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Epidermal growth factor increases claudin-4 expression mediated by Sp1 elevation in MDCK cells
Akira Ikari1, Kosuke Atomi, Ayumi Takiguchi
1Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan. ikari@u-shizuoka-ken.ac.jp
Summary
Epidermal growth factor (EGF) activates the MEK/ERK pathway, increasing Sp1 expression and subsequently elevating claudin-4 levels in MDCK cells. This study elucidates the molecular mechanisms behind EGF-induced claudin-4 upregulation.
Area of Science:
- Cell biology
- Molecular biology
- Biochemistry
Background:
- Epidermal growth factor (EGF) is known to influence cellular processes.
- Claudins are crucial tight junction proteins involved in epithelial barrier function.
- Understanding the regulation of claudin-4 expression is important for epithelial biology.
Purpose of the Study:
- To investigate the regulatory mechanisms underlying EGF-induced claudin-4 expression in Madin-Darby canine kidney (MDCK) cells.
- To identify the specific signaling pathways and transcription factors involved in this process.
Main Methods:
- Utilized Western blotting to assess protein levels of claudin-4, claudin-1, and phosphorylated ERK1/2 (p-ERK1/2).
- Employed MEK inhibitor (U0126) and Sp1 inhibitor (mithramycin) to block specific signaling pathways.
- Used Sp1 small interfering RNA (siRNA) to knockdown Sp1 expression.
- Performed immunocytochemistry to determine Sp1 localization.
Main Results:
- EGF treatment led to a transient increase in claudin-4 mRNA and a sustained increase in claudin-4 protein, without affecting claudin-1.
- EGF increased p-ERK1/2 levels, indicating MEK/ERK pathway activation, which was confirmed by using a constitutively active MEK.
- EGF also increased Sp1 expression, which was found to be localized in the nucleus.
- Inhibition of MEK/ERK or Sp1 abolished the EGF-induced elevation of claudin-4.
Conclusions:
- The MEK/ERK signaling pathway is critically involved in the EGF-induced upregulation of claudin-4.
- Increased Sp1 expression, mediated by the MEK/ERK pathway, is essential for the elevation of claudin-4.
- EGF-induced claudin-4 expression is regulated by the activation of the MEK/ERK/Sp1 axis.
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