Fig. 6: Akt phosphorylates Yki in vitro and in vivo.
From: Metabolic control of progenitor cell propagation during Drosophila tracheal remodeling
a Comparative analysis of Drosophila Yorkie, human (h), mouse (m) and zebrafish (z) YAP proteins. b The GST-Yki proteins are phosphorylated by Akt. The amount of each GST–YAP protein was detected by Coomassie staining. c Akt phosphorylates GST-Yki but not GST-YkiS168A. d Akt phosphorylates Yki in vivo. Immunoprecipitation assay of FLAG-Yki expressed in fly trachea. Phosphorylated proteins were detected using antibodies that recognize phospho-Akt substrates or phospho-AMPK substrates. e Yki is phosphorylated at Ser168 by Akt in the trachea. Western blot analysis of precipitated FLAG-Yki and YkiS168A-HA with antibodies against phospho-Akt substrates. b–e Three independent experiments were repeated with similar results. f, g Akt enhances signal of YAP-SPARK in the trachea. h–m Akt attenuates Yki-dependent activation of tracheal progenitors. h, i The incorporation of EdU in the tracheal progenitors of control (h) and UAS-Akt (i). j Box plot depicts the signal of YAP-SPARK reporter in control (n = 16) and UAS-Akt-expressed trachea (n = 31). Six biologically independent experiments were performed. Data are presented as median with minima and maxima. 25th–75th percentile (box) and 5th–95th percentile (whiskers) as well as outliers are indicated in the box plots. p = 9.46e-12. k Bar graph showing the number of EdU incorporation in control (n = 17) and UAS-Akt-expressed trachea (n = 8). Three biologically independent experiments were performed. Data are presented as mean values ± SD. ****p = 6.38e-5. l–m” Expression of UAS-Akt suppresses the migration of tracheal progenitors. n Scatter plot showing the velocity of migrating progenitors in control (n = 8; p = 4.69e-5) and UAS-Akt-expressed trachea (n = 8; p = 4.53e-4). Eight biologically independent experiments were performed. Data are presented as mean values ± SD. j, k, n Unpaired two-tailed t-test was used for all statistical analyses. No adjustments were made for multiple comparisons. Scale bars: 100 μm (f, g), 30 μm (h, i) and 300 μm (l–m”). Genotypes: (d) btl-Gal4/+; UAS-FLAG-yki/+; (f) btl-Gal4/+; UAS-YAP-SPARK/tub-Gal80ts; (g) btl-Gal4/UAS-Akt; UAS-YAP-SPARK/tub-Gal80ts; (h, l–l”) btl-Gal4/+; P[B123]-RFP-moe/ tub-Gal80ts; (i, m-m”) btl-Gal4/UAS-Akt; P[B123]-RFP-moe/tub-Gal80ts. Source data for (j, k, n) are provided as a Source Data file.
