Extended Data Figure 3: Reductive citrate metabolism in anchorage-independent spheroid culture.
From: Reductive carboxylation supports redox homeostasis during anchorage-independent growth
a, Citrate m+5 may be generated in several ways from [U-13C]glutamine, including through reductive (left) or oxidative (middle) pathways. To test whether citrate m+5 arises from oxidative or reductive metabolism, spheroids were cultured with [1-13C]glutamine. Glutamine-C1 (green circle) is released as CO2 by α-ketoglutarate dehydrogenase in the oxidative TCA cycle, but is transferred to citrate via reductive metabolism. Citrate mass isotopologues in H460 cells cultured with [1-13C]glutamine (right). The m+1 fraction in this experiment is comparable to the m+5 fraction from [U-13C]glutamine (~20%), indicating that reductive labelling was enhanced in spheroids (n = 3 cultures from a representative experiment). b, Time-dependent evolution of succinate, fumarate and malate mass isotopologues in spheroids cultured with [U-13C]glutamine (n = 2 cultures for each time point). c, Citrate labelling from [U-13C]glutamine in immortalized, non-transformed bronchial epithelial cells (HBEC30) and lung cancer cells (HCC4017) from the same patient (n = 3 cultures from two experiments). All data represent mean ± s.d. *P < 0.05, Welch’s unequal variances t-test. All experiments were repeated 3 times or more.
