Setup for the wavelength choice test.

<p>(a) The experimental arena used for the wavelength choice test (modified from [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.ref022" target="_blank">22</a>]). Two dodecagonal clear plastic plates (top and bottom) were separated by six semicircular black spacers. A piece of filter paper was placed on the bottom plate. A hole was made in the center, through which the fly could climb into the experimental arena via the plastic tube. Six LED lights of different colors were placed on the open sides of the arena. We used UV (365 nm), VL (405 nm), BL (450 nm), GR (525 nm), OR (590 nm), and RD (660 nm) LEDs. The light-emitting surface was made of an optical diffusion filter framed with an aluminum seal. Dashed lines indicate imaginary borders, separating each LED area from the central hexagonal area. (b) Irradiance spectra of the LEDs. The integral of the spectra was adjusted so that the photon fluxes were equivalent for all six LEDs. (c, d) Example trajectories of the flies. Several trials were merged in each Fig The color of a trajectory reflects the peak wavelength of the LED that was eventually chosen by the individual. (c) Example trajectories of the flies that chose UV or VL. (d) Example trajectories of the flies that chose LEDs other than UV or VL.</p

Wavelength preference of <i>Exorista japonica</i>.

<p>(a) Unmated males, (b) mated males, (c) unmated females, and (d) mated females. For each experimental group, <i>n</i> = 50.</p

Regression coefficients and <i>AIC</i> for each model in mated males.

<p><i>ΔAIC</i> was defined as <i>AIC</i> subtracted from the minimum <i>AIC</i>. The row was sorted according to <i>AIC</i>.</p

Regression coefficients and <i>AIC</i> for each model in unmated males.

<p><i>ΔAIC</i> was defined as <i>AIC</i> subtracted from the minimum <i>AIC</i>. The row was sorted according to <i>AIC</i>.</p

Compound-eye-based modeling of wavelength preference.

<p>(a) Spectral sensitivity of the <i>Exorista japonica</i> compound eye. Mean ± standard error. (b) The relative quantum catch of the compound eye. (c-f) The compound eye quantum catch versus the choice frequency for LEDs in (a) unmated males, (b) mated males, (c) unmated females, and (d) mated females. Circles indicate the measured choice frequency, and the dashed line is the value fitted by the model (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.e002" target="_blank">Eq 2</a>) as a function of <i>Q</i>_CE. The coefficients are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.t001" target="_blank">Table 1</a>.</p

Regression coefficients in the compound-eye-based model for each experimental group.

<p>Regression coefficients in the compound-eye-based model for each experimental group.</p

Photoreceptor-based modeling of wavelength preference.

<p>(a) Spectral sensitivities of the <i>Musca domestica</i> photoreceptors ([<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.ref011" target="_blank">11</a>]; the numerical data were obtained from [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.ref028" target="_blank">28</a>]). (b) The relative quantum catch of individual photoreceptors, <i>Q</i>_i. (c-f) Linear predictor in the best photoreceptor-based model versus choice frequency in (c) unmated males, (d) mated males, (e) unmated females, and (f) mated females. Circles indicate the measured choice frequency, and the dashed line is the value fitted by the best model as a function of the linear predictor η. The coefficients are shown in Tables <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.t002" target="_blank">2</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160441#pone.0160441.t005" target="_blank">5</a>.</p

Generation of transgenic flies carrying hGBA variants.

<p>(A) Sequence of hGBA. Blue and red fonts show R120W and RecNciI mutations, respectively. (B) Expression levels of hGBA mRNA confirmed by quantitative RT-PCR (n  =  about 30 fly heads per transgenic combination) with dRpL32 as internal control. Error bars represent SE. (C) Levels of hGBA protein confirmed by Western blotting (n  =  about 100 fly heads per transgenic combination). Total amounts of hGBA protein were decreased in hGBA^R120W, and significantly decreased in hGBA^RecNciI transgenic combinations, compared with hGBA^WT transgenic combination.</p

Feeding of ambroxol ameliorates neurodevelopmental defects and ER stress in the mutated hGBA induced <i>Drosophila</i> eye.

<p>Ambroxol can recover morphological defects and decrease ER stress in transgenic flies. (A) Less fluorescence emitted by the eye imaginal discs of hGBA^RecNciI transgenic combinations treated with, than without 1 mM Ambroxol. (B) Values generated by different transgenic combinations at fixed quantities of fluorescence intensity (n = 12–43 eye imaginal discs of third instar larvae per transgenic combination). Error bars represent SE. *Significant difference compared with controls (all without Ambroxol) (***P<0.001; Student's t test). (C) Ambroxol (1 mM) decreases expression levels of dBiP mRNA in the heads of hGBA^RecNciI transgenic combinations (n  =  about 30 fly heads per transgenic combination). Internal control was dRpL32. Error bars represent SE. (D) Eye phenotypes of hGBA^RecNciI transgenic combinations incubated without or with 1 mM Ambroxol. Size and shape of ocelli were uniform, and layout uniformity was more similar to that of normal fly eyes treated with 1 mM Ambroxol. (E) Size histograms of ocelli in hGBA^RecNciI transgenic combinations treated with or without 1 mM Ambroxol. (n = 6–10 flies per transgenic combination; about 400 ocelli each). Dispersion analysis showed significant differences from hGBA^RecNciI transgenic combinations treated with and without 1 mM Ambroxol (F = 2.07–3.35; P<0.001; Levene's test).</p

Neurodevelopmental defects in the <i>Drosophila</i> eye caused by expression of hGBA carrying the RecNciI mutation.

<p>We investigated the effects of overexpression to mutated hGBAs in fly eyes. (A) Phenotype of eyes overexpressing hGBA^WT transgenic combination do not significantly differ from those of GMR control. Phenotype of eyes overexpressing hGBA^R120W transgenic combinations occasionally differed in terms of morphology in some flies compared with control. Eye morphology is obviously affected in hGBA^RecNciI transgenic combinations compared with control. (B) Size histograms of ocelli in transgenic combinations (n = 3–5 flies each, about 100 ocelli each). Dispersion analysis showed obvious differences in variance of the sizes of ocelli between the hGBA^RecNciI transgenic combinations and the GMR control (F = 29.50–37.19; P<0.001; Levene's test).</p