Fig. 6: CTSL-BNIP3 network promotes tumor metastatic potential.
A Human PPI network of CTSL-BNIP3 neighbors colored by log2 FC (tumor versus matched normal lung) in TCGA LUAD. B Immunofluorescence of CTSL (green) and BNIP3 (red) in LUAD tissue microarray cores: normal adjacent, n = 22; tumor cores, n = 128. Bar graph shows proportions of BNIP3-high/-low within the CTSL-high and CTSL-low tumors. C Kaplan–Meier overall survival for TCGA LUAD stratified by combined CTSL and BNIP3 expression (CTSL-BNIP3-high, red, n = 183; CTSL-BNIP3-low, black, n = 156). D GSEA plots of ALONSO_METASTASIS_UP in CTSL-BNIP3 high versus low groups in TCGA LUAD (left) and in TB mice (right). E Representative WB of indicated proteins in TB muscle. Two mice per group are shown (n = 4 per group); quantification of all four mice in Supplementary Fig. 16A. Samples derive from the same experiment; different gels for Ctsl, N-Cad, Gapdh, and another for Bnip3 were processed in parallel. F WB of indicated proteins in A549 cells treated with or without CTSLi; GAPDH as loading control; quantification from four independent biological replicates in Supplementary Fig. 16B. Samples derive from the same experiment; different gels for CTSL and another for N-Cad, Bnip3, GAPDH, were processed in parallel. Migration (G) and invasion (H) assays for A549 cells treated with or without CTSLi. Bar graphs represent quantified gap closure (G) and invaded cells (H); biological triplicates per group. WB of indicated proteins in A549 cells transfected with si-CTSL (I) or si-BNIP3 (J); quantification from four independent biological replicates in Supplementary Fig. 16E, F. For I, samples derive from the same experiment; different gels for CTSL and another for N-Cad, Bnip3, GAPDH, were processed in parallel. For J, samples derive from the same experiment; different gels for CTSL, N-cad, GAPDH, and another for BNIP3 were processed in parallel. Quantified migration after si-CTSL (K) or si-BNIP3 (L) transfection; biological triplicates per group. M Working model: combined αPD-L1 and CTSLi therapy suppresses tumorigenesis. For B, one-sided Fisher’s exact test; for C, log-rank test; for D, GSEA; for G, H, K, L, data are mean ± SEM, two-sided unpaired t-test. Source data are provided as a Source Data file.
