Sequence specificity of aminoglycoside-induced stop codon readthrough: Potential implications for treatment of Duchenne muscular dystrophy

As a result of their ability to induce translational readthrough of stop co dons, the aminoglycoside antibiotics are currently being tested for efficac y in the treatment of Duchenne muscular dystrophy patients carrying a nonse nse mutation in the dystrophin gene. We have undertaken a systematic analys is of aminoglycoside-induced readthrough of each stop codon in human tissue culture cells using a dual luciferase reporter system. Significant differe nces in the efficiency of aminoglycoside-induced readthrough were observed, with UGA showing greater translational readthrough than UAG or UAA. Additi onally, the nucleotide in the position immediately downstream from the stop codon had a significant impact on the efficiency of aminoglycoside-induced readthrough in the order C > U > A greater than or equal to G. Our studies show that the efficiency of stop codon readthrough in the presence of amin oglycosides is inversely proportional to the efficiency of translational te rmination in the absence of these compounds. Using the same assay, we analy zed a 33-base pair fragment of the mouse dystrophin gene containing the mdx premature stop codon mutation UAA (A), which is also the most efficient tr anslational terminator. The additional flanking sequences from the dystroph in gene do not significantly change the relatively low-level aminoglycoside -induced stop codon readthrough of this stop codon. The implications of the se results for drug efficacy in the treatment of individual patients with D uchenne muscular dystrophy or other genetic diseases caused by nonsense mut ations are discussed.