S ince transferrin is required for cellular proliferation, we investigated transferrin synthesis by a small cell lung cancer line (NCI-H510) that survives in serum-free media without added transferrin. Immunoassays for human transferrin demonstrated that these cells contained immunoreactive human transferrin. Immunofluorescence studies showed that the protein is expressed on the surface of cells, presumably bound to transferrin receptor. Media conditioned by NCI-H510 cells support proliferation of human leukemic cells that would not survive in media lacking transferrin. [35S]Methionine incorporation documented transferrin synthesis by NCI-H510 cells as well as three other small cell lines. Transferrin synthesis by NCI-H510 cells increased more than 10-fold when cells entered active phases of the cell cycle, and this increase was seen before large increases in transferrin-receptor expression. Further experiments examining the effects of agents that affect iron metabolism show that the addition of transferrin-iron or hemin to the media is associated with a more rapid initial rate of proliferation and lower rates of transferrin synthesis than control cells. Gallium salts, which inhibit iron uptake, inhibited proliferation of these cells. If the cells recovered from this effect, transferrin synthesis remained greatly increased compared to control. We conclude that transferrin synthesis by these malignant cells is ultimately related to an iron requirement for cellular proliferation. It appears that this synthesized transferrin acts as part of an important autocrine mechanism permitting proliferation of these cells, and perhaps permitting tumor cell growth in vivo in areas not well vascularized.