Fig. 6.

Nuclear GRP78 regulates the expression of genes important for cell migration and invasion in human lung cancer cells. (A) Volcano plot for differentially expressed genes in H1975 cells expressing F-78(WT) vs. F-78(NLS Mut). (B) Heatmap and clustering of differentially expressed genes from (A). (C and D) Enrichment of cell components and molecular functions of up-regulated genes in F-78(WT) vs F-78(NLS Mut) using ClusterProfiler. (E) Representative confocal fluorescence images of H1975 cells transfected with F-78(WT) (Upper) or F-78(NLS Mut) constructs (Lower) for 48 h and stained for F-actin with phalloidin (green). DAPI (blue) represents nuclei staining. (Scale bars, 10 μm.) (F) Representative confocal fluorescence images of DAPI-stained cells remaining on the microporous membrane (M.M.) on the left and cells migrated to the lower compartment (L.C.) on the right in transwell migration assay of H1975 cells transfected with si78 16 h, followed by transfection with F-78(WT) or F-78(NLS Mut) constructs for 48 h. (G) Quantification of the number of cells remaining on the M.M. and cells migrated to the L.C. in (F). Data are presented as mean ± SEM. *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001 (Student’s t test). (H) Proposed model for nuclear GRP78-mediated transcriptional regulation via interaction with ID2 under ER stress. In nonstressed cell (Left), ID2 binds to E protein and prevents its interaction with bHLH transcription factor. In cancer or stressed cells (Right), GRP78 is up-regulated and translocates to the nucleus where it binds and sequesters ID2, relieving the inhibitory effects on transcription leading to activation of genes important for migration and invasion. See also SI Appendix, Figs. S6 and S7.