TGFβ1-induced Amelotin gene expression is downregulated by Bax expression in mouse gingival epithelial cells

Abstract

Amelotin (AMTN) is induced upon initiation of apoptosis by transforming growth factor beta1 (TGFβ1) and is mediated by Smad3 in gingival epithelial cells (GE1 cells). This upregulation of AMTN gene expression is temporary, and the mechanism responsible is still unclear. The present study investigated the transcriptional downregulation of TGFβ1-induced AMTN gene expression in GE1 cells during the progression of apoptosis. To examine time-dependent changes in the levels of AMTN, Smad3 and Bax mRNA induced by TGFβ1, real-time PCR analyses were performed. Immunocytochemistry was carried out to detect the expression of Smad3 and Bax. Transient transfection analyses were performed using mouse AMTN gene promoter constructs of various lengths including Smad response elements (SBEs), in the presence or absence of TGFβ1. Changes in Smad3 binding to SBEs resulting from overexpression of Bax were examined using ChIP assays. Overexpression of Bax dramatically downregulated the levels of TGFβ1-induced AMTN mRNA and transcription of the AMTN gene. Smad3 binding to SBEs in the mouse AMTN gene promoter was induced by overexpression of Smad3 or TGFβ1, and this was inhibited by Bax overexpression. These results show that the levels of AMTN mRNA induced by TGFβ1 and Smad3 are decreased by robust expression of Bax in gingival epithelial cells.

C/EBPβ and YY1 bind and interact with Smad3 to modulate lipopolysaccharide‐induced amelotin gene transcription in mouse gingival epithelial cells

“…Amtn gene transcription is tightly regulated and is affected by the activity of inflammatory cytokines and their byproducts. AMTN can lead to pathologies including inflammation , apoptosis and cancer . In this study we have shown that Pg LPS induced Amtn gene transcription that was mediated through C/EBPβ, YY1 and Smad3 binding to C/EBP1, C/EBP2 and YY1 elements, which was accompanied by physical interaction between C/EBPβ, YY1 and Smad3.…”

“…We have previously shown a relation between inflammatory cytokines and expression of JE‐specific genes, such as amelotin (AMTN) , odontogenic ameloblast‐associated protein (ODAM) and follicular dendritic cell‐secreted protein (FDC‐SP) , during dental development and regeneration of JE in gingival epithelial cells and a Porphyromonas gingivalis ( Pg )‐ and Aggregatibacter actinomycetemcomitans ‐infected mouse periodontitis model . Moreover, we have reported that Amtn gene expression was temporarily increased via Smad3 binding to Smad binding elements (SBEs) at the initiation of transforming growth factor β1 (TGF‐β1)‐induced apoptosis in gingival epithelial cells .…”

Negative feedback by SNAI2 regulates TGFβ1‐induced amelotin gene transcription in epithelial–mesenchymal transition

“…Therefore, these data suggest the existence of hypersensitive sites from SBE5 to E4 elements in the mouse AMTN gene promoter, additional research is required to confirm these observations. TGFβ1-induced Smad3 bindings to SBEs resulted in upregulation of AMTN gene expression in GE1 cells (Nakayama et al, 2016(Nakayama et al, , 2018. In this study, TGFβ1-induced AMTN expression levels were not completely abrogated by SNAI2 expression (Figure 10c).…”

“…The fixed cells were rinsed with wash buffer (10 mM PMSF and complete protease inhibitor cocktail in the PBS) on ice twice, collected by scrape and centrifuged for 5 min at 4°C. Following procedures were briefly described because they were carried out in the same manner as the previous study (Nakayama et al, 2018).…”

Recombinant adiponectin peptide promotes neuronal survival after intracerebral haemorrhage by suppressing mitochondrial and ATF4‐CHOP apoptosis pathways in diabetic mice via Smad3 signalling inhibition

“…Smad3 is a transcription factor; once phosphorylated and activated, it translocates to the nucleus to bind to Smad-binding elements (SBEs) in target gene promoters, influencing the transcription of the target genes and thus regulating various cellular processes. 35,36 Previous research used sequence analysis to reveal the presence of SBEs in the promoters of PGC-1α and ATF4 and found that activated Smad3 signalling could transcriptionally decrease PGC-1α levels 37 and increase ATF4 levels 38 in adipose tissue. This led us to question whether Smad3 played a distinctive role in regulating the mitochondrial and ATF4-CHOP apoptosis pathways in diabetic mice that contribute to aggravated neural apoptosis following ICH.…”