Conservation of shibire and RpII215 temperature-sensitive lethal mutations between Drosophila and Bactrocera tryoni

Abstract

The sterile insect technique can suppress and eliminate population outbreaks of the Australian horticultural pest, Bactrocera tryoni, the Queensland fruit fly. Sterile males mate with wild females that produce inviable embryos, causing population suppression or elimination. Current sterile insect releases are mixed sex, as the efficient removal of unrequired factory-reared females is not yet possible. In this paper, we assessed the known Drosophila melanogaster temperature-sensitive embryonic lethal alleles shibire (G268D, shi^ts1) and RNA polymerase II 215 (R977C, RpII215^ts) for potential use in developing B. tryoni genetic sexing strains (GSS) for the conditional removal of females. Complementation tests in D. melanogaster wild-type or temperature-sensitive genetic backgrounds were performed using the GAL4-UAS transgene expression system. A B. tryoni wild-type shibire isoform partially rescued Drosophila temperature lethality at 29°C by improving survivorship to pupation, while expressing B. tryoni shi^ts1 failed to rescue the lethality, supporting a temperature-sensitive phenotype. Expression of the B. tryoni RpII215 wild-type protein rescued the lethality of D. melanogaster RpII215^ts flies at 29°C. Overexpressing the B. tryoni RpII215^ts allele in the D. melanogaster wild-type background unexpectedly produced a dominant lethal phenotype at 29°C. The B. tryoni shibire and RpII215 wild-type alleles were able to compensate, to varying degrees, for the function of the D. melanogaster temperature-sensitive proteins, supporting functional conservation across species. Shibire and RpII215 hold potential for developing insect strains that can selectively kill using elevated temperatures; however, alleles with milder effects than shi^ts1 will need to be considered.

Keywords: RNA polymerase II 215; embryo lethality; shibire; temperature sensitivity; transgenic complementation test.

Figure 1

Expressing Bactrocera tryoni shi wild-type (UAS-Qshi⁺ ) and mutant (UAS-Qshi^ts1 ) transgenes driven by the shi^ts2; nSyb-GAL4/TM6B driver. In each standard food vial, F₀ crosses were set up by placing three males that were hemizygous for the temperature-sensitive shi^ts2 mutant allele on the X chromosome, heterozygous for nSyb-GAL4, and balancer on the third chromosome (shi^ts2/Y; +/+; nSyb-GAL4/TM6B) and five females of either control wild type (w¹¹¹⁸ ) ( i ), Qshi⁺ in the wild-type background (shi⁺; UAS-Qshi⁺ ) ( ii ), Qshi^ts1 in the wild-type background (shi⁺; UAS-Qshi^ts1 ) ( iii ), Qshi⁺ in the temperature-sensitive background (shi^ts2; UAS-Qshi⁺ ) ( iv ), Qshi^ts1 in the temperature-sensitive background (shi^ts2; UAS-Qshi^ts1 ) ( v ), or control temperature-sensitive (shi^ts2 ) ( vi ). F₀ parents were allowed to lay eggs for 24 h at 25°C, then eggs in vials were reared at either 18°C, 22°C, 25°C, or 29°C. In the F₁, chromosomes are presented in order: sex determination chromosomes, chromosome 2, and chromosome 3. F₁ flies with non-TM6B expressed UAS-Qshi⁺ or UAS-Qshi^ts1 , while TM6B flies did not express these transgenes. The number of replicates and total counted F₁ flies for each cross at each temperature were indicated. Box plots represent the interquartile range, and the median value is indicated. Error bars represent 1.5 times the interquartile range.

Figure 2

Expressing Bactrocera tryoni RpII215 wild-type (UAS-QRpII215⁺ ) and mutant (UAS-QRpII215^ts ) transgenes with the da-GAL4 driver. In each food vial, F₀ crosses were set up by placing three males that were hemizygous for the temperature-sensitive RpII215^ts mutant allele on the X chromosome, heterozygous for da-GAL4, and balancer on the third chromosome (RpII215^ts/Y; +/+; da-GAL4/TM6B) and five females of either control wild type (w¹¹¹⁸ ) (i), QRpII215⁺ in the wild-type background (RpII215⁺; UAS-QRpII215⁺ ) (ii), QRpII215^ts in the wild-type background (RpII215⁺; UAS-QRpII215^ts ) (iii), QRpII215⁺ in the temperature-sensitive background (RpII215^ts; UAS-QRpII215⁺ ) (iv), or QRpII215^ts in the temperature-sensitive background (RpII215^ts; UAS-QRpII215^ts ) (v). F₀ parents were allowed to lay eggs for 24 h at 25°C, then eggs in vials were reared at either 18°C, 22°C, 25°C, or 29°C. In the F₁, chromosomes are presented in order: sex determination chromosomes, chromosome 2, and chromosome 3. F₁ flies with non-TM6B expressed UAS-QRpII215⁺ or UAS-QRpII215^ts , while TM6B flies did not express these transgenes. The number of replicates and total counted F₁ flies for each cross at each temperature were indicated. Box plots represent the interquartile range, and the median value is indicated. Error bars represent 1.5 times the interquartile range.