Identification and tissue distribution of a novel variant of 11 beta-hydroxysteroid dehydrogenase 1 transcript.
A novel variant of 11 beta-hydroxysteroid dehydrogenase 1 (11 beta-HSD1) mRNA was identified from the ovine liver by reverse transcription-polymerase chain reaction (RT/PCR), and was named 11 beta-HSD1C mRNA. Sequence analysis of the RT-PCR product revealed that 11 beta-HSD1C mRNA was the product of an alternative exon-splicing within the 11 beta-HSD1 gene in which exon 5 was spliced out. Although it caused a deletion of 48 amino acids in the deduced 11 beta-HSD1 protein, this alternative splicing did not result in a shift within the predicted open reading frame of 11 beta-HSD1 cDNA. Thus, 11 beta-HSD1C mRNA was predicted to code for a protein of 244 amino acids. Using RT-PCR, we also examined the expression of 11 beta-HSD1C mRNA in ovine fetal organs and in maternal myometrium, endometrium, chorion, amnion and placenta. The 11 beta-HSD1C mRNA was expressed ubiquitously, similar to 11 beta-HSD1A mRNA, but at a lower abundance. Furthermore, since levels of 11 beta-HSD1C mRNA were directly related to those of 11 beta-HSD1A mRNA, there is no tissue-specificity for this shorter transcript and the only factor regulating its production appears to be 11 beta-HSD1A mRNA itself. To determine whether 11 beta-HSD1C mRNA encoded a functional enzyme, we inserted the cDNA into the expression vector pRc/CMV, and transfected the construct into Chinese hamster ovary cells. The transfected cells expressed a mRNA of expected size but contained no detectable 11 beta-HSD activity. When combined with cellular extracts of 11 beta-HSD1A cDNA transfected cells, they also did not alter either the dehydrogenase or reductase activity. The functional significance of the 11 beta-HSD1 transcript lacking exon 5 (11 beta-HSD1C mRNA) remains to be determined.
D042842:11-beta-Hydroxysteroid Dehydrogenases; D017398:Alternative Splicing; D000650:Amnion / Q000201:enzymology; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D016466:CHO Cells; D002823:Chorion / Q000201:enzymology; D006224:Cricetinae; D017931:DNA Primers; D004717:Endometrium / Q000201:enzymology; D005091:Exons; D005260:Female; D005333:Fetus; D015870:Gene Expression; D014644:Genetic Variation; D006913:Hydroxysteroid Dehydrogenases / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D009215:Myometrium / Q000201:enzymology; D016366:Open Reading Frames; D009928:Organ Specificity; D010920:Placenta / Q000201:enzymology; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis; D012756:Sheep; D014158:Transcription, Genetic; D014162:Transfection
D017931:DNA Primers; D012333:RNA, Messenger; D011994:Recombinant Proteins; D006913:Hydroxysteroid Dehydrogenases; D042842:11-beta-Hydroxysteroid Dehydrogenases