Enhanced AP-1 and NF-κB activities and stability of interleukin 8 (IL-8) transcripts are implicated in IL-8 mRNA superinduction in lung epithelial H292 cells

Abstract

Inhibition of protein synthesis may result in superinduction of short-lived transcripts and has been attributed variably to stabilization of transcripts and/or increased gene transcription. Little is known about the kinetics of these processes and relevant transcriptional elements have not been identified. In this study, we describe superinduction of interleukin 8 (IL-8) mRNA, an important inflammatory mediator, in lung epithelial-like H292 cells and identify the underlying molecular mechanisms and their kinetics. Cycloheximide (CHI, 10 microg/ml), an inhibitor of protein synthesis, maximally increased IL-8 mRNA levels 30-fold in H292 cells. Tumour necrosis factor alpha (TNF-alpha), which induced IL-8 mRNA 3-fold, synergized with CHI causing a 150-fold increase at 6 h. CHI early on increased the stability of IL-8 mRNA (from 40 min in cells cultured with medium to more than 4 h with CHI). CHI also increased transcription as shown by transfection with IL-8 promoter constructs. Truncated and mutated constructs identified NF-kappaB and AP-1 binding sites as primary cis-acting elements in IL-8 gene transcription and IL-8 mRNA superinduction. Electrophoretic mobility shift assays indicated that CHI increased NF-kappaB and prolonged AP-1 DNA-binding activities and that the synergism of TNF-alpha and CHI on IL-8 mRNA expression was paralleled by a further increase of AP-1 DNA-binding activity. This synergism was still noticed when 4 h elapsed between the addition of CHI and that of TNF-alpha. Taken together, our results indicate that CHI interferes with both post-transcriptional and transcriptional repressive mechanisms of IL-8 mRNA expression.

ERK1/2 Regulates Epidermal Chemokine Expression and Skin Inflammation

“…4B) and a 2.5-fold increase in the rate of transcript decay (Fig. 4F), in keeping with previous lines of evidence (34,35). The same results were obtained with RNA polymerase II inhibition by DRB (data not shown).…”

Chlamydial infection of polarized HeLa cells induces PMN chemotaxis but the cytokine profile varies between disseminating and non-disseminating strains

“…The IL-11 and IL-6 genes, like the genes for ICAM-1, IL-8 or GM-CSF (Wertheimer et al, 1992;Xing et al, 1993), belong to the group of superinducible genes for which an enhancement of transcript accumulation is induced by protein synthesis inhibitors, albeit via different mechanisms (Maier et al, 1993;Faggioli et al, 1997;Roger et al, 1998a). The IL-8 gene that was superinduced by cycloheximide treatment (10 mg ml 21 ) in lung epithelial H292 cells (Roger et al, 1998b) was not overexpressed in C. trachomatis-infected HeLa cells exposed to 1 mg ml 21 inhibitor, at least at the protein level. Rasmussen et al (1998) showed that, in HeLa cells infected with C. trachomatis serovar L2, the late maximal induction of cytokine resulted from a NF-kB-independent process.…”

Metalloproteinase-dependent Transforming Growth Factor-α Release Mediates Neurotensin-stimulated MAP Kinase Activation in Human Colonic Epithelial Cells

“…Our group has previously shown that inhibition of MAP kinase activation does not affect NF-B DNA binding activity in response to H. pylori infections (56) and NT (20). Instead, the effect of the MAP kinase pathway in IL-8 gene expression may be mediated through increased transcriptional activity of AP-1, one of three transcriptional factors important for IL-8 gene transcription (22,57,58).…”