The Signaling Pathway Involved in Neutrophil Elastase–Stimulated MUC1 Transcription

Abstract

We previously reported that neutrophil elastase (NE) stimulated MUC1 gene expression in A549 lung epithelial cells through binding of Sp1 to the MUC1 promoter element. The current study was undertaken to elucidate the complete signaling pathway leading to Sp1 activation. Using a combination of pharmacologic inhibitors, dominantnegative mutant, RNA interference, and soluble receptor blocking techniques, we identified a protein kinase Cd (PKCd) / dual oxidase 1 (Duox1) / reactive oxygen species (ROS) / TNF-a-converting enzyme (TACE) / TNF-a / TNF receptor (TNFR)1 / extracellular signal-regulated kinase (ERK)1/2 / Sp1 pathway as responsible for NE-activated MUC1 transcription. This cascade was identical up to the point of TACE with the signaling pathway previously reported for NE-stimulated MUC5AC production. However, unlike the MUC5AC pathway, TNF-a, TNFR1, ERK1/2, and Sp1 were unique components of the MUC1 pathway. Given the anti-inflammatory role of MUC1 during airway bacterial infection, up-regulation of MUC1 by inflammatory mediators such as NE and TNF-a suggests a crucial role for MUC1 in the control of excessive inflammation during airway bacterial infection.Keywords: neutrophil elastase; MUC1; signaling; airway MUC1 (MUC1 in human and Muc1 in nonhuman species) is a transmembrane mucin-like glycoprotein expressed on the surface of epithelial cells lining various mucosal surfaces, including the respiratory tract (1-3). We recently showed that mice deficient in Muc1 expression exhibited both enhanced airway inflammation and bacterial clearance during Pseudomonas aeruginosa (PA) airway infection (4), suggesting an anti-inflammatory role for Muc1 and the importance of Muc1 levels during airway bacterial infection. The anti-inflammatory activity of MUC1/Muc1 after treatment with bacterial products also was demonstrated in various in vitro studies (4). How the levels of MUC1/Muc1 are regulated during airway bacterial infection is unknown, but recent evidence indicates the involvement of neutrophil elastase (NE) (5).NE is present in micromolar concentrations in airway surface liquid of patients with cystic fibrosis and patients with chronic bronchitis (6, 7), and stimulates mucin release and mucin gene expression by cultured airway epithelial cells through the proteolytic activity (8, 9). Using a co-culture system containing hamster neutrophils and tracheal surface epithelial (TSE) cells, we demonstrated that activation of the neutrophils by fMLP or cytochalasin B resulted in the stimulation of mucin release (10), indicating that the concentrations of NE produced by activation of neutrophils are sufficient to release mucins. To date, NE is the most potent mucin secretagogue that has been described (11).In addition to gel-forming mucins such as MUC5AC, NE also induces the expression of the membrane-bound MUC1 and MUC4 mucins (5, 12). Our prior report showed that MUC1 protein synthesis by A549 cells was enhanced after treatment with NE, and this effect was blocked by pretreatment with actinomycin D or cycloheximide (...

Antiinflammatory Role of MUC1 Mucin during Infection with NontypeableHaemophilus influenzae

“…TNF-a stimulates the expression of MUC1 in A549 cells (39,40), human uterine epithelial cells (41), and human prostate cells (42). The molecular mechanism of the TNF-a-induced up-regulation of MUC1 was described elsewhere (39).…”

Suppression of IL-8 production in gastric epithelial cells by MUC1 mucin and peroxisome proliferator-associated receptor-γ

“…Second, TGN contains an ␣-tocopheroyl moiety, potentially giving it antioxidant activity (40). However, this alternative mechanism of action also appears unlikely based on a prior study demonstrating decreased MUC1 expression following treatment of epithelial cells with antioxidants (29). Finally, phosphorylation of ERK1/2 through the TNF-␣ receptor is known to activate MUC1 expression (25,29), and Bolden et al (8) recently demonstrated that treatment of human prostate cancer cells with TGN induced ERK1/2 phosphorylation via a PPAR␥-independent pathway.…”

“…However, this alternative mechanism of action also appears unlikely based on a prior study demonstrating decreased MUC1 expression following treatment of epithelial cells with antioxidants (29). Finally, phosphorylation of ERK1/2 through the TNF-␣ receptor is known to activate MUC1 expression (25,29), and Bolden et al (8) recently demonstrated that treatment of human prostate cancer cells with TGN induced ERK1/2 phosphorylation via a PPAR␥-independent pathway. However, this effect was only observed at relatively high TGN concentrations (40 M) and no effect on ERK1/2 activation was seen at the doses used in the present study (Ͻ 1.0 M).…”

NADPH oxidases in lung biology and pathology: Host defense enzymes, and more

“…Moreover, the airway epithelium also generates airway mucins that facilitate mucociliary clearance of inhaled particulates or microorganisms (103,104). Recent studies suggest that DUOX may participate in these responses as well, as illustrated by DUOX1-mediated production of epithelial mucins, such as MUC1 and MUC5AC, in response to inflammatory mediators such as neutrophil elastase or tumor necrosis factor (TNF)-α (105,106). Similarly, DUOX1 also mediates epithelial production of the neutrophil chemokine IL-8 in response to stimulation by bacterial lipopolysaccharide (LPS) (107).…”