Fig 1.
JHAMT-Gal4 driver directs expression in the corpora allata (CA).
Frozen sections of adult JHAMT-Gal4/UAS-lacZ males were incubated with an anti-ßGal antibody (red) and with an anti-JHAMT antibody (green). Overlapping expression in the CA (marked by arrow) was observed. H:Head; T:Thorax
Fig 2.
Expression of JHAMT-RNAi lowers the courtship index and can be rescued by application of the JH analog Methoprene.
Graphs show the courtship index CI (fraction of time males spend courting during the observation period) ± SEM of the indicated genotypes (A, C), or the performance of males in a control activity assay (# of line crossings ± SEM) (B). Data were analyzed by ANOVA followed by Bonferroni multiple comparisons. A) Expression of UAS-JHAMT RNAi using our CA-specific JHAMT-Gal4 driver significantly reduces male courtship in comparison to the control males. N = 20. (p < 0.001). Genotypes used were UAS-dicer/Y; UAS-JHAMT-RNAi/+; JHAMT-GAL4/+; and control genotypes UAS-dicer/Y; UAS-JHAMT-RNAi/+; +/+ and X/Y; +/+; JHAMT-GAL4/+. B) Activity levels in the mutants and in control flies (genotypes as in A). Mutants are not different from the normally courting X/Y; +/+; JHAMT-Gal4 control. (N = 10). C) Mature males of the genotypes described in (A) were treated with Methoprene in acetone or acetone alone, and tested four hours later. The experimental genotype shows complete rescue of the courtship index following Methoprene treatment compared to acetone treatment alone. (N = 20).
Fig 3.
Conditional reduction of JHAMT in mature males causes courtship defects that can be rescued by application of the JH analog Methoprene.
Graphs show the courtship index CI (fraction of time males spend courting during the observation period) ± SEM of the indicated genotypes (A, C), or the performance of males in a control activity assay (# of line crossings ± SEM (B). Data were analyzed by ANOVA followed by Bonferroni multiple comparisons. (A) Courtship index of induced and un-induced experimental (X/Y; UAS-JHAMT-RNAi/+; hsp70-GAL4/+) and control (X/Y; +/+; hsp70-GAL4/+; and X/Y; UAS-JHAMT-RNAi/+;+/+) genotypes. The experimental genotype shows a significant reduction in courtship index (p = 0.0007). (N = 20). Induced flies were heat-shocked at 37°C for 1 hour and let recover for four hours. (B) Activity assay of the induced and un-induced genotypes; genotypes as in (A) (n = 10). (C) Genotypes as in (A); one hour after induction, flies were treated with Methoprene in acetone, or with acetone alone, and courtship was examined 4 hours later. The experimental genotype shows complete rescue (p< 0.003) (N = 20). (Un-induced (-), Induced (+).
Fig 4.
Conditional adult ablation of the corpora allata (CA) causes courtship defects that can be rescued by application of the JH analog Methoprene.
The conditional Gal80ts system was used to express UAS-DTI in adult males. The flies were created and maintained at 18°C. For induction, mature males were placed at 30°C for 2 days and then kept at room temperature for one day. The induced and un-induced experimental and control genotypes were subjected to courtship assay. Graphs show the courtship index CI (fraction of time males spend courting during the observation period) ± SEM of the indicated genotypes (A, C), or the performance of males in a control activity assay (# of line crossings ± SEM (B). Data were analyzed by ANOVA followed by Bonferroni multiple comparisons. (A) Courtship assay of induced and un-induced experimental X/Y; Gal80ts/Gal80ts; JHAMT-GAL4/ UAS-DTI and control genotypes X/Y; Gal80ts/+; UAS-DTI/ +; and X/Y; Gal80ts/+; JHAMT-GAL4/+. (N = 15). Experimental flies had significantly lower courtship than the controls (p<0.01). (B) Activity assay of the flies described in (A) (N = 10). (C) Methoprene in acetone or acetone alone was applied to induced males four hours prior to testing for courtship. Methoprene application completely rescued the courtship defect of experimental flies (P<0.001). (N = 20). (Un-induced (-), Induced (+).