Upregulation of MMP-20 expression by P70T mutation in amelogenin

Amelogenin is the predominant extracellular matrix protein in developing enamel and is rapidly processed by matrix metalloproteinase (MMP-20) after secretion. Transgenic mice carrying a proline-to-threonine (P70T) substitution of amelogenin display a hypomineralized type of amelogenesis imperfecta (AI), similar to that in the patients with the identical mutation. Our previous in vitro studies found that this specific proline change in amelogenin reduced protein hydrolysis by MMP-20. More amelogenin protein is present in P70T mice as compared to control mice. Objectives: The aim of this study is to determine the effect of the P70T mutation in amelogenin on MMP-20 expression by ameloblasts. Methods: Total RNA was extracted from mandibles of 5-day-old male mice and MMP-20 expression was quantified by real-time PCR. MMP-20 protein levels in the P70T and control M180 mice were compared by immunocytochemistry, Western blotting and quantitative mass spectrometry. Results: MMP-20 mRNA expression was increased 1.9-fold in P70T mice compared with the control M180 mice. The MMP-20 protein level was also found to be greatly up-regulated in P70T mice. Conclusions: The expression of MMP-20 was up-regulated in mice with reduced amelogenin hydrolysis. This suggests that a compensatory feedback may contribute to MMP-20 regulation. However, higher amelogenin retention in enamel matrix of these mutant mice indicates that this up-regulation appears not to fully compensate for reduced hydrolysis of amelogenin resulting from P70T mutation. These findings provide better understanding of AI and amelogenin-protease interactions in tooth enamel development. Supported by grant R01DE015821, R01DE011089 and R01DE13508 from NIDCR.