Mutant Mos1 mariner transposons are hyperactive in Aedes aegypti.
The development of genetic strategies to control the spread of mosquito-borne diseases through the use of class II transposons has been hampered by suboptimal rates of transformation and the absence of post-integration mobility for all transposons evaluated to date. Two Mos1 mariner transposase mutants were produced by the site-directed mutagenesis of amino acids, E137 and E264, to K and R, respectively. The effects of these mutations on the transpositional activities of Mos1-derived transposon constructs were evaluated by interplasmid transposition assays in Escherichia coli and Aedes aegypti. The transpositional activities of two Mos1 transposons, one with imperfect wild type inverted terminal repeats (ITRs) and another that contained two perfectly matched 3' ITRs, were increased when the mutant transposases were supplied in trans in E. coli. The use of the perfect repeat transposon with wild type transposase did not result in an increase in transposition frequency in Ae. aegypti. However, an improvement in the integrity of the transposition process did occur, as evidenced by a lower rate of recombination events in which the transgene was transferred. An increase in the transpositional activity of the perfect repeat transposon was observed in the mosquito in the presence of either mutant transposase, and in the case of the E264R transposase, the observed increase in transposition frequency was also accompanied by a further improvement in the integrity of transposition. We discuss the possible contributions of these mutant residues to the transposition of the perfect repeat Mos1 transposon, the implications of these results with respect to the molecular evolution of Mos1, and the potential uses of the perfect repeat transposon and mutant transposases for the improvement of Mos1 mediated germ line transformation of Ae. aegypti.
Pledger
David W
DW
Department of Biology (MSC-158), Texas A&M University, Kingsville, TX 78363, USA.
Coates
Craig J
CJ
D000330:Aedes / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004251:DNA Transposable Elements / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016366:Open Reading Frames; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D019895:Transposases
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; C111652:mariner transposases; D019895:Transposases