The Homeobox Gene

Abstract

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 (P d ) in the first 5-UT exon and a proximal promoter (P p ) located in the "intron" upstream of the first coding exon. The P d was active in placenta, ovary, tumor cell lines, and to a lesser extent in skeletal muscle. In contrast, transcripts from the P p 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.

Localization and Stability of Introns Spliced from thePem Homeobox Gene

“…We also considered the possibility that IVS1 and IVS2 preferentially leak out of the nucleus during RNA preparation in vitro because they are smaller than Pem pre-mRNAs. To test this notion, we examined the localization of Pem IVS3, as it is much larger (2.4 kb) than the other Pem introns (25). We found that IVS3 was present in the cytoplasmic fraction at a level approximately equal to that in the nuclear fraction, when an equal amount of RNA from each fraction was loaded (Fig.…”

“…In the present communication, we report on the fate, localization, and stability of spliced introns from the Pem homeodomain gene, a mammalian gene that contains typical mammalian introns ranging in length from ϳ0.2 to 2.4 kb (25,26). Unlike the other genes from which spliced introns have been studied, the Pem gene is not expressed in a cell type-specific manner.…”

“…Its ϳ1.5-kb size indicates that m2 is polyadenylated at the consensus site at the end of the last Pem exon (25). In contrast, the shorter m1 transcript is a mature mRNA whose 3Ј terminus is probably generated by use of the polyadenylation signal consensus sequence (AAUAAA) present in the 5Ј portion of IVS2 (25). The evidence for this assignment includes the following: its ϳ0.9-kb size is consistent with a transcript that polyadenylates in the beginning of IVS2; it hybridized with a 5Ј IVS2 probe ( Fig.…”

“…25; GenBank TM accession number U52034). To study the half-life of the Pem introns, we placed the Pem gene downstream of the tet-regulated promoter (Fig.…”

“…Unlike the other genes from which spliced introns have been studied, the Pem gene is not expressed in a cell type-specific manner. Instead, Pem is expressed by different cell types in many different fetal and adult tissues, as well as in tumor cells from several different lineages (25)(26)(27)(28)(29)(30)(31)(32). Because the Pem gene is not cell type-specific and its introns appear to be typical, we hypothesized that Pem would be a good candidate to provide information on the metabolism of mammalian introns in general.…”