Amelotin gene expression is temporarily being upregulated at the initiation of apoptosis induced by TGF[beta]1 in mouse gingival epithelial cells

• Published in: Apoptosis (London) Vol. 21; no. 10; p. 1057
• Main Authors: Nakayama, Yohei, Matsui, Sari, Noda, Keisuke, Yamazaki, Mizuho, Iwai, Yasunobu, Matsumura, Hiroyoshi, Izawa, Takashi, Tanaka, Eiji, Ganss, Bernhard, Ogata, Yorimasa
• Format: Journal Article
• Language: English
• Published: London Springer Nature B.V 01.10.2016
• Subjects: Deoxyribonucleic acid; DNA
• ISSN: 1360-8185; 1573-675X
• DOI: 10.1007/s10495-016-1279-5

Amelotin (AMTN) is expressed and secreted by ameloblasts in the maturation stage of amelogenesis and persist with low levels in the junctional epithelium (JE) of erupted teeth. The purpose of this study is to investigate the transcriptional regulation of the AMTN gene by transforming growth factor beta1 (TGF[beta]1) in gingival epithelial (GE1) cells in the apoptosis phase. Apoptosis was evaluated by the fragmentation of chromosomal DNA and TUNEL staining. A real-time PCR was carried out to examine the AMTN mRNA levels induced by TGF[beta]1 and Smad3 overexpression. Transient transfection analyses were completed using the various lengths of mouse AMTN gene promoter constructs with or without TGF[beta]1. Chromatin immunoprecipitation (ChIP) assays were performed to investigate the Smad3 bindings to the AMTN gene promoter by TGF[beta]1. TGF[beta]1-induced apoptosis in GE1 cells were detected at 24 and 48 h by DNA fragmentation and TUNEL staining. AMTN mRNA levels increased at 6 h and reached maximum at 24 h in GE1 cells. Luciferase activities of the mouse AMTN gene promoter constructs were induced by TGF[beta]1. The results of the ChIP assays showed that there was an increase in Smad3 binding to Smad-binding element (SBE)#1 and SBE#2 after stimulation by TGF[beta]1. Immunohistochemical localization of AMTN was detected in the JE, and the AMTN protein levels in Smad3-deficient mice were decreased compared with wild-type mice. AMTN mRNA levels were induced at the initiation of apoptosis by TGF[beta]1, which mediated through the Smad3 bindings to SBEs in the mouse AMTN gene promoter.