Figure 2. Physiological serine concentrations approach a range that is limiting for cell proliferation in culture.
(A) Western blot showing CRISPRi-mediated knockdown of PHGDH expression in cells derived from an autochthonous breast tumor arising in a BRCAfl/fl; Trp53+/−; MMTV-Cre mouse. Cells with sgRNA targeting PHGDH (sgPHGDH-1) or a non-targeting sgRNA (sgctrl) are shown. (B) Fraction of 13C-labeled serine (fully labeled, M+3) in control (sgctrl) or PHGDH knockdown (sgPHGDH-1) cells cultured in U-13C glucose. The difference in the fraction of labeled serine between the cell types is significant (p< 0.0001) by an unpaired, Welch’s t test. (C) Proliferation rate of control (sgctrl) and PHGDH knockdown (sgPHGDH-1) cells. There is no significant change in proliferation rate by unpaired, two-tailed Welch’s t test (p = 0.2820). (D) Serine concentration in plasma of C57BL/6J mice collected at 11 AM. n=60 mice. (E) Proliferation rate of control (sgctrl) and PHGDH knockdown (sgPHGDH-1) breast cancer cells cultured in media containing the indicated amounts of serine. Grey box indicates the range of plasma serine concentrations measured in (D). (F) Plasma serine, glycine, and lysine concentrations measured in the indicated mouse strains fed ad lib on a control diet, fasted starting at 5 AM, or fed a serine and glycine free diet for one week prior to measurement. Plasma was collected at the specified times. n=5 mice on each diet. (G) Plasma serine and glycine concentrations in C57BL/6J mice fed a serine and glycine free diet, a control diet, or a high serine diet for the indicated time. Plasma was collected at 11 AM. n=5 mice on each diet. (H) Mouse weight for animals fed a serine and glycine free diet, a control diet, or a high serine diet for the indicated time. n=5 mice on each diet.
Mean +/− SD is shown for all panels.