Submitted on April 11, 2008
Revised on August 25, 2008
Accepted on August 28, 2008
Oxytocin and vasopressin gene expression and RNA splicing patterns in the rat supraoptic nucleus
Chunmei Yue1, Todd A Ponzio1, Raymond L Fields1, and Harold Gainer1*
1 National Institutes of Health
* To whom correspondence should be addressed. E-mail: gainerh{at}mail.nih.gov.
In this paper, we test the hypothesis that there are differential splicing patterns between the expressed oxytocin (OT) and vasopressin (VP) genes in the rat supraoptic nucleus (SON). We quantified the low abundance, intron-containing heteronuclear RNAs (hnRNAs) and the higher abundance mRNAs in the SON using two-step, quantitative SYBR Green real-time reverse transcription (RT)-PCR and external standard curves constructed using synthetic 90nt sense-strand oligonucleotides.The levels of OT and VP mRNA in the SON were found to be similar, around 108 copies/SON pair, whereas the copy numbers of VP hnRNAs containing intron 1 or 2 and the OT hnRNA containing intron 1 are much lower, i.e., about 102 to103 copies/rat SON pair. However, the estimated copy number of the intron 2-containing OT hnRNA is much larger, about106 copies/SON pair. The relative distributions of all the OT and VP RNA species were invariant and independent of the physiological status of the rats (e.g, osmotically stimulated or lactating rats). Using intron-specific riboprobes against hnRNAs, we demonstrate by fluorescence in situ hybridization (FISH) strong signals of OT hnRNA containing intron 2 predominantly in the cytoplasm, in contrast to the localization of the VP hnRNA found only in the nuclei. Taken together, these data support the view that the splicing patterns between OT and VP gene transcripts are different, and show that there is a selective cytoplasmic retention of OT intron 2.
