Fig. 3: RNF41 directly interacts with and ubiquitinates NUDC. | Cell Death & Disease

Fig. 3: RNF41 directly interacts with and ubiquitinates NUDC.

From: Loss of RNF41 promotes bladder cancer metastasis through increasing NUDC stability to enhance tubulin polymerization

Fig. 3

A Proteins interacting with RNF41 were detected using co-immunoprecipitation (co-IP) and silver staining in T24 cells. B, C Peptide fragment mass spectrum for RNF41 and NUDC protein. D Fluorescence microscopy revealed co-localization of RNF41 (red) and NUDC (green) in T24 cells, with nuclear staining by 4’, 6-diamidino-2-phenylindole (DAPI; blue). Scale bar, 10 μm. E Protein docking map illustrating the predicted interaction between RNF41 and NUDC. F Endogenous co-immunoprecipitation results showing the interaction of RNF41 and NUDC in T24 cells. G Co-immunoprecipitation of FLAG-RNF41 and MYC-NUDC in T24 cells, transfected with FLAG-RNF41 and MYC-NUDC plasmids for 36 h. H HEK293T cells co-transfected with the indicated shRNA underwent immunoprecipitation (IP) with anti-NUDC antibody, followed by immunoblotting (IB) with antibodies against NUDC, RNF41, and β-actin. Cells were treated with 20 μM MG132 for 6 h prior to harvesting. I HEK293T cells co-transfected with Myc-NUDC, HA-ubiquitin (HA-Ub), and Flag-tagged RNF41 WT or RNF41 L163Q, with lysates subjected to IP using anti-NUDC antibody followed by IB with antibodies against Myc-tagged, Flag-tagged proteins, and β-actin. Cells were treated with 20 μM MG132 for 6 h before collection. J HEK293T cells were co-transfected with Myc-NUDC and Flag-tagged full-length RNF41 or its deletion mutants, followed by immunoprecipitation with Flag beads and immunoblotting with antibodies against Myc and Flag.

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