Expression and function of OD314, Apin protein during ameloblast differentiation and amelogenesis

Article information

Restor Dent Endod. 2006;31(6):437-444

Publication date (electronic) : 2006 November 30

doi : https://doi.org/10.5395/JKACD.2006.31.6.437

Jong-Tae Park ¹, Yong-Seok Choi ¹, Heung-Joong Kim ¹, Moon-Jin Jeong ¹, Hyun-Ju Oh ¹, In-Cheol Shin ¹, Joo-Cheol Park ¹, Ho-Hyun Son ²

¹Department of Oral Histology, College of Dentistry, Chosun University, Korea.

²Department of Conservative Dentistry, School of Dentistry, Seoul National University, Korea.

Corresponding author: Ho-Hyun Son. Department of Conservative Dentistry, School of Dentistry, Seoul National University, 22 Yeongun-dong, Chongro-gu, Seoul, Korea. Tel: 82-2-2072-2652, Fax: 82-2-2072-3859, hhson@snu.ac.kr

Received 2006 August 24; Revised 2006 September 19; Accepted 2006 September 20.

Abstract

This study was aimed to elucidate the biological function of OD314 (Apin protein), which is related toameloblast differentiation and amelogenesis. Apin protein, calcifying epithelial odontogenic (pindborg) tumors (CEOTs)-associated amyloid, were isolated from CEOTs, and has similar nucleotide sequences to OD314. We examined expression of the OD314 mRNA using in-situ hybridization during tooth development in mice. Expression of OD314 and several enamel matrix proteins were examined in the cultured ameloblast cell line up to 28 days by reverse transcription-polymerase chain reaction (RT-PCR) amplification. After inactivation and over-expression of the OD314 gene in ameloblast cell lines using U6 vector-driven RNA interference and CMV-OD314 construct, RT-PCR were performed to evaluate the effect of the OD314 during amelogenesis.

The results were as follows:

1. In in-situ hybridization, OD314 mRNAs were more strongly expressed in ameloblast than odontoblast.

2. When ameloblast cells were cultured in the differentiation and mineralization medium for 28 days, the tuftelin mRNA expression was maintained from the beginning to day 14, and then gradually decreased to day 28. The expressions of amelogenin and enamelin were gradually decreased according to the ameloblast differentiation.

3. Inactivation of OD314 by U6-OD314 siRNA construct down-regulated the expression of OD314, MMP-20, and tuftelin, whereas over-expression of OD314 by CMV-OD314 construct up-regulated the expression of OD314 and MMP-20 without change in tuftelin.

These results suggest that OD314 is considered as an ameloblast-enriched gene and may play the important roles in ameloblast differentiation and mineralization.

Keywords: Apin protein; OD314; Mineralization; Ameloblast differentiation; Amelogenesis

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Figure 1

Maxillary molar of 1-week-old mouse hybridized with antisense cRNA probes of OD314. Intense signal for OD314 mRNA is detected in ameloblast (arrow).

Figure 2

RT-PCR amplification of OD314 in the cultured ameloblast cell line up to 28 days.

Figure 3

RT-PCR amplification of enamel matrix proteins in the cultured ameloblast cell line up to 28 days.

Figure 4

RT-PCR amplification of OD314 in ameloblast cell line after over-expression with CMV-OD314 plasmid and inactivation with U6-OD314 siRNA.

Figure 5

RT-PCR amplification of matrix protein in ameloblast cell line after over-expression with CMV-OD314 plasmid and inactivation with U6-OD314 siRNA.

Table 1

Nucleotide sequences of the primers used for RT-PCR