Extended Data Fig. 7: UGDH promotes tumour metastasis by relieving the inhibition of HuR by UDP-Glc.

a, Transwell migration assays were performed in A549 cells treated with increasing concentrations of UDP-Glc for 16 h. b–d, A549 cells stably expressing luciferase were implanted into randomized athymic nude mice by tail-vein injection (six mice per group). Then, 14 days after inoculation, mice were injected with or without UDP-Glc (200 mg per kg, every 2 days) by tail-vein injection. b, Left, 28 or 30 days after inoculation, bioluminescence imaging of the tumour-implanted mice was carried out and representative images of lung metastasis are shown. Right, the luciferase intensities of metastatic tumours in the lung were statistically analysed. The relative luciferase intensities were normalized to those of the mice implanted with A549 cells without UDP-Glc treatment at day 28. Data represent the mean ± s.d. of the luciferase intensities in six mice (two-tailed Student’s t-test). c, Top, representative images of H&E-stained sections in dissected lungs 35 days after inoculation are shown. Bottom, the metastatic nodules were quantified based on the H&E-stained lung sections. Data represent the mean ± s.d. of the metastatic nodules per mouse in six mice (two-tailed Student’s t-test). d, Kaplan–Meier survival analysis of a different group of mice (nine mice per group) implanted with tumour cells with or without UDP-Glc administration (two-tailed log-rank test). Upwards tick mark represents censored (alive at last follow-up) mice. e, f, UGP2-depleted A549 or H1299 cells were rescued with or without rUGP2. e, The expression of UGP2 was examined. f, The intracellular UDP-Glc levels were determined. g, h, A549 or H1299 cells stably expressing shNT or shUGDH were infected with or without the lentivirus expressing shUGP2. g, The expression of UGP2 and UGDH was examined. h, Transwell migration assays were performed with cells treated with EGF (100 ng ml⁻¹) for 16 h (A549 cells) or 24 h (H1299 cells). i, j, A549 cells stably expressing shNT or shUGDH were infected with or without the lentivirus expressing shUGP2. Cells were stably infected with the lentivirus expressing luciferase and then implanted into randomized athymic nude mice by tail-vein injection (six mice per group). i, Left, 25 days after inoculation, bioluminescence imaging of tumour-implanted mice was carried out and representative images of lung metastasis are shown. Right, the luciferase intensities of metastatic tumours in lung were statistically analysed. The relative luciferase intensities were normalized to those of the mice implanted with A549 cells expressing shNT. Data represent the mean ± s.d. of the luciferase intensities in six mice. j, The metastatic nodules were quantified based on the H&E-stained lung sections. Data represent the mean ± s.d. of the metastatic nodules per mouse in six mice (two-tailed Student’s t-test). k, l, ELAVL1-depleted A549 or H1299 cells rescued with rHuR(WT) or rHuR(N134A) were infected with or without the lentivirus expressing shUGDH. k, The expression of HuR and UGDH was examined. l, Transwell migration assays were performed with cells treated with EGF (100 ng ml⁻¹) for 16 h (A549 cells) or 24 h (H1299 cells). e, g, k, Immunoblotting experiments were performed with the indicated antibodies. Data are representative of at least three independent experiments. a, f, h, l, Data represent the mean ± s.d. of three biologically independent experiments (two-tailed Student’s t-test).

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