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. 2014 Nov;34(22):4165–4176. doi: 10.1128/MCB.00722-14

FIG 5.

FIG 5

PKA phosphorylates and regulates ATGL-1 functions. (A) Potential PKA phosphorylation site prediction results obtained using the GPS2.1 software were sorted according to the scores indicated. (B) In vitro kinase assay with wild-type (WT) ATGL-1 and S303A ATGL-1 recombinant proteins. Purified GST-fused ATGL-1 proteins were incubated with the PKA catalytic subunit and 32P-labeled ATP before SDS-PAGE. (C) Confocal images of atgl-1(hj67) worms after L4440 control and kin-2 RNAi. (D and E) Representative images and quantitation of ATGL-1::GFP intensities of atgl-1(hj67) worms under control or forskolin (FSK; 100 μM) treatment. (F) Quantitation of Western blots of C. elegans ATGL-1 proteins in Cos-1 cells with or without forskolin (50 μM, 3 h) treatment. (G) Western blot analysis of mammalian ATGL protein levels in differentiated 3T3-L1 adipocytes treated with forskolin (20 mM) for the indicated time periods. The GAPDH protein was used as a loading control. Scale bars, 20 μm. White lines indicate the boundaries of worm bodies. Error bars represent standard deviations. **, P < 0.01.