The growth hormone regulated serine protease inhibitor (SPI) 2.1 and 2.2 gene promoters have been shown to contain a response element similar to the gamma-interferon activated sequence (GAS) family of signal transducer and activator of transcription (STAT) response elements. We have investigated the STAT and cytokine specificity of the SPI 2.1 STAT responsive element using a luciferase (LUC) reporter construct and a cDNA complementation strategy in the COS 7 cell line. Growth hormone was found to stimulate SPI-LUC reporter gene expression via activation of STAT 5, but not STATs 1 or 3, which indicates that the SPI 2.1 STAT responsive element is STAT 5 specific. In addition to the growth hormone receptor, the receptors for prolactin and erythropoietin enhanced gene transcription via the SPI 2.1 STAT responsive element, which indicates that this element is, on the other hand, not cytokine specific. Activation of STAT 5 was also observed after growth hormone treatment of cells transfected with cDNA expression plasmids for several different truncated growth hormone receptor mutants, although this activation was less efficient than with the wild type receptor. Point mutation of individual tyrosines in the growth hormone receptor intracellular domain to phenylalanines had no significant effect on signal transduction via STAT 5. These data, taken together with results from experiments using the phosphatase inhibitor sodium orthovanadate, suggest that STAT 5 may not have an absolute requirement for specific phosphorylated receptor tyrosine docking sites. That receptor tyrosine residues in a variety of amino acid contexts, or phosphorylated Janus kinase (JAK) 2 alone, can facilitate STAT 5 activation could explain the observed lack of cytokine specificity in STAT 5 activation.