A Locus for Autosomal Recessive Hypodontia with Associated Dental Anomalies Maps to Chromosome 16q12.1

To the Editor: Hypodontia is characterized by the congenital absence of one or more teeth and may include deciduous and/or permanent dentition. The permanent teeth most frequently missing are second premolars, maxillary lateral incisors, and third premolars (Schulze, 1970Schulze C Developmental abnormalities of the teeth and jaws.in: Gorlin RJ Goldman HM Thoma's oral pathology. CV Mosby, St Louis1970: 96-183Google Scholar). Congenitally missing deciduous teeth are uncommon, but, when occurring, usually involve the maxillary lateral incisors, mandibular lateral incisors, and mandibular cuspids (Grahnen and Granath, 1961Grahnen H Granath LE Numerical variations in primary dentition and their correlation with the permanent dentition.Odont Rev. 1961; 12: 348-357Google Scholar). Segregation analyses in many family studies have suggested that hypodontia is a single-gene defect, often transmitted as an autosomal dominant trait with incomplete penetrance and variable expressivity (Burzynski and Escobar, 1983Burzynski NJ Escobar VH Classification and genetics of numeric anomalies of dentition.Birth Defects. 1983; 19: 95-106PubMed Google Scholar; Svinhufvud et al., 1988Svinhufvud E Myllarniemi S Norio R Dominant inheritance of tooth malpositions and their association to hypodontia.Clin Genet. 1988; 34: 373-381Crossref PubMed Scopus (52) Google Scholar). In this study, we report a consanguineous kindred from Pakistan, with hypodontia associated with various dental anomalies, transmitted as an autosomal recessive trait. Clinical investigation of the affected family members demonstrated hypodontia associated with dental anomalies such as malformation, enamel hypoplasia, and failure of eruption of teeth, leading prematurely to the edentulous state. To localize the gene responsible for the disease, we performed genomewide screening with a panel of 386 markers spaced at 10-cM intervals. We demonstrated linkage to a 10-cM region on chromosome 16q12.1, with a maximum two-point LOD score of 5.76 for marker D16S3140. Tooth development begins as an epithelial bud, which undergoes complex morphogenesis and is regulated by interactions between the epithelial and mesenchymal tissue layers. The first step in tooth development is the migration of mesenchymal cells from the neural crest into the tooth-forming zone of the vertebrate jaw, known as the “maxillary and mandibular arches.” When the mesenchymal cells arrive, they induce the overlying dental epithelium to thicken, forming the dental lamina. The epithelium subsequently invaginates into the mesenchyme to form the dental organ, around which the mesenchyme then proliferates and condenses into the dental papilla. Together, the dental organ and the dental papilla form the tooth germ (also known as the “tooth organ”; Thesleff, 1996Thesleff I Two genes for missing teeth.Nat Genet. 1996; 13: 379-380Crossref PubMed Scopus (31) Google Scholar; Lewin, 1997Lewin DI Evolution: mammalian tooth development.J NIH Res. 1997; 9: 75-80Google Scholar). Secreted signaling molecules that transmit the sequential and reciprocal inductive interactions between the epithelium and mesenchyme have been identified, and transcription factors that are needed for the signaling cascades (Thesleff, 1996Thesleff I Two genes for missing teeth.Nat Genet. 1996; 13: 379-380Crossref PubMed Scopus (31) Google Scholar), such as the homeobox genes (MSX) 1 and 2, the epidermal growth factor (EGF) and its receptor (EGFR), and transforming growth factor (TGF) B1 (Vainio et al., 1993Vainio S Karavanova I Jowett A Thesleff I Identification of BMP-4 as a signal mediating secondary induction between epithelial and mesenchymal tissues during early tooth development.Cell. 1993; 75: 45-58Abstract Full Text PDF PubMed Scopus (790) Google Scholar), are now known. Recent studies (Vaahtokari et al. Vaahtokari et al.Vaahtokari A Aberg T Jernvall J Keranen S Thesleff I The enamel knot as a signaling center in the developing mouse tooth.Mech Dev. 1996a; 54: 39-43Crossref PubMed Scopus (297) Google Scholar,Vaahtokari et al.Vaahtokari A Aberg T Thesleff I Apoptosis in the developing tooth: association with an embryonic signaling center and suppression by EGF and FGF-4.Development. 1996b; 122: 121-129PubMed Google Scholar) have demonstrated that the enamel knot, the transient cluster of epithelial cells in the center of the developing tooth germ, may represent a major signaling center regulating tooth-shape development by expressing growth factors such as bone morphogenetic proteins (BMP) 2, 4, and 7, sonic hedgehog, and fibroblast growth factor 4. Yet, despite a large body of scientific investigation, only one mutation in a single gene, MSX1, has been identified, in one family with an autosomal dominant form of hypodontia (Vastardis et al., 1996Vastardis H Karimbux N Guthua SW Seidman JG Seidman CE A human MSX1 homeodomain missense mutation causes selective tooth agenesis.Nat Genet. 1996; 13: 417-421Crossref PubMed Scopus (518) Google Scholar). To search for the gene defect in hypodontia, we studied a single highly consanguineous family with an autosomal recessive form of hypodontia and other dental anomalies (fig. 1). The family described in this report resides in Sind, Pakistan. The family members rarely marry outside the family, and consequently consanguineous unions are common. The pedigree was drawn by interviewing the elders of the family. Information was checked and rechecked by interviewing different persons. Living affected and unaffected subjects were clinically examined at Abbassi Shaheed Hospital, Karachi, Pakistan, and blood samples were collected, with informed consent, at the local hospital. Clinical status was based on dental history, intraoral examination, and panoramic radiographs. Dental examinations of the affected subjects in our family demonstrated a range of dental anomalies involving several maxillary and mandibular teeth. The main clinical problems encountered were delays in the formation and eruption of the teeth (fig. 2). The skin, hair, nails, and sweat glands were normal, and no evidence for an ectodermal dysplasia was observed. The pedigree (fig. 1) provided convincing evidence for an autosomal recessive mode of inheritance, and consanguineous loops accounted for all the affected persons being homozygous for the mutant allele. To elucidate the gene defect in the family, we initially searched for linkage by using polymorphic markers within several candidate genes—including MSX1 (4p16), MSX2 (5q34–35), EGF (4q25–27), and EGFR (7p12–14)—previously reported to be involved in early tooth morphogenesis (Partanen et al., 1985Partanen A-M Ekblom P Thesleff I Epidermal growth factor inhibits morphogenesis and cell differentiation in cultured mouse embryonic teeth.Dev Biol. 1985; 111: 84-94Crossref PubMed Scopus (84) Google Scholar; Kronmiller et al., 1991Kronmiller JE Upholt WB Kollar EJ EGF antisense oligodeoxynucleotides block murine odontogenesis in vitro.Dev Biol. 1991; 147: 485-488Crossref PubMed Scopus (64) Google Scholar; Mackenzie et al. Mackenzie et al., 1991Mackenzie A Leming GL Jowett AK Ferguson MWJ Sharpe PT The homeobox gene Hox 7.1 has specific regional temporal expression patterns during early murine craniofacial embryogenesis, especially tooth development in vivo and in vitro.Development. 1991; 111: 269-285PubMed Google Scholar, Mackenzie et al., 1992Mackenzie A Ferguson MWJ Sharpe PT Expression patterns of the homeobox gene, Hox 8, in the mouse embryo suggest a role in specifying tooth initiation and shape.Development. 1992; 115: 403-420PubMed Google Scholar). When linkage to these candidate genes was excluded, we undertook a genomewide search.Figure 2A, Subject V-9, a 15-year-old male, showing mixed dentition, with the first permanent molars, maxillary central incisors, and mandibular right cuspid erupted. In addition, the maxillary right lateral incisor and canine and the mandibular incisors and left canine from primary dentition were present. Maxillary central incisors had erupted only halfway and were widely spaced. The mucosa and other oral structures appeared normal in color and texture. B, Panoramic radiograph showing a number of unerupted teeth, including the maxillary lateral incisors, cuspids, bicuspids, and second molars and the mandibular left cuspid and bicuspids, right bicuspids, and second molars. All teeth, representing various stages of development, were malpositioned and malformed, showing only crown formation. Maxillary and mandibular last molars and left second molars were completely missing. Of the teeth that had erupted, the maxillary central incisors had open apices with wide pulp chambers and a loss of distinction between the chamber and the canal. The erupted maxillary and mandibular permanent molars had tapering cone-shaped fused roots curved toward the apices, and their pulp chambers and canals were wide, merging together, with loss of detail. C, Subject V-10, a 14-year-old male, showing the presence of maxillary central incisors and first molars and of mandibular incisors and first molars. The only deciduous tooth present was the maxillary right canine. D, Panoramic radiograph showing the presence of a number of unerupted teeth, including the maxillary lateral incisors, cuspids, bicuspids, and second molars and the mandibular cuspids, bicuspids, and second molars. The widely spaced maxillary central incisors had wide pulp chambers and canals with open apices. The pulp canals of the mandibular incisors were distorted, with complete loss of detail. All permanent first molars had fused roots and open apices and showed little distinction between the pulp chamber and the canal. E, Upper jaw of subject IV-17, a 38-year-old male, containing only six teeth, including the central incisors, canines, right second bicuspid, and right first molar with roots visible above the mucosal level. The lower jaw showed only nine erupted teeth, including the central incisors, right lateral incisor, right cuspid, both second bicuspids, and first molars. F, Panoramic radiograph showing a few malformed unerupted teeth, which might have corresponded to the maxillary left bicuspids, mandibular right and left canines, and left first bicuspid.View Large Image Figure ViewerDownload Hi-res image Download (PPT) An initial genomewide screen with microsatellite markers evenly spaced at 10-cM intervals was conducted by use of the DNA from three of the affected individuals (III-7, IV-12, and IV-17). The initial genomewide search involving the three affected individuals was carried out at Research Genetics, Inc. In the course of screening 386 markers, 17 genomic regions were found to be homozygous in all three affected subjects and were tested further in the rest of the affected and unaffected members of the family. A marker on chromosome 16q12.1 (D16S3112) was found to be homozygous in only the affected individuals. Further analysis with markers from this region resulted in the identification of homozygosity, in affected individuals, for markers D16S3112 and D16S3140. By use of the FASTLINK 3.0P package (Schaffer, 1996Schaffer AA Fast linkage analysis computations for pedigrees with loops or unused alleles.Hum Hered. 1996; 46: 226-235Crossref PubMed Scopus (104) Google Scholar), which enables retention of all inbreeding loops in the family, a two-point LOD score of 5.76 at .00 recombination was obtained for marker D16S3140 (table 1). Autosomal recessive inheritance with complete penetrance was assumed, using a disease-allele frequency of .0001. The order of the markers was derived from on-line genetic mapping data at the Genome Medical Center, University of Wisconsin (http://www.marshmed.org/genetics/). The LOD score remains significant over a range of allele-frequency estimates. Recombination events observed in individuals IV-19 and IV-18, with markers D16S492 and D16S2620, respectively, placed the disease locus in a 10-cM interval between these two markers.Table 1LOD Scores for Linkage of the Hypodontia Locus to Chromosome 16q12.1 MarkersLod Score at Recombination Fraction ofMarker.00.01.05.1.2.3.4D16S492∞3.313.613.42.651.77.86D16S31125.445.344.914.363.232.06.9D16S31405.765.655.214.653.492.311.12D16S4084.073.993.673.272.441.58.73D16S30712.642.582.352.061.47.91.4D16S2620∞3.533.83.542.71.72.76 Open table in a new tab Physical mapping data for this region include a single linked YAC contig, WI-16.4, and radiation hybrid mapping data have placed a number of expressed sequence tags (ESTs) in the region between D16S492 and D16S2620. Currently, there are no potential candidate genes for the disease; however, two ESTs show strong DNA homology to the drosophila son of sevenless gene and the Saccharomyces cerevisiae gene sequences for the general negative regulator of transcription subunit 1. Discovery of the first gene directly implicated in the pathogenesis of inherited autosomal recessive hypodontia associated with various forms of dental anomalies could generate a new direction of scientific investigation into the tooth bud as a model for genetic control of morphogenesis and development, particularly of epithelial-mesenchymal signaling interactions. We sincerely thank the family members who participated in this study. The Pakistan Science Foundation provided the support necessary for M.A. to visit the family members. This work was supported, in part, by National Institutes of Health–National Human Genome Research Institute grant HG-00008 (to J.O.) and by a grant from the National Institutes of Health–National Institute of Arthritis, Musculoskeletal and Skin Diseases, Skin Disease Research Center (to J.O. and A.M.C.).