Mutations in a novel gene, TMIE, are associated with hearing loss linked to the DFNB6 locus

Abstract

We have identified five different homozygous recessive mutations in a novel gene, TMIE (transmembrane inner ear expressed gene), in affected members of consanguineous families segregating severe-to-profound prelingual deafness, consistent with linkage to DFNB6. The mutations include an insertion, a deletion, and three missense mutations, and they indicate that loss of function of TMIE causes hearing loss in humans. TMIE encodes a protein with 156 amino acids and exhibits no significant nucleotide or deduced amino acid sequence similarity to any other gene.

Figure 1

Families showing linkage to DFNB6 and TMIE mutation analyses. A, Family I-51, from India, defined the minimum interval for DFNB6 as being between markers D3S3582 and D3S1613. Sequence chromatograms from a noncarrier (II:4) and a deaf individual (IV:2) show the wild-type sequence, the place of insertion (arrowhead), and the insertion of CGCC (bracketed) in exon 2, respectively. B, 250C→T transition in exon 3 in an affected individual (II:1) from family 5020-22, compared with the wild-type sequence. C, The chromatograms show sequence of the intron 1–exon 2 boundary in PKSN21, from a noncarrier and an affected individual with the deletion of seven bases (bracketed in wild type). The site of the deletion is indicated by an arrowhead, along with an insertion of cytosine (arrow). D, The sequence traces of a wild type and an affected individual (IV:2) in family PKSR22B with the 241C→T transition (arrows). E, Chromatograms from an unaffected control individual and an affected individual (IV:1) from family PKDF76, showing the 274C→T transition (arrows).

Figure 2

Diagrammatic representations of TMIE and TMIE. A, The boxed regions depict exons of TMIE. Slashed lines represent introns. The length of each exon and intron is given in base pairs (bp), within boxes (for exons) and below slashed lines (for introns). “ATG” denotes the initiation codon, and the asterisk (*) marks the stop codon. Mutations found in the five families showing linkage to DFNB6 are shown above the respective exons. We obtained a 1023-bp 3′ UTR followed by a poly A tail. BLAST analysis of the TMIE cDNA sequence identified ESTs corresponding to a 1202-bp 3′ UTR because of utilization of a different polyadenylation signal. B, Clustal W alignment of deduced amino acid sequences of Tmie and TMIE. The shared amino acid identity between Tmie (mouse) and TMIE (human) is indicated by shaded boxes: dark gray for absolute identity and light gray for conservative changes. Dashes (—) show gaps in the alignment. TMIE has two potential sites of signal peptide cleavage (arrows), and two predicted transmembrane regions (black bars on top of amino acid residues). Cleavage of the signal peptide would result in a protein with an extracellular amino terminus, one transmembrane segment, and an intracellular carboxy terminus. The C-terminus has many positively charged amino acids interspersed with negatively charged residues (charge indicated on top of each residue) and three potential protein kinase phosphorylation sites (bold underlines). Arrowheads indicate arginine residues in TMIE mutated in affected individuals in three families linked to DFNB6.