Environmental cystine drives glutamine anaplerosis and sensitizes cells to glutaminase inhibition

Many cancer cell lines depend on extracellular glutamine as a major tri-carboxylic acid (TCA) cycle anaplerotic substrate to support proliferation in vitro. However, recent studies have suggested that some cells that depend on glutamine anaplerosis in culture rely much less on glutamine catabolism to proliferate in vivo, with environmental differences between tumors and cell culture influencing the extent of glutamine catabolism. Here we sought to better understand the environmental differences that cause differential dependence on glutamine for TCA cycle anaplerosis. We find that cells cultured in adult bovine serum, a condition that more closely reflects the nutrients available to cells in vivo, leads to decreased glutamine catabolism and reliance on glutamine anaplerosis compared to standard tissue culture conditions. By analyzing the nutrient differences between bovine serum and media, we find that levels of a single nutrient, cystine, can account for the differential dependence on glutamine in these different environmental contexts. Further, we show that cystine levels dictate glutamine dependence via the cystine/glutamate antiporter xCT/SLC7A11, and that environmental cystine levels in conjunction with xCT/SLC7A11 expression is necessary and sufficient to drive increased glutamine anaplerosis, defining important determinants of glutamine anaplerosis and glutaminase dependence in cancer cells.