Fig. 1: VSTM2L is a VDAC1 interacting protein in PCa cells.

A The top ten proteins, identified by co-IP/MS, was elucidated in association with VDAC1 in LNCaP cells. B, C HEK293T cells were transfected with Myc-VDAC1 and 3 × Flag-VSTM2L expression plasmids as indicated, whole cell lysates were extracted and immunoprecipitated with anti-Flag (B) or anti-Myc (C) antibodies. D, E DU145 (D) and 22Rv1 (E) cells overexpressed Myc-VDAC1-pLVX or control cells were subjected to immunoprecipitation with anti-Myc antibody and followed by western blot analysis. F The direct interaction between VSTM2L and VDAC1 was confirmed by GST pull-down assay. GST-VSTM2L fusion proteins were purified and incubated with PC3 cell lysates and analyzed by western blotting. Purified GST proteins served as negative controls. G A representative image of an SDS-PAGE gel stained with Coomassie blue. LNCaP cells transfected with 3 × Flag-VSTM2L were collected for immunoprecipitation with anti-Flag antibodies, and separated by SDS-PAGE gel. The gel was subsequently stained with Coomassie blue to visualize the distinct binding bands. H Venn diagram showing the 574 proteins obtained by intersecting the IP-MS data of VDAC1 and VSTM2L. I The top 15 positively enriched KEGG pathways of the 574 proteins are shown. Hypergeometric test was used for P-value calculation. J Western blotting analysis of the expression of VSTM2L in mitochondria and cytoplasm of PC3, DU145 and 22Rv1 cells, respectively. Mitochondria and cytoplasmic protein levels were normalized to COX IV and α-Tubulin, respectively. The samples were derived from the same experiment and the gels/blots were processed in parallel. K Representative images of immunofluorescence staining for VSTM2L and MitoTracker in PC3, DU145, and 22Rv1 cells. The nucleus was stained with Hoechst. Scale bars, 8 μm. Source data are provided as a Source Data file.