Colony-stimulating factors (CSFs) are glycoproteins that stimulate the growth of hematopoietic progenitors and enhance the functional activity of mature effector cells. Human granulocyte/macrophage colony-stimulating factor (GM-CSF) is a 22-kDa glycoprotein that stimulates the growth of myeloid and erythroid progenitors in vitro and increases the responsiveness of neutrophils, monocytes, and eosinophils to physiologic stimuli. Elucidation of the cell and tissue sources of CSFs, as well as study of their regulation of expression, is required to understand their role in physiologic and pathophysiologic states. An extensive survey of normal and neoplastic human tissues did not reveal constitutive production of detectable levels of GM-CSF mRNA in any of the 64 samples studied. Antigen- or lectin-activated T lymphocytes have been shown to produce GM-CSF; therefore, to elucidate the genetic sequences required, we constructed recombinant plasmids containing 5' flanking DNA of the GM-CSF gene linked to the marker chloramphenicol acetyltransferase gene. The recombinant constructs were transfected into a human T-cell leukemia virus type I (HTLV)-infected T-lymphoblast cell line that can be stimulated to produce high levels of GM-CSF. We show here that the 5' flanking sequences of the GM-CSF gene can direct increased expression of the chloramphenicol acetyltransferase gene in activated T-lymphoblast cells.