The structure of the rat ameloblastin gene and its expression in amelogenesis.
Ameloblastin (also designated amelin and sheathlin) is an enamel matrix protein expressed within the ameloblast lineage. In this study we analyzed the structure of the rat ameloblastin gene and characterized its subtypes. The promoter sequence contains several E-box-like elements, and consensus sequences for AP1 and SP1. The gene is about 6 kb in length and contains 12 exons. Exon 1 was mapped by primer extension and encodes 90 bp of 5' untranslated leader sequence, followed by the coding sequences of exon 2 (309 bp), alternatively spliced exon 3a (45 bp), exon 3b (198 bp), exon 4 (36 bp), exon 5 (60 bp), exon 6 (45 bp), exon 7 (150 bp), and exon 8 (448 bp) containing coding sequence (426 bp) and 3' untranslated sequence (22 bp), followed by exon 9 (39 bp); exon 10 (143 bp); exon 11 (342 bp); and exon 12. Exon 3a, encoding YEYSLPVHPPPLPSQ, has a potential SH3 binding domain. Analysis of ameloblastin subclones showed that exon 3a and 11 were potential alternative splicing sites, producing 4 types of ameloblastin mRNA, from which ameloblastin I and II could be translated. Using in situ hybridization, immunohistochemistry, Western blot and RT-PCR methods we found that ameloblastin II, containing exon 3a, was more strongly expressed at the late maturation stage of ameloblasts than at the secretory stage, while a common probe for both ameloblastin subtypes showed wide expression throughout the presecretory, secretory and postsecretory stages. From the above results we propose that ameloblastin II plays an important role in the mineralization of ameloblasts during the maturation stages.
Lee
Suk Keun
SK
Department of Pathology, Seoul National University, Seoul 110-744, Korea.
Kim
Soung Min
SM
Lee
Young Joon
YJ
Yamada
Kenneth M
KM
Yamada
Yoshihiko
Y
Chi
Je G
JG
D017398:Alternative Splicing / Q000235:genetics; D000566:Amelogenesis / Q000235:genetics*; D020816:Amino Acid Motifs / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D003746:Dental Enamel Proteins / Q000096:biosynthesis / Q000235:genetics*; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
C098519:Ambn protein, rat; D003746:Dental Enamel Proteins; D020033:Protein Isoforms