Gal4/UAS mediated RNA interference in <i>Drosophila.</i>

<p>(A) Schematic representation of <i>Drosophila</i> phosphatidylinosytol 3 kinase class III (PI3K(III)) composed of dVps34 (a catalytic subunit, CG5373) and dVps15 (an adaptor subunit, CG9746). PI3K C2 domain, PI kinase conserved domain (PIK domain), PI3K catalytic domain, serine threonine protein kinase domain, WD 40 domain and N-terminal myristoylation are indicated. Underlining indicates regions used to construct dsRNA inducible vectors. (B) dVps34 and dVps15 mRNA abundance were reduced by transfection of the respective dsRNA (+) in S2 cells. RNAs was quantified by RT-PCR. γ-tubulin was used as a control. (C) Adult wings in which dsRNAs for either <i>dVps34</i> (dVps34 RNAi) or <i>dVps15</i> (dVps15 RNAi) were expressed by the <i>sd</i>-Gal4 driver. The severe malformation was completely rescued by co-expression of wild-type cDNA with dsRNA for <i>dVps34</i> (dVps34 Rescue), compared with the wild-type wing (Control). (D) Confocal immunodetection showing XY and XZ sections in the plane of pupal wings at 32 h APF. Double-headed arrows indicate the regions where only GFP-2xFYVE, or both GFP-2xFYVE and dsRNA for <i>dVps15</i>, were expressed using the <i>dpp</i>-Gal4 driver. GFP-2xFYVE (green) was localized to dot-like structures in the wild type but dispersed in the cytoplasm in the <i>dVps15</i>-knockdown cells. Wing hairs stained by rhodamine phalloidin (red) were pointed and orderly in the wild type but deformed and irregularly pointed in the <i>dVps15</i>-knockdown cells. The scale bars represent 10 µm.