The Pem gene encodes an atypical homeodomain protein, distantly related to Prd/Pax family members, that we demonstrate is regulated in a complex transcriptional and post-transcriptional manner. We show that the rat Pem genomic structure includes three 5'-untranslated (5'-UT) exons and four coding exons, three of which encode the homeodomain. Several alternatively spliced transcripts were identified, including one that skips an internal coding exon, enabling this mRNA to express a novel form of the Pem protein. Other alternatively spliced mRNAs were characterized that possess different 5'-UT regions, including a muscle-specific transcript. The different 5'-UT termini present in Pem transcripts conferred different levels of translatability in vitro. Two promoters containing multiple transcription initiation sites were identified: a distal promoter (Pd) in the first 5'-UT exon and a proximal promoter (Pp) located in the "intron" upstream of the first coding exon. The Pd was active in placenta, ovary, tumor cell lines, and to a lesser extent in skeletal muscle. In contrast, transcripts from the Pp were only detectable in testis and epididymis and were only expressed in epididymis in the presence of testosterone. To our knowledge no transcription factors have previously been identified that exhibit androgen-dependent expression in the epididymis.