Fig. 4: DLAT binds to MTHFD2 in the mitochondria.

a Top 10 mitochondrial interactors of DLAT beyond PDC in A549^cisR cells with or without DLAT knockdown. DLAT binding proteins were co-immunoprecipitated using DLAT antibody, and peptides were measured by LC-MS/MS. The normalized peak intensity value for each protein is shown. b Co-immunoprecipitation showing candidate interactions with DLAT. c Representative MS spectrum of MTHFD2 peptide fragments is shown. d Cisplatin-induced apoptotic cell death and cell viability in cancer cells without or with knockdown of MTHFD2. Stable knockdown cells were treated with sublethal doses of cisplatin (A549^cisR 2 μg/ml and KB-3-1^cisR 5 μg/ml) for 48 h. Effect of MTHFD2 inhibitors on cisplatin-mediated apoptotic cell death and cell viability. Cells were treated with either 10 μM DS18561882 (e) or 10 μM LY345899 (f) and sublethal doses of cisplatin for 48 h. g Endogenous interaction between MTHFD2 and DLAT, independent of other subunits of the PDC, was determined by MTHFD2 co-immunoprecipitation in cancer cells. h Immunofluorescence assay shows the co-localization of DLAT and MTHFD2 in A549^cisR cells. i Mitochondrial localization of DLAT and MTHFD2 in A549^cisR and KB-3-1^cisR cells is shown by immunoblotting of mitochondria and cytosolic fractions prepared using a mitochondria isolation kit. Tom40 and α-tubulin were used as control markers for mitochondria and cytosol, respectively. Immunofluorescence staining demonstrates the localization of DLAT (j) and MTHFD2 (k) with mitochondrial marker MitoTracker in A549^cisR cells. Scale bars represent 10 μm for (h, j, k). Data are mean ± SD from 3 independent biological replicates for (d–f). P values were determined by one-way ANOVA. Source data are provided as a Source Data file.