Figure 1. Expression of EGFP-L10a and assays of function in clock cells.

(A–C) Expression of EGFP-L10a in a large neurosecretory cell. Nu, nucleolus; N, Nucleus; C, Cytoplasm. Staining for a nuclear protein called LARK (red signal) is used to identify the nucleus. (D) Schematic representation of the structure of the nucleolus. FC, Fibrillar Center; DFC, Dense Fibrillar Components; GC, Granular Components. GC is the location of ribosome assembly. (E) Expression pattern of EGFP-L10a in the brain and ventral ganglion using the elav-Gal4 pan-neuronal driver. (F) Expression of EGFP-L10a in all clock cells driven by tim-Gal4. (G) Restricted expression of EGFP-L10a to clock neuron but not glia using a combination of tim-Gal4 and repo-Gal80. (H) Expression of EGFP-L10a in clock cells does not disrupt normal circadian behavior. Left panels shows representative free-running actograms of control flies and flies expressing EGFP-L10a in either PDF neurons (using pdf-Gal4) or all clock cells (using tim-Gal4). Right panels show the corresponding correlograms. (I) TRAP is capable of detecting changes in mRNA translation, as assayed by changes in the translational status of Ferritin 1 Heavy Chain Homolog (Fer1HCH) mRNA in response to overexpression of the Iron Regulatory Protein (IRP). Control, w¹¹¹⁸; act5C-Gal4/tub-Gal80^ts; UAS-EGFP-L10a/+. IRP overexpression, w¹¹¹⁸; act5C-Gal4/tub-Gal80^ts; UAS-EGFP-L10a/UAS-IRP. (J) Circadian changes in the translation of period (per) and timeless (tim) mRNAs. Genotype of the flies assayed, elav-Gal4; UAS-EGFP-L10a/+. Error bar represents standard error of the mean (SEM). *p<0.01; **p<0.001 (Student's t test).