Extended Data Fig. 7: The Gstt1High macrometastatic subpopulation regulates cell cycle progression and shares dissemination gene signatures with latent DTCs.
From: The glutathione S-transferase Gstt1 drives survival and dissemination in metastases
(A) A panel of metastatic cell lines (independently derived from PDAC liver and lung metastases) expressing either Control or shGstt1 were subjected to RNA-Seq. DAVID biological pathway analysis on Control vs shGstt1 differentially expressed gene signatures (95 UP, 217 DN) (>2FC, FDR 0.01) (GO_TERMs). Pathways were ranked by p-value(Log10) using an unpaired, two-sided t-test with post-hoc Bonferroni correction. (B) Expression (Log2CPM) of ‘Cell Cycle’ GO_TERM genes enriched in Control vs shGstt1 from (A) (>2FC, FDR 0.01). (C) Metastatic-derived PDAC cells stably expressing control, shGstt1 #1 or shGstt1 #2 were grown for 5 days, fixed, and analyzed for cell cycle stages using propidium iodide and flow cytometry. Cell cycle stages were analyzed using FlowJo. Data represents n = 3 independent experiments. Data are represented as mean s.d. ANOVA with Brown-Forsythe post-hoc test for multiple comparisons was performed to determine statistical significance between groups (*P = 0.0247). (D) Dissemination gene panel identified by Hosseini et al, 2018 was analyzed in mCherry populations and DTCs. Data represented as genes commonly enriched in mCherryhigh cells and DTC populations compared to mCherrylow. Data are represented as mean s.e.m. Two-way ANOVA with a post-hoc Tukey’s multiple comparisons test was used to determine statistical significance between groups (Log2CPM, ****P < 0.0001). (E) Dissemination gene panel (n = 5 genes) identified in Hosseini et al, 2018 from (D) were analyzed for impact on overall survival in pancreatic cancer (KMPlot). Data represents overall survival (OS) in N = 177 patient dataset based on mean expression of selected genes. A long-rank test was used to determine significance (*P = 0.0022). (F) Proliferation genes commonly downregulated in mCherryhigh cells and DTC populations compared to mCherrylow. Data are represented as mean s.e.m. Two-way ANOVA with a post-hoc Tukey’s multiple comparisons test was used to determine statistical significance between groups (Log2CPM, ****P < 0.0001).
