Cytogenetic approaches to finding auditory genes.

摘要

Gene discovery is a crucial step in gaining understanding into developmental processes. In the auditory system, the majority of genes involved have been identified through genetic linkage studies, positional candidate and candidate gene approaches. Another positional method for gene discovery is to ascertain deaf individuals with balanced translocations and identify disrupted or disregulated genes at the site of chromosomal rearrangement. We report herein the use of fluorescence in situ hybridization (FISH) to map the breakpoint regions on each derivative chromosome of a balanced translocation in two deaf individuals. The first individual, designated DGAP090, has a de novo translocation between chromosomes 8 and 9, t(8;9)(q12.1;p21.3). The chromosome 9 breakpoint disrupts a known gene, methylthioadenosine phosphorylase (MTAP). As seen by immunohistochemistry, MTAP mRNA localizes to the stria vascularis in the inner ear. A knock-out mouse model for MTAP deficiency has been created and demonstrates a small, but significant level of hearing loss across all frequencies. Disruption of MTAP is predicted to lead to deafness due to the role of MTAP in metabolizing an inhibitor of polyamine synthesis.; The second individual, designated DGAP056, has a translocation between chromosomes 2 and 13, t(2;13)(p24.1;q22.3). The chromosome 2 breakpoint disrupts a hypothetical gene, FLJ21820. This gene contains an alpha/beta-hydrolase fold, a protein domain found in a number of enzyme families. As seen by in situ hybridization, FLJ21820 mRNA localizes specifically to the supporting cells in the organ of Corti as well as to cells in the spiral limbus. The expression of FLJ21820 in these important cells in the inner ear, as well as its disruption in a deaf individual, suggests that the gene plays a fundamental role in the auditory system.; Another method of gene discovery employs a tissue specific approach. Through characterization of a human fetal cochlear cDNA library, a cochlear clone of unknown function was identified to be highly expressed. Eventually, this expressed sequence became known to represent mimecan/osteoglycin. Mimecan is a member of a class of proteoglycans (PGs) called small leucine-rich proteoglycans (SLRPs) that are involved in matrix assembly, cellular growth and migration. Herein, we report our studies of observations of a significant loss at high frequencies of mimecan-deficient mice.