The activity of asparagine synthetase decreased almost 50% during dexamethasone-induced mouse myeloid leukemia M1 cell differentiation. This enzyme activity also declined significantly during differentiation of the human myelogenous leukemic cell lines, HL-60 and U-937, induced by either macrophage culture supernatant or retinoic acid. The decline of asparagine synthetase activity closely paralleled the expression of various maturation markers, but could also be induced by serum starvation. These results suggest that asparagine synthetase or L-asparagine has some biological function in growth regulation of these leukemia cell lines.