The exon 6ABC region of amelogenin mRNA contribute to increased levels of amelogenin mRNA through amelogenin protein-enhanced mRNA stabilization.
We recently demonstrated that the reuptake of full-length amelogenin protein results in increased levels of amelogenin mRNA through enhanced mRNA stabilization (Xu, L., Harada, H., Tamaki, T. Y., Matsumoto, S., Tanaka, J., and Taniguchi, A. (2006) J. Biol. Chem. 281, 2257-2262). Here, we examined the molecular mechanism of enhanced amelogenin mRNA stabilization. To identify the cis-regulatory region within amelogenin mRNA, we tested various reporter systems using a deletion series of reporter plasmids. A deletion at exon 6ABC of amelogenin mRNA resulted in a 2.5-fold increase in the amelogenin mRNA expression level when compared with that of full-length mRNA, indicating that a cis-element exists in exon 6ABC of amelogenin mRNA. Furthermore, Northwestern analysis demonstrated that amelogenin protein binds directly to its mRNA in vitro, suggesting that amelogenin protein acts as a trans-acting protein that specifically binds to this cis-element. Moreover, recombinant mouse amelogenin protein extended the half-life of full-length amelogenin mRNA but did not significantly alter the half-life of exon 6ABC-deletion mutant mRNA. The splice products produced by deletion of exon 6ABC are known as leucine-rich amelogenin peptides and have signaling effects on cells. Our findings also suggest that the regulation of full-length amelogenin protein expression differs from the regulation of leucine-rich amelogenin peptide expression.
Xu
Liming
L
Cell Engineering Technology Group, Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044, Japan.
Harada
Hidemitsu
H
Taniguchi
Akiyoshi
A
D053523:Amelogenin / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D003609:Dactinomycin / Q000494:pharmacology; D004847:Epithelial Cells / Q000378:metabolism; D005091:Exons; D017930:Genes, Reporter; D006207:Half-Life; D008156:Luciferases / Q000378:metabolism; D010957:Plasmids; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection
D053523:Amelogenin; D012333:RNA, Messenger; D011994:Recombinant Proteins; D003609:Dactinomycin; D008156:Luciferases