Fig. 8: Vaha functions as a DAG lipase and dietary supplementation of TAG, DAG, or MAG increases insulin secretion from IPCs. | Nature Communications

Fig. 8: Vaha functions as a DAG lipase and dietary supplementation of TAG, DAG, or MAG increases insulin secretion from IPCs.

From: A nutrient responsive lipase mediates gut-brain communication to regulate insulin secretion in Drosophila

Fig. 8

a Silver stained SDS-PAGE gel showing purification steps. Lane 1: 50% ammonium sulfate precipitate of culture supernatant from Vaha V5 stable cells, lane 2: nickel affinity chromatography, lane 3: gel filtration by FPLC. Western blot of the purified protein with V5 antibody. n = 2. b–d Purified Vaha protein is incubated with increasing concentrations of (b)TAG (c) DAG or (d) MAG (cold + radiolabeled) as a lipid substrate and the lipase activity is determined by measuring the released fatty acid. The substrate concentration curve is used to calculate Km, kcat and kcat/Km. For each concentration, 3 independent replicates are performed. Data are presented as mean ± SD. e, f DAG (e) or MAG levels estimated in fly homogenates of 30(f) 35-day-old control and vaha mutant by LC-MS/MS (f). n = 3, 100 flies per replicate. Data assessed by two-tailed Student’s t-test and presented as mean ± SD. g Quantification of ILP2HF fluorescence intensity in the IPCs of control (w-; +/+; gd2HF/gd2HF) flies, and fed either TAG, DAG, or MAG. Each data point represents fluorescent intensity in one IPC, n = 35 (control), n = 39 (TAG), n = 39 (DAG), n = 40 (MAG). Data assessed by One-way ANOVA followed by Tukey’s multiple comparison test and horizontal line indicates median. h Quantification of ILP2HF fluorescence intensity in the IPCs of control (w-; +/+; gd2HF/gd2HF) and vaha mutant flies (w-; vaha/vaha; gd2HF/gd2HF) fed either TAG, DAG, or MAG. Each data point represents fluorescent intensity in one IPC, n = 47 (control TAG), n = 45 (control DAG), n = 46 (control MAG), n = 47 (mutant TAG), n = 44 (mutant DAG), n = 48 (mutant MAG). Data assessed by One-way ANOVA followed by Tukey’s multiple comparison test and horizontal line indicates median. i A model of Vaha-mediated inter-organ communication in ILP2 release. Vaha (dark blue circles) secreted from R4 region of the midgut (light blue) concentrates in IPCs (green) in a process requiring LTP (black circles) from the fat body (yellow) to regulate ILP2 release. Source data are provided for 8b–h as a source data file.

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