Fig. 5: USP46 promotes melanocyte malignant transformation and tumor progression through stabilizing NRAS mutants.

a hTERT/CDK4^R24C/p53^DD human melanocytes expressing FLAG-NRAS Q61R were infected with lentivirus encoding shScramble (shScr) or shUSP46 (#1 and #2) and indicated proteins were examined by western blotting. b The colony formation ability of hTERT/CDK4^R24C/p53^DD melanocytes as in (a) was measured. c, d Cells as in (a) were subcutaneously implanted into nude mice (5–6 weeks, n = 6). Tumors were collected (c) and tumor weights were analyzed (d). e Cells as in (a) were transfected with empty vector (pLV5), USP46 WT or C44S mutant and western blotting was performed with indicated antibodies. f The colony formation assay of melanocytes as in (e) was performed. g, h Cells as in (e) were subcutaneously implanted into nude mice (5–6 weeks, n = 6). Tumors were collected (g) and weights were measured (h). i SK-MEL-103 cells stably expressing shScramble (shScr) or shUSP46 (#1 and #2) were transfected with empty vector (pLV3) or FLAG-NRAS Q61R and western blotting was performed with indicated antibodies. j Cell proliferation of SK-MEL-103 cells in (i) was examined. k The migration and invasion abilities of SK-MEL-103 as in (i) were measured by Transwell migration and invasion assays and results were quantified in low panel. l SK-MEL-103 cells as in (i) were treated with indicated concentrations of dacarbazine (DTIC) and cell survival was determined. m, n SK-MEL-103 cells as in (i) were subcutaneously implanted into nude mice (5–6 weeks, n = 6). When tumors reached around 150–200 mm³ in size, mice were treated with saline or DTIC (8 mg/kg). Tumors were collected (m) and weights were measured (n). The results represent the mean ± s.d. of data from six mice. Data were presented as mean ± SD of three independent experiments (b, f, j–l). Data were analyzed by two-sided one-way ANOVA in (b, d, f, h, j, k, n). Source data are provided as a Source Data file.