.
Rotavirus is the major etiologic agent of diarrhea in children and the most common cause of severe pediatric gastroenteritis. Rotavirus infection is limited to mature enterocytes that line the villi of the small intestine. Gut epithelial cells, upon infection and cytokine stimulation, are able to produce chemokines, a family of small chemotactic cytokines that regulate the migration and activation of leukocytes. We have previously shown that rotavirus infection of the intestinal epithelial cell line HT-29 induces increased expression of the CXC chemokine interleukin- (IL) 8. Mechanisms responsible for the transcriptional regulation of the IL-8 gene in intestinal epithelial cells during viral infections have not been fully elucidated. Therefore, the purpose of this study was to define the molecular mechanisms of IL-8 gene expression in HT-29 cells infected with rotavirus. Transient transfection analysis of 5' deletions and mutations of the IL-8 promoter driving expression of luciferase reporter gene indicates that the activating protein- (AP) 1 and nuclear factor- (NF) kappaB elements are necessary for IL-8 promoter activation during rotavirus infection. The importance of NF-kappaB activation for IL-8 gene expression was further demonstrated by the inhibition of rotavirus-induced IL-8 gene transcription and protein synthesis following blockade of degradation of the NF-kappaB cytoplasmic inhibitor IkappaB-alpha. Rotavirus infection of HT-29-induced IkappaB kinase (IKK) activation and overexpression of a dominant negative mutant of IKK-beta greatly reduced rotavirus-induced IL-8 promoter activation and NF-kappaB-driven transcription, indicating that IKK is involved in rotavirus-induced IL-8 gene expression and NF-kappaB activation.
Casola
Antonella
A
Department of Pediatrics, University of Texas Medical Branch, Galveston 77555-0366, USA. ancasola@utmb.edu
Garofalo
Roberto P
RP
Crawford
Sue E
SE
Estes
Mary K
MK
Mercurio
Frank
F
Crowe
Sheila E
SE
Brasier
Allan R
AR
D004847:Epithelial Cells / Q000276:immunology / Q000821:virology*; D005786:Gene Expression Regulation; D006801:Humans; D051550:I-kappa B Kinase; D016209:Interleukin-8 / Q000235:genetics* / Q000378:metabolism; D007422:Intestines / Q000821:virology*; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D012401:Rotavirus / Q000276:immunology*; D018808:Transcription Factor AP-1 / Q000378:metabolism
D016209:Interleukin-8; D016328:NF-kappa B; D018808:Transcription Factor AP-1; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human