fly, the human tumor cell lines of the cancer screening panel were grown in RPMI 1640 medium containing 5% FBS and 2 mM Lglutamine. For a typical screening experiment, cells were inoculated into 96 well microtiter plates in 100 ml at plating densities depending on the doubling time of individual cell lines. After cell inoculation, the microtiter plates are incubated at 37uC, 5% CO2 and 100% relative humidity for 24 h. Following incubation, aliquots of 100 ml of compound at different dilutions were added to the appropriate microtiter wells and were further incubated for 48 h. At the assay end-point, cells were fixed with trichloroacetic acid followed by Sulforhodamine B staining for cellular protein content. Sulforhodamine B absorbance was read at a wavelength of 515 nm as a measurement of cell LY341495 density. Mitochondria Isolation and Detergent Solubilization Functional mitochondria were isolated from mouse liver by differential centrifugation method. Briefly, a mouse was starved overnight before sacrificed by cervical dislocation. The liver was harvested promptly and rinsed with ice-cold mitochondria isolation buffer until blood-free. The liver was then cut into small pieces in a beaker using scissors while keeping in an ice-bath. The buffer was replaced with 5 ml of fresh isolation buffer and the liver was homogenized with a Polytron probe until smooth. The homogenate was centrifuged at 1000 g for 15 min at 4uC. The pellet was discarded and the supernatant was centrifuged at 12000 g for 15 min at 4uC to pellet the mitochondria. The mitochondria were washed twice by resuspending in 4 volumes of isolation buffer containing 16protease cocktail inhibitor. The concentration of the mitochondrial protein was determined using the Bradford method. The mitochondria were frozen in 10 mg/ml aliquot at 280uC. Detergent solubilization of the mitochondria proteins was done prior to measurement of the oxidative phosphorylation complexes activity. The NP1617 and NP3558 P-element transposons are inserted in the 2nd intron of the HDAC4 gene, 4838 and 3144 bp downstream from the 3′ end of the 2nd exon, respectively. The next closest gene is >7 kb from the P-element insertions. To 10381762 examine the expression pattern driven by the two p lines, we crossed each of them to a line harbouring UAS-CD8::GFP and UAS-Redstinger. Redstinger is a nuclear localized dsRED and CD8::GFP is a plasma membrane-targeted GFP, which together allow for visualisation of GFP in neuronal processes that surround the dsRED-filled nucleus. NP1617 drove expression primarily in the MB, with minimal expression elsewhere in the brain. The MB is a structure critical for memory formation in Drosophila. The intrinsic neurons of the MB are the Kenyon cells, of which there are 
three specific classes. The axons of the three neuronal subtypes are bundled to together to form a ventrally projecting peduncle, which then splits to form lobes. The / and ‘/’ axons both bifurcate to form the vertical and ‘ lobes and the medial and ‘ lobes, while the neuron axons form a single medial lobe. NP1617 drove expression in the / and lobes, but not the ‘/’ lobes, as shown by a lack of co-expression in these lobes with Trio, which is expressed in ‘/’ and neurons. NP3558-driven expression of nls.dsRED caused lethality at the pupal stage, therefore the expression pattern driven by NP3558 was characterised using CD8::GFP only. The expression level 3986806 achieved with this driver was much higher than NP1617, with CD8::GFP protein p
