Tooth enamel is formed in a typical biomineralization process under the guidance of specific organic components. Amelotin (AMTN) is a recently identified, secreted protein that is transcribed predominantly during the maturation stage of enamel formation, but its protein expression profile throughout amelogenesis has not been described in detail. The main objective of this study was to define the spatiotemporal expression profile of AMTN during tooth development in comparison with other known enamel proteins. A peptide antibody against AMTN was raised in rabbits, affinity purified and used for immunohistochemical analyses on sagittal and transverse paraffin sections of decalcified mouse hemimandibles. The localization of AMTN was compared to that of known enamel proteins amelogenin, ameloblastin, enamelin, odontogenic ameloblast-associated/amyloid in Pindborg tumors and kallikrein 4. Three-dimensional images of AMTN localization in molars at selected ages were reconstructed from serial stained sections, and transmission electron microscopy was used for ultrastructural localization of AMTN. AMTN was detected in ameloblasts of molars in a transient fashion, declining at the time of tooth eruption. Prominent expression in maturation stage ameloblasts of the continuously erupting incisor persisted into adulthood. In contrast, amelogenin, ameloblastin and enamelin were predominantly found during the early secretory stage, while odontogenic ameloblast-associated/amyloid in Pindborg tumors and kallikrein 4 expression in maturation stage ameloblasts paralleled that of AMTN. Secreted AMTN was detected at the interface between ameloblasts and the mineralized enamel. Recombinant AMTN protein did not mediate cell attachment in vitro. These results suggest a primary role for AMTN in the late stages of enamel mineralization.

Keywords:
Amelotin, Amelogenesis, Protein expression, Enamel
1.
Beniash, E., Z. Skobe, J.D. Bartlett (2006) Formation of the dentino-enamel interface in enamelysin (MMP-20)-deficient mouse incisors. Eur J Oral Sci 114(suppl 1): 24–29, discussion 39–41, 379.
2.
Caterina, J.J., Z. Skobe, J. Shi, Y. Ding, J.P. Simmer, H. Birkedal-Hansen, J.D. Bartlett (2002) Enamelysin (matrix metalloproteinase 20)-deficient mice display an amelogenesis imperfecta phenotype. J Biol Chem 277: 49598–49604.
3.
Fukumoto, S., T. Kiba, B. Hall, N. Iehara, T. Nakamura, G. Longenecker, P.H. Krebsbach, A. Nanci, A.B. Kulkarni, Y. Yamada (2004) Ameloblastin is a cell adhesion molecule required for maintaining the differentiation state of ameloblasts. J Cell Biol 167: 973–983.
4.
Gao, Y., W. Wang, Y. Sun, J. Zhang, D. Li, Y. Wei, T. Han (2010) Distribution of amelotin in mouse tooth development. Anat Rec (Hoboken) 293: 135–140.
5.
Gibson, C.W., Z.A. Yuan, B. Hall, G. Longenecker, E. Chen, T. Thyagarajan, T. Sreenath, J.T. Wright, S. Decker, R. Piddington, G. Harrison, A.B. Kulkarni (2001) Amelogenin-deficient mice display an amelogenesis imperfecta phenotype. J Biol Chem 276: 31871–31875.
6.
Hu, J.C., Y.H. Chun, T. Al Hazzazzi, J.P. Simmer (2007) Enamel formation and amelogenesis imperfecta. Cells Tissues Organs 186: 78–85.
7.
Hu, J.C., Y. Hu, C.E. Smith, M.D. McKee, J.T. Wright, Y. Yamakoshi, P. Papagerakis, G.K. Hunter, J.Q. Feng, F. Yamakoshi, J.P. Simmer (2008) Enamel defects and ameloblast-specific expression in Enam knock-out/LacZ knock-in mice. J Biol Chem 283: 10858–10871.
8.
Iwasaki, K., E. Bajenova, E. Somogyi-Ganss, M. Miller, V. Nguyen, H. Nourkeyhani, Y. Gao, M. Wendel, B. Ganss (2005) Amelotin – a novel secreted, ameloblast-specific protein. J Dent Res 84: 1127–1132.
9.
Kinumatsu, T., S. Hashimoto, T. Muramatsu, H. Sasaki, H.S. Jung, S. Yamada, M. Shimono (2009) Involvement of laminin and integrins in adhesion and migration of junctional epithelium cells. J Periodontal Res 44: 13–20.
10.
Kodaka, T., M. Kuroiwa, S. Higashi (1991) Structural and distribution patterns of surface ‘prismless’ enamel in human permanent teeth. Caries Res 25: 7–20.
11.
Lu, Y., P. Papagerakis, Y. Yamakoshi, J.C. Hu, J.D. Bartlett, J.P. Simmer (2008) Functions of KLK4 and MMP-20 in dental enamel formation. Biol Chem 389: 695–700.
12.
Lyngstadaas, S.P., C.B. Moinichen, S. Risnes (1998) Crown morphology, enamel distribution, and enamel structure in mouse molars. Anat Rec 250: 268–280.
13.
McKee, M.D., A. Nanci (1995) Postembedding colloidal-gold immunocytochemistry of noncollagenous extracellular matrix proteins in mineralized tissues. Microsc Res Tech 31: 44–62.
14.
Moffatt, P., C.E. Smith, R. St-Arnaud, D. Simmons, J.T. Wright, A. Nanci (2006) Cloning of rat amelotin and localization of the protein to the basal lamina of maturation stage ameloblasts and junctional epithelium. Biochem J 399: 37–46.
15.
Moinichen, C.B., S.P. Lyngstadaas, S. Risnes (1996) Morphological characteristics of mouse incisor enamel. J Anat 189: 325–333.
16.
Nakata, A., T. Kameda, H. Nagai, K. Ikegami, Y. Duan, K. Terada, T. Sugiyama (2003) Establishment and characterization of a spontaneously immortalized mouse ameloblast-lineage cell line. Biochem Biophys Res Commun 308: 834–839.
17.
Nanci, A. (2007) Ten Cate’s Oral Histology: Development, Structure, and Function. St. Louis, Mosby.
18.
Paine, M.L., M.L. Snead (2005) Tooth developmental biology: disruptions to enamel-matrix assembly and its impact on biomineralization. Orthod Craniofac Res 8: 239–251.
19.
Paine, M.L., D.H. Zhu, W. Luo, P. Bringas Jr., M. Goldberg, S.N. White, Y.P. Lei, M. Sarikaya, H.K. Fong, M.L. Snead (2000) Enamel biomineralization defects result from alterations to amelogenin self-assembly. J Struct Biol 132: 191–200.
20.
Pender, N., C.A. McCulloch (1991) Quantitation of actin polymerization in two human fibroblast sub-types responding to mechanical stretching. J Cell Sci 100: 187–193.
21.
Risnes, S., C.B. Moinichen, D. Septier, M. Goldberg (1996) Effects of accelerated eruption on the enamel of the rat lower incisor. Adv Dent Res 10: 261–269.
22.
Shimono, M., T. Ishikawa, Y. Enokiya, T. Muramatsu, K. Matsuzaka, T. Inoue, Y. Abiko, T. Yamaza, M.A. Kido, T. Tanaka, S. Hashimoto (2003) Biological characteristics of the junctional epithelium. J Electron Microsc (Tokyo) 52: 627–639.
23.
Simmer, J.P., Y. Hu, R. Lertlam, Y. Yamakoshi, J.C. Hu (2009) Hypomaturation enamel defects in Klk4 knockout/LacZ knockin mice. J Biol Chem 284: 19110–19121.
24.
Siqueira, W.L., H.C. Margolis, E.J. Helmerhorst, F.M. Mendes, F.G. Oppenheim (2010) Evidence of intact histatins in the in vivo acquired enamel pellicle. J Dent Res 89: 626–630.
25.
Sire, J.Y., T. Davit-Beal, S. Delgado, X. Gu (2007) The origin and evolution of enamel mineralization genes. Cells Tissues Organs 186: 25–48.
26.
Sudo, H., H.A. Kodama, Y. Amagai, S. Yamamoto, S. Kasai (1983) In vitro differentiation and calcification in a new clonal osteogenic cell line derived from newborn mouse calvaria. J Cell Biol 96: 191–198.
27.
Thesleff, I., T. Aberg (1997) Tooth morphogenesis and the differentiation of ameloblasts. Ciba Found Symp 205: 3–12, discussion 12–17.
28.
Timpl, R. (1989) Structure and biological activity of basement membrane proteins. Eur J Biochem 180: 487–502.
29.
Warshawsky, H. (1971) A light and electron microscopic study of the nearly mature enamel of rat incisors. Anat Rec 169: 559–583.
30.
Wazen, R.M., P. Moffatt, S.F. Zalzal, Y. Yamada, A. Nanci (2009) A mouse model expressing a truncated form of ameloblastin exhibits dental and junctional epithelium defects. Matrix Biol 28: 292–303.
31.
Weile, V., K. Josephsen, O. Fejerskov (1993) Scanning electron microscopy of final enamel formation in rat mandibular incisors following single injections of 1-hydroxyethylidene-1,1-bisphosphonate. Calcif Tissue Int 52: 318–324.
32.
Zeichner-David, M., T. Diekwisch, A. Fincham, E. Lau, M. MacDougall, J. Moradian-Oldak, J. Simmer, M. Snead, H.C. Slavkin (1995) Control of ameloblast differentiation. Int J Dev Biol 39: 69–92.
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2011
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