Myosin VIIa is the product of the gene USH1B, the locus of genetic mutations in Usher syndrome type 1B. Features associated with the syndrome include congenital deafness and gradual blindness. The normal protein is expressed in hair cells of the vestibular system and cochlea, in photoreceptors, retinal pigment epithelium (RPE) cells, and in kidney and testis. In photoreceptors and hair cells, it may be involved in constructing and maintaining their different cytoskeletal specializations. Myosin VIIa is the first cloned gene with a demonstrated functional requirement in hair cells; an understanding of how its expression is controlled could reveal transcriptional controls that operate more generally in hair cell and/or retinal ell differentiation. The investigator aims to define the regulatory elements that specify the tissue-specific expression of myosin VIIa. Using deletion analysis of the putative promoter region, the primary focus is to define promoter elements that are required for transcription. Regions of cloned genomic DNA encompassing some 4. 5kb upstream of and including the transcriptional start site are to be joined to a reporter gene encoding luciferase. This construct, and derivatives in which progressively larger segments have been deleted, are to be transiently transfected into a retinal pigment epithelium cell line that also expresses myosin VIIa. The luciferase activity (light output) in cell extracts is to be quantitated. Preliminary assays have demonstrated activation of reporter gene expression by the full-length construct and derivatives, but not cell type-specific expression; the same relative activation was observed in myosin-expressing an non- expressing cells. Initial studies suggest multiple start sites for transcription in testis. A combination of RT-PCR and RNAse protection assays are proposed to resolve this issue. DNA sequences required for transcription are to be screened for mutations in Usher syndrome type I families.
| Orten, D J; Weston, M D; Kelley, P M et al. (2000) Erratum: analysis of DNA elements that modulate myosin VIIa expression in humans. Hum Mutat 15:114-5 |
| Orten, D J; Weston, M D; Kelley, P M et al. (1999) Analysis of DNA elements that modulate myosin VIIA expression in humans. Hum Mutat 14:354 |
