Figure 6. Extracellular actin-induced TotM expression requires Shark and Src42A in the fat body.

(a)TotM expression levels in flies that lack Shark ubiquitously (Tub-Gal80^ts; Tub-Gal4 > UAS-Shark IR R2/Fr) or in control flies lacking a driver (w¹¹¹⁸ > UAS-Shark IR R2/Fr), 24 hr after injection with either buffer or actin. Data are representative of three independent experiments with 5–10 flies/sample with duplicate samples. (b) Flies lacking Shark selectively in the fat body (c564-Gal80^ts > UAS-Shark IR R2/Fr) or control flies lacking a driver (w¹¹¹⁸ > UAS-Shark IR R2/Fr) were injected with either buffer or actin. Relative TotM levels 24 hr post injection are depicted. Data are representative of two independent experiments with 5–10 flies/sample and triplicate samples. (c) The relative TotM expression in a constitutive fat body driver line (r4-Gal4) crossed to control (w¹¹¹⁸) or UAS-Shark IR (R2/R3), 24 hr after injection with actin. Data are representative of three independent experiments with 5–10 flies/sample and duplicate samples. (d) Relative TotM levels in flies lacking Src42A either ubiquitously (Tub-Gal80^ts > UAS-Src42A IR) or selectively in the fat body (c564-Gal80^ts > UAS-Src42A IR) compared to control flies lacking a driver (w¹¹¹⁸ > UAS-Src42A IR), 24 hr after injection with either buffer or actin. Data are representative of three independent experiments with 5–10 flies/sample with duplicate samples. (e) Relative TotM levels in flies expressing a dominant negative version of Src42A ubiquitously (Tub-Gal80^ts > UAS-Src42A^DN), within the fat body (c564-Gal80^ts > UAS-Src42A^DN) or in the absence of a driver (w¹¹¹⁸ > UAS-Src42A^DN), 24 hr after injection with either buffer or actin. No driver control refers to Tub-Gal80^ts; UAS-Src42A^DN/TM6C.Sb¹. No UAS control refers to c564-Gal4; Tub-Gal80^ts/TM6C.Sb¹. Data are representative of two independent experiments with 5–10 flies/sample with triplicate samples. (f) Relative TotM levels between untreated (-) fat body driver line crossed to constitutively active Src42A (c564-Gal80^ts > UAS-Src42A^CA); untreated control flies (c564-Gal80^ts > w¹¹¹⁸) and actin-injected control flies (c564-Gal80^ts > w¹¹¹⁸). Data are representative of three independent experiments with 5–10 flies/sample with triplicate samples. TotM relative levels were calculated using the housekeeping gene Rp49 as a reference gene. Bars represent mean ± SEM. Statistical analysis was performed using one-way (c, f) or two-way (a, b, d, e) ANOVA with Sidak’s multiple comparison test as post-test for pairwise comparisons. Results of Sidak’s multiple comparison test are shown (ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001). All data points are plotted even where no bars are visible.

DOI: http://dx.doi.org/10.7554/eLife.19662.011

Figure 6—figure supplement 1. Extracellular actin-induced TotM expression is independent of Draper.

(a) Control (w¹¹¹⁸) fruit flies or Draper null mutants (drpr^Δ5) were injected with buffer or actin. The relative expression of TotM was measured 24 hr post injection. The data are representative of two independent experiments with 5–10 flies/sample and duplicate samples. (b, c) Draper was ectopically expressed in a deficient background, either by use of a heat shock driver (w; UAS-drpr/Hs-Gal4; drpr^Δ5) or two different fat body drivers (w; UAS-drpr/c564-Gal4; drpr^Δ5or w; UAS-drpr/FB-Gal4; drpr^Δ5). Relative TotM (b) or drpr (c) expression levels were measured 24 hr post actin injection and compared to a control line that lacked a driver (w; UAS-drpr/+; drpr^Δ5). The data are representative of three independent experiments with 5–10 flies/sample and duplicate samples. (d) The relative expression of TotM in flies in which draper was knocked down in the fat body (Tub-Gal80^ts; c564-Gal4 > UAS-drpr IR), measured 24 hr post actin injection and compared to control flies (Tub-Gal80^ts; c564-Gal4 > w¹¹¹⁸). Data are representative of two independent experiments with 5–10 flies/sample and triplicate samples. TotM relative levels were calculated using the housekeeping gene Rp49 as a reference gene. Bars represent mean ± SEM. Statistical analysis was performed using one-way (b, c, d) or two-way (a) ANOVA with Sidak’s multiple comparison test as post-test for pairwise comparisons. Results of Sidak’s multiple comparison test are shown (ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001). All data points are plotted even where no bars are visible.

DOI: http://dx.doi.org/10.7554/eLife.19662.012