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Fig. 2. Akhr mutants have abnormal lipid and carbohydrate homeostasis. All experiments were performed with 1-week-old male flies unless otherwise noted. In the bar graphs data for Akhr^null flies are shown in blue and those for Akhr^rev flies in yellow. (A) Akhr^null flies have higher total body triglyceride content than Akhr^rev (ad libitum fed). Triglyceride values are averages of triplicate measurements, with corresponding standard deviations. (B) Akhr^null flies had larger lipid cells than control flies, consistent with the total body triglyceride measurements. Representative images are shown for Nile Red staining of fat body tissue from 1-week-old male Akhr^null (right panel) and Akhr^rev (left panel) flies. Scale bars, 10 µm. (C) Akhr^null flies have higher glycogen content (measurements obtained from whole body homogenates). Differences between Akhr^null and Akhr^rev flies are accentuated after 24 h of starvation (Student's t-test, ^*P<0.05). (D) Genetic rescue experiments demonstrate that glycogen levels decrease when Akhr is re-introduced in an otherwise Akhr^null background. Data from Gal4 flies are shown as white bars and those for UAS control flies as black bars; data from flies containing both Gal4 and UAS transgenes are shown in grey. Gal4 and UAS control flies have a higher glycogen content than flies that contain both transgenes (which can now express Akhr; Student's t-test, ^*P<0.05). (E) Tissue and cell size in Akhr^null and Akhr^rev were indistinguishable. Measurements for wing size, wing cell size, mesothorax size and foreleg femur length are shown. (F) Akhr^null flies are not hyperphagic. The ingestion of FD&C No. 1 blue was quantified for both fed and starved (18–24 h) 1-week-old male flies. Food intake is shown in arbitrary units which are proportional to the measured absorbance of ingested dye as per a published protocol (Libert et al., 2006). With prior starvation, Akhr^null flies had lower food ingestion during the first 30 min (Student's t-test, ^*P<0.05).

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