Root is essential for neuron-specific behaviors. (A) GAL4 drivers used for Root rescue and their expression patterns. (B) Root is essential for negative geotaxis. The percentages of Root66 mutant flies that passed the negative geotaxis assay are significantly lower compared with controls (white bars). The mutants are rescued, to different degrees, by expressing GFP-Root with different GAL4 drivers. Expression in the entire nervous system (elav, insc) or mainly in the ChOs (tilB+nan, JO15-2) conferred complete or significant rescue, whereas expression limited to the central nervous system (CNS; wor) did not. (C) Root66 larvae lack sensitivity to touch, compared with the controls (white bars). Touch sensitivity is best restored by expressing GFP-Root in both ChOs and EsOs (elav, insc), expression limited to mainly ChOs (tilB+nan) also rescues the defect significantly, but expression of GFP-Root in CNS (wor) did not rescue the phenotype. (D) Root66 flies show reduced taste responses to sucrose compared with Root66/TM6B. The proboscis extension reflex (PER) taste response is significantly restored by driving GFP-Root expression in the entire PNS (elav, Insc), although the rescue was more thorough in females than in males. Significance was measured between Root66 and rescue groups at each sucrose concentration. (E) Root66 flies show significantly reduced hearing response indicated by SEPs, compared with Root66/TM6B (white bars). GFP-Root expression driven by elav-GAL4 rescues the hearing impairment of Root66 significantly but not completely. (F) In the JO, ectopically expressed GFP-Root localizes to the normal-sized, ∼15- to 25-µm rootlets in Root66/TM6B (arrows); however, the GFP-Root rescue in the Root66 mutant organizes much shorter rootlets at ∼2–8 µm (arrowheads). Bars, 10 µm. (G) Root66 males have impaired fertility. More than 90% of the control males are fertile (white bars); in contrast, none of Root66 and less than 40% of Root66/Df males produce progeny. UAS-GFP-Root; Root66 showed some fertility, probably because of leaky expression of the transgene. The fertility of Root66 males is fully rescued by expressing GFP-Root in the nervous system with elav-, Insc-, or tilB+nan-GAL4. (H) Root66 males produce mature sperm with normal tail length. For all charts, numbers of flies/larvae/sperm assayed are indicated in/near the bars, and error bars represent SEM. ns, P > 0.05; *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001. See also Video 1 and Fig. S2.
