Amelogenesis imperfecta (AI) is a collection of rare genetic conditions affecting tooth enamel. The affected enamel can be of insufficient quantity and/or altered quality, impacting structural content, surface integrity and coloration. Heterozygous mutations in ENAM result in hypoplastic AI without other syndromic phenotypes, with variable expressivity and reduced penetrance, unlike other AI-associated genes. In this study, we recruited a Caucasian family with hypoplastic AI. Mutational analysis (using whole exome sequencing) revealed a splicing donor site mutation (NM_031889.3: c. -61 + 1G > A). Mutational effects caused by this variant were investigated with a minigene splicing assay and in vitro expression analysis. The mutation resulted in a retention of intron 1 and exon 2 (a normally skipped exon), and this elongated 5' UTR sequence attenuated the translation from the mutant mRNA. Structure and translation predictions raised the possibility that the long complex structures-especially a hairpin structure located right before the translation initiation codon of the mutant mRNA-caused reduced protein expression. However, there could be additional contributing factors, including additional uORFs. For the first time, we determined that a mutation altered the ENAM 5' UTR, but maintained the normal coding amino acid sequence, causing hypoplastic AI.
Keywords: ENAM; amelogenesis imperfecta; hereditary enamel defects; intron retention; splicing donor site mutation; whole exome sequencing.