The relationship between expression of the pituitary-specific transcription factor, GHF-1, and activation of the growth hormone and prolactin genes during mouse anterior pituitary development was investigated. While GHF-1 transcripts were detected within 24 hr of the first observable events in anterior pituitary differentiation, no GHF-1 protein could be detected until about 3 days later. The appearance of GHF-1 protein showed good temporal and spatial correlation with activation of the growth hormone gene. Prolactin gene expression, on the other hand, was observed transiently during embryonic day 16 in two different populations of cells, of which the major one does not contain GHF-1 or growth hormone. These results suggest that expression of GHF-1 is controlled both transcriptionally and posttranscriptionally. The spatial and temporal correlation between the appearance of GHF-1 protein and growth hormone gene activation suggests that GHF-1 is responsible for this very last step in the specialization of somatotrophic cells.