Extended Data Figure 4: HUWE1 is required for hyperubiquitination of MFN2 in response to C18:0 withdrawal.
From: Regulation of mitochondrial morphology and function by stearoylation of TFR1
a, Short interfering RNA (siRNA) depletion of other ubiquitin ligases targeting MFN (besides HUWE1, shown in Fig. 3) does not rescue the mitochondrial fragmentation induced by removal of C18:0 (top). Bottom, quantification. n = 15. b, siRNA depletion of PARK2 in HEK293 cells, as in HeLa cells (a), does not rescue the mitochondrial fragmentation induced by removal of C18:0 (top). Bottom, quantification. n = 15. c, HUWE1 knockdown efficiency controlled by detecting HUWE1 protein levels. See Supplementary Fig. 17 for image of the uncropped full western blot. d, Survival to pupation of Elovl6 mutants is fully rescued by ubiquitous expression (daughterless-GAL4) of RNAi targeting Huwe1 (CG8184). Elovl6 mutants expressing HUWE1 RNAi survive to pupation at expected Mendelian frequencies (χ2 test, 0.86 < 3.841 = χ2, where P = 0.05). Flies were grown on antifungal-free food. Values represent average of four biological replicates. Scale bars, 10 μm. a, b, d, *P < 0.05, **P < 0.01, ***P < 0.001, not significant (NS) P ≥ 0.05, two-tailed t-test. Error bars represent s.d.
