Fig. 5. Purine depletion triggers cell elongation and an EMT transcriptional program through an increase in cellular serine levels.

a Representative images of actin cytoskeleton staining with Phalloidin (green) and nuclei (blue) from A375 cells treated with MTX (200 nM, 24 h) in the presence or absence of inosine (50 µM, 24 h). Quantification of cell length from seven independent cells is presented relative to vehicle control. b As in (a), but from A375 cells that were treated with LTX (800 nM, 24 h) or AG2037 (100 nM, 24 h). c Relative mRNA expression of the indicated EMT markers upon treatment with MTX (200 nM, 24 h) in the presence or absence of inosine (50 µM, 24 h). d As in (a), but from A375 cells that were treated with MTX (200 nM, 24 h) in the presence or absence of PHGDH inhibitor (PHGDHi, BI-4924, 15 µM, 24 h). e Relative mRNA expression of the indicated EMT markers upon treatment with MTX (200 nM) in the presence or absence of PHGDH inhibitor (PHGDHi, BI-4924, 15 µM, 24 h). f Relative mRNA expression of the indicated EMT markers as in c, but from wild-type or ΔPHGDH A375 cells treated with either vehicle (DMSO) or MTX (200 nM, 24 h). g As in (a), but from A375 cells that were treated with MTX (200 nM, 24 h) in the presence or absence of TEPP-46 (100 µM, 24 h). h As in (a), but from A375 cells that were treated with serine (400 nM, 24 h) or Formate (2 mM, 24 h) in normal DMEM media. i Relative mRNA expression of the indicated EMT markers upon treatment with serine (400 nM, 24 h) or Formate (2 mM, 24 h) in normal DMEM media. a–i Data are presented as the mean ± s.d from n = 3 (c, e, f, i) or n = 7 (a, b, d, g, h) of biologically independent samples. a–i One-way ANOVA, Turkey’s post-hoc test, multiple comparison *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. a, b, d, g, h) Scale bars are indicated. Data are representative of n = 2 (c, e, f, i) or n = 3 (a, b, d, g, h) independent experiments. Source data and exact p-values are provided as a Source Data file.