Identification of the gene for oral-facial-digital type I syndrome

Abstract

Oral-facial-digital type 1 syndrome (OFD1 [MIM 311200]) is transmitted as an X-linked dominant condition with lethality in males and is characterized by malformations of the face, oral cavity, and digits, and by a highly variable expressivity even within the same family. Malformation of the brain and polycystic kidneys are commonly associated with this disorder. The locus for OFD1 was mapped by linkage analysis to a 12-Mb interval, flanked by markers DXS85 and DXS7105 in the Xp22 region. To identify the gene responsible for this syndrome, we analyzed several transcripts mapping to the region and found mutations in OFD1 (formerly named "Cxorf5/71-7a"), encoding a protein containing coiled-coil alpha-helical domains. Seven patients with OFD1, including three with familial and four with sporadic cases, were analyzed. Analysis of the familial cases revealed a missense mutation, a 19-bp deletion, and a single base-pair deletion leading to a frameshift. In the sporadic cases, we found a missense (de novo), a nonsense, a splice, and a frameshift mutation. RNA in situ studies on mouse embryo tissue sections show that Ofd1 is developmentally regulated and is expressed in all tissues affected in OFD1 syndrome. The involvement of OFD1 in oral-facial-digital type I syndrome demonstrates an important role of this gene in human development.

Figure 1

A and B, Upper- and lower-digit abnormalities in a 12-year-old girl with OFD1. Note brachydactyly and syndactyly. C, Ultrasound scan of kidney of the same individual who is depicted in A and B. She had chronic renal excretory failure and polycystic kidneys: note the cysts (white arrows), up to 1 cm in diameter, located in the parenchyma of the organ.

Figure 2

Analysis of family 4. Left panel, Sequencing chromatograms showing the genomic sequence of the affected individual (P) and of wild-type (W) and mutant (M) alleles. Note the presence of double peaks in the patient’s DNA, downstream of the deletion due to the presence of both normal and out-of-frame alleles. In the wild-type allele, the 19-bp deletion is indicated by a horizontal bracket. Single alleles have been separated by cloning the PCR products. Right panel, Segregation of the 19-nucleotide deletion with the disease phenotype in family 4. After visualization of the PCR products on a 4% agarose gel, two different bands, corresponding to the wild-type (316-bp) and the mutant (297-bp) alleles, appear in the affected individuals (lanes 1 and 2), whereas only the wild-type band is visible in the unaffected individuals (lanes 3–5). In the next-to-last lane, a normal female control is depicted. In the last lane, the PCR-negative control (no DNA) is reported. The lozenge in the genealogical tree indicates an aborted fetus of unknown sex.

Figure 3

Mutation of OFD1 in family 10. The DNA-sequence fluorogram of the patient’s DNA for exon 11 displays eight double peaks that result from the presence of both the wild-type and the mutant alleles (top). After the PCR product was cloned, the wild-type allele (bottom) and the mutant allele (middle) were analyzed separately. The corresponding amino acid sequence is shown above the nucleotide sequence; the interchange of nine nucleotides results in a change of the three amino acids shown in boldface type.

Figure 4

Expression analysis of Ofd1, revealed by in situ hybridization