Cancer cells depend on environmental lipids for proliferation when electron acceptors are limited.

Production of oxidized biomass, which requires regeneration of the cofactor NAD + , can be a proliferation bottleneck that is influenced by environmental conditions. However, a comprehensive quantitative understanding of metabolic processes that may be affected by NAD + deficiency is currently missing. Here, we show that de novo lipid biosynthesis can impose a substantial NAD + consumption cost in proliferating cancer cells. When electron acceptors are limited, environmental lipids become crucial for proliferation because NAD + is required to generate precursors for fatty acid biosynthesis. We find that both oxidative and even net reductive pathways for lipogenic citrate synthesis are gated by reactions that depend on NAD + availability. We also show that access to acetate can relieve lipid auxotrophy by bypassing the NAD + consuming reactions. Gene expression analysis demonstrates that lipid biosynthesis strongly anti-correlates with expression of hypoxia markers across tumor types. Overall, our results define a requirement for oxidative metabolism to support biosynthetic reactions and provide a mechanistic explanation for cancer cell dependence on lipid uptake in electron acceptor-limited conditions, such as hypoxia.