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Journal of Histochemistry and Cytochemistry, Vol. 50, 1229-1236, September 2002, Copyright © 2002, The Histochemical Society, Inc.

ARTICLE

Expression of Alternatively Spliced RNA Transcripts of Amelogenin Gene Exons 8 and 9 and Its End Products in the Rat Incisor

Otto Babaa, Nobuyuki Takahashia, Tatsuo Terashimaa, Wu Lib, Pamela K. DenBestenb, and Yoshiro Takanoa
a Biostructural Science, Department of Hard Tissue Engineering, Division of Bio-Matrix, Graduate School of Tokyo Medical and Dental University, Tokyo, Japan
b Department of Growth and Development, University of California at San Francisco, San Francisco, California

Correspondence to: Yoshiro Takano, Biostructural Science, Dept. of Hard Tissue Engineering, Div. of Bio-Matrix, Graduate School of Tokyo Medical and Dental University, 5-45, Yushima 1-chome, Bunkyo-ku, Tokyo 113-8549, Japan. E-mail: takanoy.bss@tmd.ac.jp

In addition to seven known exons of the amelogenin gene, recent studies have identified two exons downstream of amelogenin exon 7 in genomic DNA of mouse and rat. Here the spatial and temporal expression of mRNAs and of the translated proteins derived from alternative splicing of the amelogenin gene ending with exon 8 and exon 9 were examined by in situ hybridization (ISH) and immunohistochemistry (IHC). RNA signals for exons 8 and 9 were expressed in the ameloblast layer extending from early presecretory to postsecretory transitional stages of amelogenesis. IHC of amelogenin proteins that include sequences encoded by these exons demonstrated identical localization of these proteins in the ameloblast layer corresponding to RNA signals identified by ISH. There was intense immunostaining of the enamel matrix secreted by these cells. Western blotting analysis of rat enamel proteins revealed three distinct protein bands with sequences encoded by the new exons. These data confirmed the existence of the transcripts of alternatively spliced mRNAs coding for exons 8 and 9 of the amelogenin gene in rat tooth germs and suggest that the translated proteins contribute to the heterogeneity of amelogenins and have some significant roles in enamel formation and mineralization.

(J Histochem Cytochem 50:1229–1236, 2002)

Key Words: amelogenin, alternative splicing, exons 8 and 9, rat, in situ hybridization, immunohistochemistry


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