SLP-2 genetically interacts with Parkin and rescues parkin mutant
phenotypes when overexpressed in the Drosophila model.
(A) Mitochondrial morphology visualized by mitoGFP transgene driven by
IFM-Gal4 in adult indirect flight muscles of wild type
(w1118) and parkin null
(park25) flies alone or in combination with
Drosophila SLP-2 (dSLP-2) or human SLP-2 (hSLP-2) overexpression
transgenes. Indirect flight muscles from thoraces of 3-days old flies were dissected
to record mitoGFP pattern. Scale bar = 50 μm. (B) Measurements of ATP
levels from thoracic muscles of 5-days old flies expressing transgenes in all tissues
under the control of tub-Gal4. The relative levels of ATP were determined by dividing
the luminescence by the total protein concentration and normalized to wild-type flies.
(C) Flight ability of wild type (w1118),
parkin and SLP-2 RNAi fly lines with or without SLP-2 transgenes expressed under the
control of tub-Gal4. 1-week old flies were used to perform the flight assay. A minimum
of 100 flies were tested each time in two independent experiments. Flight score of 6
indicates a wild type flight function, score of 0 represents a flightless phenotype.
(D) Quantification of abnormal wing posture phenotype in wild type,
parkin and SLP-2 RNAi flies with or without SLP-2 transgenes. The
percentage of flies displaying abnormal wing posture was determined in 2-weeks old F1
males. (E) Quantification of DA neurons in PPL1 cluster. The numbers of
DA neurons in the PPL1 cluster visualized by TH > StingerGFP on the posterior side
of the brain were counted. (F) Measurement of motor function using
climbing assay. 4-weeks old males aged at 25 °C were subjected to climb an 8 cm mark,
and the percentage of flies that crossed this mark in 10sec was determined. A minimum
of 100 flies were tested in repeated trails as described in the methods section.
Transgenes were expressed under the control of TH-Gal4. Statistical differences were
calculated by one-way ANOVA followed by Tukey’s post hoc test to
correct for multiple comparisons. Data is plotted as ± SEM.
***P ≤ 0.001, **P ≤ 0.01,
*P ≤ 0.05.