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

Abstract

S U M M A R YIn 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.

Molecular Evolution of Amelogenin in Mammals

“…This gene is autosomal in monotremes (prototherians) and in marsupials (metatherians) (Watson et al 1992). The amelogenin gene is generally known to be composed of seven exons, but two extra exons, exons 8 and 9, exist at least in human and rodents (Li et al 1998;Baba et al 2002). Exon 1 is not translated and most of the exon 2 sequence codes for the leader peptide (Delgado et al 2001).…”

Amelogenin in cranio‐facial development: the tooth as a model to study the role of amelogenin during embryogenesis

“…The gene contains 9 exons ( Li et al, 1998;Baba et al, 2002) which undergo extensive alternative mRNA splicing (Simmer and Fincham, 1995;Veis, 2003;Papagerakis et al, 2005). The most abundant amelogenin isoform in the mouse enamel is M180, which is encoded by exons 1-7, skipping exon 4.…”

Amelogenin expression in long bone and cartilage cells and in bone marrow progenitor cells

“…The most abundant isoform of the native protein secreted into the enamel matrix lacks the internal region encoded by exon 4 (Salido et al, 1992). Two additional exons downstream of exon 7 have been identified in a rare alternatively spliced RNA transcript of amelogenin (Li et al, 1998;Baba et al, 2002). The relatively large number of amelogenin alternatively spliced mRNA isoforms, code for several proteins with different structures and characteristics, which might have different functions (Veis, 2003).…”