Evaluation of the myosin VIIA gene and visual function in patients with Usher syndrome type I.

Usher syndrome type I (USH1) is a recessively-inherited disorder consisting of retinitis pigmentosa, profound congenital deafness, and vestibular ataxia. It can be caused by mutations in at least six different loci (USH1A-1F). The gene encoding human myosin VIIA (MYO7A) is the USH1B locus. In this study, 66 unrelated patients with USH1 were evaluated for defects in MYO7A using single-strand conformation polymorphism analysis and direct genomic sequencing. Twenty-nine per cent of cases were found to have likely pathogenic MYO7A mutations. A total of 22 likely pathogenic changes were identified, 18 of which were novel. Cosegregation analysis of mutations in five available families showed that the MYO7A changes segregated with the disease in an autosomal recessive fashion. Average visual function as measured by visual acuity, visual field area, and ERG amplitude was not significantly different between the group of patients with likely pathogenic MYO7A changes and the group in which no likely pathogenic MYO7A changes were detected.

Interaction of palytoxin with red cells: structure-function studies.

Palytoxin (PTX), a potent toxin isolated from the marine soft coral Palythoa tuberculosa increases the cationic permeability of red cell membranes and inhibits the (Na+,K+)-activated ATPase, effects that are completely reversed by ouabain. It has also been shown to compete with ouabain for a binding site and it has been suggested that it binds to the Na+,K(+)-pump molecule in cells. In a search for analogues of PTX that would bind covalently and could thus be used to identify and characterize the binding site, we have used compounds which differed from PTX at one end or at both ends simultaneously. In order to determine whether these derivatives could be used to replace palytoxin, we tested their potency to induce an increased cation flux, complete with ouabain for its binding site, and inhibit the isolated, purified (Na+,K(+)-ATPase). The results obtained indicate that departures from the PTX structure at one end or at both ends simultaneously substantially decrease the ability of the compounds to increase the cationic permeability of red blood cells and to inhibit the (Na+,K(+)-ATPase). These actions were found to be completely reversed by ouabain, but the analogues are two to three orders of magnitude less potent than PTX in competing with ouabain for its binding site. These results suggest that both ends of the palytoxin molecule participate in the interactions of the toxin with its receptor and that modifications in these regions of the molecule produce significant alterations in its binding and subsequent action on red cell membranes.

Inhibition of the actions of palytoxin on the K+ efflux from red cells and on the Na/K-ATPase activity by a monoclonal antibody.

Palytoxin (PTX) binds to the Na/K pump, inhibits the (Na/K)-ATPase and forms Na and K permeable channels in human red cells. Here, we report that a monoclonal antibody raised against a derivative of PTX (Bignami, G.S. et al. (1992) Toxicon 30, 687-700) inhibits these effects. The observations are consistent with a model in which (a) the antibody binds and, thus reduces the concentration of free PTX available to react with the Na/K pump, and, (b) the PTX-antibody complex also binds to the PTX receptor on the Na/K pump but in such a way that a cation permeable channel is not formed, probably by reducing the concentration of free PTX. Using this model, we estimate that the apparent dissociation constant for the binding of PTX to antibody is 0.2 nM.