Although the mouse protamine 1 gene (mP1) is first transcribed in round spermatids, its mRNA is not translated until about 1 week later in elongating spermatids. To determine what mP1 sequences are important for its transcriptional and translational regulation, we have constructed fusions between mP1 and the human growth hormone (hGH) structural gene and analyzed their expression in transgenic mice. We show that mP1 sequences 5' to the start of transcription are sufficient to confer spermatid-specific expression on the hGH gene. We also show that 156 nucleotides of mP1 3'-untranslated sequence is sufficient to confer mP1-like translational regulation on the hGH mRNA. Interestingly, the subcellular localization of hGH was dependent on the time during spermiogenesis that it was made. Synthesis of hGH in early round spermatids resulted in localization in the acrosome, whereas synthesis in late elongating spermatids resulted in intracellular, but not acrosomal, localization.