Extended Data Fig. 2: 1-Deoxysphingolipid synthesis and degradation influence spheroid growth in vitro.
From: Serine restriction alters sphingolipid diversity to constrain tumour growth
a, Total (hydrolysed) sphinganine and deoxysphinganine (deoxySA) abundances in adherent and spheroid cultures of HCT116 cells (n = 3 culture wells each). b, Free deoxysphinganine abundances in HCT116 cultures in normal medium or medium without serine and glycine but containing 1 mM alanine (−SG+A) (n = 3 culture wells each). c, Deoxysphinganine/sphinganine (SA) molar ratio in HCT116 cells in cultures from (b) (n = 3 culture wells each). d, HCT116 spheroid growth in the presence or absence of 10 nM deoxysphinganine and treated with vehicle (DMSO) or UK5099 (n = 3 culture wells each). e, Adherent (n = 4 culture wells each) and spheroid (n = 3 culture wells each) biomass of HCT116 cells treated with vehicle or 50 nM sphingoid bases. SO, sphingosine; deoxySO, deoxysphingosine. f, Free deoxysphinganine, summed deoxyDHCER and summed deoxyCER in spheroid cultures from (e) (n = 3 culture wells each). g, Sphinganine and deoxysphinganine abundances in HCT116 spheroid cultures treated with vehicle or 10 nM myriocin (n = 3 culture wells each). h, Deoxysphinganine/ sphinganine molar ratio in HCT116 adherent and spheroid cultures treated with vehicle or 10 nM myriocin (n = 3 culture wells each). i, Free deoxysphinganine abundances in HCT116 spheroid cells in the presence or absence of 1 mM alanine and treated with vehicle or 10 nM myriocin (n = 3 culture wells each). j, Spheroid growth of cell lines cultured in the presence or absence of 1 mM alanine (red), 10 nM myriocin (blue) or both (red outline with checkered blue fill) (n = 3 culture wells each condition). k, A549 spheroid growth under 5 d of culture in myriocin or 10 μM fumonisin B1 (FuB1) (n = 3 culture wells each). l, m, HCT116 spheroid growth (l) and free deoxysphinganine (m) in the presence (+SG) or absence (−SG) of 0.4 mM serine and glycine and treated with DMSO, fenofibrate (FeF), 1 mM alanine or both (n = 3 culture wells for each condition). Two-sided Student’s t-test (a, g), two-way ANOVA (b–d, h, i, l, m) or one-way ANOVA (e, f, j, k) was performed for each comparison with no adjustment for multiple comparison. Similar results obtained in two (b–e) or three (a, g–i) independent experiments. Data are mean ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.0001, #P < 0.0001.
