Extended Data Fig. 4: NHE activity is required for early G1 progression.
From: Cytosolic pH regulates proliferation and tumour growth by promoting expression of cyclin D1
(a, b) Elevated pHc is required within the first 8 hours of the cell-cycle. Extracts of cells as in Fig. 1g were blotted for RB phosphorylation and cyclin A expression as markers for cell-cycle progression. Blots representative of three independent experiments are shown. (b) Elevated pHc is required to drive cell-cycle progression before initiation of S phase. Cells were starved (-/-) and treated with FCS glucose for the indicated time points and BrdU incorporation was measured to follow DNA synthesis. Results of a representative experiment from three independent experiments with percentage of cells in G1, S and G2/M phase are shown. (c, d) pHc regulates a subset of FCS and glucose dependent genes required for G1 progression. Cells were starved for 24 hours, treated with FCS glucose in the presence or absence of DMA for 5 hours and analyzed by RNAseq. (c) Venn diagram indicating the overlap of starvation and DMA-regulated genes. P-value demonstrating statistical significance of co-regulated genes based on a hypergeometric distribution is shown. (d) Genes repressed upon starvation and DMA treatment and co-regulated genes in both conditions were analyzed by GO-term enrichment using ShinyGO (http://bioinformatics.sdstate.edu/go/). Significantly enriched GO-terms were identified for the individual groups and plotted as -log10(FDR).
