Helicobacter pylori and toll-like receptor agonists induce syndecan-4 expression in an NF-kappaB-dependent manner.
The syndecans are a family of transmembrane heparan sulfate proteoglycans (HSPG) that have been implicated in a wide variety of biological functions including the regulation of growth factor signaling, adhesion, tumorigenesis, and inflammation. In the current studies, we examined the regulation of syndecan-4 gene expression in gastric epithelial cells and macrophages in response to infection with live Helicobacter pylori and purified toll-like receptor (TLR) agonists. H. pylori, PAM3CSK4 (a TLR2 agonist), and Escherichia coli flagellin (a TLR5 agonist) all induced the rapid expression of syndecan-4 mRNA in MKN45 gastric epithelial cells. Similarly, lipopolysaccharide (LPS) (a TLR4 agonist) also induced the expression of syndecan-4 in macrophages. The H. pylori- and TLR-induced increase in syndecan-4 mRNA was blocked by the proteosome inhibitor MG-132 suggesting a role for nuclear factor kappaB (NF-kappaB) in the regulation of syndecan-4 gene expression. An 895-bp fragment of the human syndecan-4 promoter was cloned upstream of the luciferase reporter. When transfected into MKN45 cells, the activity of this promoter was inducible by H. pylori and TLR agonists. Inducible activity of the syndecan-4 promoter was blocked by cotransfection with a dominant negative IkappaBalpha expression plasmid. Electrophoretic mobility shift assays (EMSA) demonstrated the presence of a highly conserved NF-kappaB-binding site. Mutation of this site within the context of the full-length syndecan-4 promoter resulted in a complete loss of responsiveness to H. pylori and TLR agonists. These results thus demonstrate that the response of the syndecan-4 gene to infectious agents, or their products, is a direct result of NF-kappaB binding to the promoter and induction of de novo transcription.
Smith
Michael F
MF
Department of Medicine, University of Virginia Health System, Charlottesville, VA 22908-0708, USA. mfs3k@virginia.edu
Novotny
Jennifer
J
Carl
Virginia S
VS
Comeau
Laurey D
LD
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism; D005753:Gastric Mucosa / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D016480:Helicobacter pylori / Q000502:physiology*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009154:Mutation / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011509:Proteoglycans / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D053671:Syndecan-4; D051193:Toll-Like Receptors / Q000819:agonists* / Q000378:metabolism
D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D011509:Proteoglycans; D012333:RNA, Messenger; C508304:SDC4 protein, human; C508306:Sdc4 protein, mouse; D053671:Syndecan-4; D051193:Toll-Like Receptors