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Amelogenin p.M1T and p.W4S Mutations Underlying Hypoplastic X-linked Amelogenesis Imperfecta
J.-W. Kim1,2,
J.P. Simmer1,
Y.Y. Hu1,
B.P.-L. Lin3,
C. Boyd4,
J.T. Wright4,
C.J.M. Yamada1,
S.K. Rayes1,
R.J. Feigal5 and
J.C.-C. Hu1,*
1 Department of Orthodontics and Pediatric Dentistry, University of Michigan Dental Research Lab, 1210 Eisenhower Place, Ann Arbor, MI 48108;
2 Seoul National University, College of Dentistry, Department of Pediatric Dentistry & Dental Research Institute, 28-2 Yongon-dong, Chongno-gu, Seoul, Korea 110-768;
3 University of Texas Health Science Center at San Antonio, Department of Pediatric Dentistry, 7703 Floyd Curl Drive, San Antonio, TX 78289-3900;
4 The University of North Carolina at Chapel Hill, School of Dentistry, Dental Research Center, Chapel Hill, NC 27599-7455; and
5 Department of Preventive Sciences, Moos Health Sciences Tower, 515 Delaware Street SE, Minneapolis, MN 55455;
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Figure 1. Strategy for mutation analysis. The structure of the human amelogenin gene is shown at the top. The exons are blocks numbered 1 through 7. Below each exon is a bar corresponding to the region amplified and a number indicating the number of nucleotides in each PCR product. The DNA sequences of the primer pairs used to generate the exon-specific PCR amplification products, written in the 5' to 3' orientation, are shown below, along with the annealing temperature used in the amplifications, provided in the table at the bottom. The PCR reactions have a five-minute denaturation at 94°C, followed by 40–50 cycles each with denaturation at 94°C for 30 sec, primer annealing at 56–61°C (as shown in the table) for 30 sec, and product extension at 72°C for 40 sec. In the final cycle, the 72°C extension was for 5 min. PCR amplification products were purified by use of the QIAquick PCR Purification Kit (Qiagen Inc., Valencia, CA, USA), and are shown on a 1% agarose gel stained with ethidium bromide, in the center of the Fig.
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Figure 2. Genotype and phenotypes of AI families. Panels A through I are from family 1 (p.M1T): oral photograph of the proband (A); the pedigree (B); bitewing radiographs of the proband, showing the very thin enamel layer that is evident only radiographically on the cusp tips (C,D); DNA sequencing chromatograms (E–H) of exon 2 from an affected male member (IV-2) showing the mutation (Mut) that changed the ATG of the wild-type (Wt) start codon into an ACG, which is normally a threonine codon; and single-stranded conformational polymorphism (SSCP) analysis of exon 2, with arrowheads pointing to the extra band that correlates with affection status (I). Panels J through P are from family 2 (p.W4S): oral photographs of the proband (J, III-2) and her mother (K, II-2), who were both affected; the pedigree of family 2 (L); and DNA sequencing chromatograms from the proband, showing the mutation in the fourth codon of exon 2 (M-P), which changed the wild-type tryptophan codon (TGG) into a serine (TCG) codon.
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Figure 3. Analysis of naturally exfoliated primary teeth from affected girls in family 1 (p.M1T). Ground sections of normal (A,C) and defective (B,D) teeth illustrate the relative thinness (1/4 of normal) of the enamel layer in the affected teeth. The enamel prism structure of the normal teeth is evident in scanning electron micrographs (E, bar = 100 µm), but is not evident, even at twice the magnification (F, bar = 100 µm), in affected teeth. At higher magnification, prism organization that only partially approaches that observed in normal enamel (G, bar = 10 µm) could be recognized in occasional areas (H, bar = 10 µm). To the unaided eye, the enamel surface of the affected teeth looked rough (I), which was due to the presence of pits along the enamel surface that varied in size from about 10 to 30 µm (F). The dentin appeared to be normal at low (J) and high (K, bar = 10 µm) magnifications, with well-defined dentinal tubules. Arrowheads indicate the DEJ.
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Journal of Dental Research, Vol. 83, No. 5,
378-383 (2004)
DOI: 10.1177/154405910408300505
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