Extended Data Fig. 8: Transformed cells display diverse capacity to utilize inosine as a carbon source.
From: Inosine is an alternative carbon source for CD8+-T-cell function under glucose restriction
a, The indicated cell lines were cultured without glucose (background), with glucose or with inosine, and cell growth curves were monitored by live cell imaging analysis (IncuCyte ZOOM™). Cell confluences of the indicated cell lines cultured with glucose at 72-96 h were set to 100%. Data are presented as mean ± SD (n=4). **, P<0.01 and ***, P<0.001 for Glucose versus Inosine. P=0.00009, 0.0001, 0.0000001, 0.00001, 0.0042, 0.6917, 0.0006, 0.00001, 0.9059, 0.7962, 0.0796, 0.6663, 0.3829 from left to right, respectively. b, The expressions of PNP in indicated cell lines were determined by immunoblot. c, B16-F10 cells were cultured in the indicated conditional media and cell growth curves were monitored and analyzed by IncuCyte ZOOM™. Data are presented as mean ± SD (n=4). ***, P<0.001 for 2 mM Glucose versus 2 mM Inosine. Data are representative of three independent experiments. d, Quantification of indicated metabolites in HeLa cells. All abbreviations are as described in Fig. 2. Values represent mean ± SEM (n=3). e, HeLa cells were incubated without glucose (background), with glucose or with inosine, as well as in combination with increased concentrations of Foro (0.1 μM, 0.5 μM, and 2 μM), and cell growth was monitored by live cell imaging analysis (IncuCyte ZOOM™). The confluences of HeLa cells cultured with glucose at 60 h were set to 100%. Data are presented as mean ± SD (n=4). f, HeLa cells were transfected with scrambled siRNA or PNP siRNA for 3 days. Cells were then switched to glucose-free (background), glucose-containing, or inosine-containing media and cell confluence was determined by live cell imaging analysis (IncuCyte ZOOM™). The confluences of HeLa cells in the presence of glucose at 96 h were set to 100%. Error bars represent standard deviation from mean of quadruplicate samples (left). NS, P=0.2022; ***, P=0.0000006 for Glucose versus Inosine. PNP protein expression levels were determined by immunoblot (right). Data are representative of two independent experiments (b, e, f). Data were analyzed by unpaired two-sided t-test (a, c, f). Sample size (n) represents biologically independent samples (a–f).
