Fig. 3

USP14 interacts with PFKL. A The exogenous expression of HA-USP14 in the pull-down elution was validated by Western blotting. B Representative images of silver-stained protein bands for mass spectrometry (MS) analysis of USP14-interacting proteins. C Venn diagram showed the numbers of mouse IgG and HA antibody binding proteins identified by immunoprecipitation coupled with mass spectrometry analysis (IP-MS). D Co-immunoprecipitation (Co-IP) analysis of the interactions between exogenous USP14 and endogenous PFKL. Whole-cell lysates from HN4 cells stably expressing HA-USP14 were immunoprecipitated and immunoblotted with antibodies against the indicated proteins. E Co-IP analysis of the interactions between endogenous USP14 and PFKL in HN4 cells. F, G Co-IP analysis of the interactions between exogenous USP14 and PFKL in HEK293T cells. Whole-cell lysates from HEK293T cells stably expressing HA-USP14 and Flag-PFKL were immunoprecipitated and immunoblotted with antibodies against the indicated proteins. H Confocal microscopic analysis of USP14 and PFKL subcellular localization. HN6 cells were fixed and immunoblotted with antibodies against the indicated proteins. Representative images from biological triplicate experiments are shown. Scale bar, 10 μM. I Nuclear and cytoplasmic proteins in three OSCC cell lines were separated by protein extraction kit and immunoblotted with antibodies against the indicated proteins