Fig. 5: LINC00941 acts as a decoy to competitively bind ANXA2 and inhibits NEDD4L-mediated ubiquitination.

A Silver-stained SDS-PAGE displayed a results of immunoprecipitated using IgG and anti-ANXA2 in ANXA2 overexpression PANC-1 cells, followed by mass spectrum. B Co-IP assay was performed to clarify the interaction between endogenous ANXA2 and endogenous NEDD4L in PANC-1 and MIA PaCa-2 cells. C, D Flag-tagged ANXA2 and HA-NEDD4L were transfected into 293T cells, followed by using Co-IP assay to clarify the interaction between exogenous NEDD4L and exogenous ANXA2. E The protein expression of NEDD4L and ANXA2 was measured in NEDD4L silenced group and negative control group by western blot. F The ubiquitination of ANXA2 upon NEDD4L or not were analyzed by Co-IP with the treatment of MG132 (10 μM) and Myc-Ubi. G The ubiquitination of ANXA2 in NEDD4L knockdown or not were analyzed by Co-IP with treatment of MG132 (10 μM) and Myc-Ubi. H The ubiquitination of ANXA2 in LINC00941 overexpression combining with NEDD4L or not were analyzed by Co-IP with the treatment of MG132 (10 μM) and Myc-Ubi. I The ubiquitination of ANXA2 in NEDD4L knockdown combining LINC00941 overexpression or not were analyzed by Co-IP with the treatment of MG132 (10 μM) and Myc-Ubi. J, K Co-IP assays were performed to clarify the effect of LINC00941 combining NEDD4L or not on the interaction between NEDD4L and ANXA2 with the treatment of MG132 (10 μM) in PANC-1 (J) and MIA PaCa-2 K cells. L The expression of ANXA2 was measured by western blot in LINC00941 overexpression combining NEDD4L knockdown in PANC-1 and MIA PaCa-2 cells. M The expression of ANXA2 was measured by western blot in LINC00941 overexpression combining NEDD4L overexpression in PANC-1 and MIA PaCa-2 cells. N, O The truncated of ANXA2 and NEDD4L was co-transfected into 293T cells, followed by Co-IP assay to clarify the interaction domain between ANXA2 and NEDD4L.