Extended Data Fig. 7: Design and functional validation of the oncofetal phenotypic reporter.
From: Oncofetal reprogramming drives phenotypic plasticity in WNT-dependent colorectal cancer
a, Design method the oncofetal (OnF) synthetic locus control regions (sLCR). b, Genome Browser view of cis-regulatory elements used in the OnF sLCR. c, Normalized distribution of OnF-GFP+ cells and eGFP fluorescence intensity measured by flow cytometry. d, eGFP geometric mean fluorescence insensitivity (MFI) quantification. Error bars, s.d; n = 3 independent cultures, p-values calculated using Fisher/s LSD post-hoc test after one-sided ANOVA (p < 0.05). e, Scaled OnF gene expression in GFPhigh and GFPNeg cells sorted from VAKSPOnF tumoroids, n = 3 independent sorts. f, Schematic of the OnF sLCR substitution with the STAR minigene. g, Representative fluorescence microscopy images of VAKSP tumoroids co-expressing both reporters (VAKSPOnF/STAR). Scale bars, 200μm. h, Volcano plot of differentially expressed genes (DEGs) in VAKSP tumoroids post-3-day FOLFIRI treatment (n = 3 independent experiments). Significant DEGs are on both sides of dashed lines (log2FC > 1; p-value < 0.01); p-values, DESeq2, Wald test (two-sided, unpaired); Benjamini-Hochberg adjustments. i, The GFP/mCherry gating strategy to sort clones used in (j-l). j-l, Quantification of predominantly cell states in response to FOLFIRI in 3 independent clones sorted from GFPhigh (j), mCherryhigh (k) or GFP/mCherryhigh (hybrid) (l) populations. p-values calculated using paired two-sided t-tests. m-r, Representative pseudocolor plots of flow-cytometry analysis of GFPhigh (m-n) mCherryhigh (o-p) and hybrid clones (q-r) before (m, o and q) and after FOLFIRI tretament for 3 days (n, p and r), related to (j-l).
