Amelotin gene expression is temporarily being upregulated at the initiation of apoptosis induced by TGFβ1 in mouse gingival epithelial cells

Yohei Nakayama, Sari Matsui, Keisuke Noda, Mizuho Yamazaki, Yasunobu Iwai, Hiroyoshi Matsumura, Takashi Izawa, Eiji Tanaka, Bernhard Ganss, Yorimasa Ogata

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

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 (TGFβ1) 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 TGFβ1 and Smad3 overexpression. Transient transfection analyses were completed using the various lengths of mouse AMTN gene promoter constructs with or without TGFβ1. Chromatin immunoprecipitation (ChIP) assays were performed to investigate the Smad3 bindings to the AMTN gene promoter by TGFβ1. TGFβ1-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 TGFβ1. 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 TGFβ1. 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 TGFβ1, which mediated through the Smad3 bindings to SBEs in the mouse AMTN gene promoter.

Original languageEnglish
Pages (from-to)1057-1070
Number of pages14
JournalApoptosis
Volume21
Issue number10
DOIs
Publication statusPublished - Oct 1 2016
Externally publishedYes

Fingerprint

Transforming Growth Factor beta1
Gene expression
Epithelial Cells
Apoptosis
Gene Expression
Genes
Epithelial Attachment
Chromatin Immunoprecipitation
In Situ Nick-End Labeling
DNA Fragmentation
Messenger RNA
Chromatin
Assays
Amelogenesis
Ameloblasts
Staining and Labeling
DNA
Luciferases
Transfection
Real-Time Polymerase Chain Reaction

Keywords

  • Amelotin
  • Apoptosis
  • Junctional epithelium
  • Smad3
  • TGFβ1

ASJC Scopus subject areas

  • Pharmacology
  • Pharmaceutical Science
  • Clinical Biochemistry
  • Cell Biology
  • Biochemistry, medical
  • Cancer Research

Cite this

Amelotin gene expression is temporarily being upregulated at the initiation of apoptosis induced by TGFβ1 in mouse gingival epithelial cells. / Nakayama, Yohei; Matsui, Sari; Noda, Keisuke; Yamazaki, Mizuho; Iwai, Yasunobu; Matsumura, Hiroyoshi; Izawa, Takashi; Tanaka, Eiji; Ganss, Bernhard; Ogata, Yorimasa.

In: Apoptosis, Vol. 21, No. 10, 01.10.2016, p. 1057-1070.

Research output: Contribution to journalArticle

Nakayama, Y, Matsui, S, Noda, K, Yamazaki, M, Iwai, Y, Matsumura, H, Izawa, T, Tanaka, E, Ganss, B & Ogata, Y 2016, 'Amelotin gene expression is temporarily being upregulated at the initiation of apoptosis induced by TGFβ1 in mouse gingival epithelial cells', Apoptosis, vol. 21, no. 10, pp. 1057-1070. https://doi.org/10.1007/s10495-016-1279-5
Nakayama, Yohei ; Matsui, Sari ; Noda, Keisuke ; Yamazaki, Mizuho ; Iwai, Yasunobu ; Matsumura, Hiroyoshi ; Izawa, Takashi ; Tanaka, Eiji ; Ganss, Bernhard ; Ogata, Yorimasa. / Amelotin gene expression is temporarily being upregulated at the initiation of apoptosis induced by TGFβ1 in mouse gingival epithelial cells. In: Apoptosis. 2016 ; Vol. 21, No. 10. pp. 1057-1070.
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AU - Nakayama, Yohei

AU - Matsui, Sari

AU - Noda, Keisuke

AU - Yamazaki, Mizuho

AU - Iwai, Yasunobu

AU - Matsumura, Hiroyoshi

AU - Izawa, Takashi

AU - Tanaka, Eiji

AU - Ganss, Bernhard

AU - Ogata, Yorimasa

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AB - 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 (TGFβ1) 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 TGFβ1 and Smad3 overexpression. Transient transfection analyses were completed using the various lengths of mouse AMTN gene promoter constructs with or without TGFβ1. Chromatin immunoprecipitation (ChIP) assays were performed to investigate the Smad3 bindings to the AMTN gene promoter by TGFβ1. TGFβ1-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 TGFβ1. 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 TGFβ1. 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 TGFβ1, which mediated through the Smad3 bindings to SBEs in the mouse AMTN gene promoter.

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