Three different Turkey luteinizing hormone receptor (tLH-R) isoforms II: Characterization of differentially regulated tLH-R messenger ribonucleic acid isoforms in the ovary

We have recently characterized three different, alternatively spliced, partial turkey LH receptor (tLH-R) cDNA isoforms by the combination of reverse transcription-polymerase chain reaction (RT-PCR) and 5'- and 3'-rapid amplification of cDNA ends. The first cDNA (intact form: tLH-R(intact)) showed 98% and 72-75% similarity with chicken and mammalian LH receptor sequences, respectively. The other two cloned cDNA isoforms (insertion and truncated forms: tLH-R(insert) and tLH-R(trunc)) could encode truncated soluble protein isoforms that lack the transmembrane region. Northern blot analysis detected two transcripts of 3.0 kilobases (kb) (tLH-R(intact)) and 1.5 kb (tLH-R(trunc)) in the turkey ovary but could not discriminate a third alternatively spliced transcript (tLH-R(insert)) due to the small 86-base pair difference in the size range of approximately 3.0-kb mRNAs. But with the combination of RNase protection assay, RT-PCR, and Northern blot analysis, three different alternatively spliced tLH-R mRNA isoforms were quantified. Differential expression of the tLH-R mRNA isoforms was demonstrated in ovarian stromal tissue during various reproductive stages and in the theca and granulosa layer through follicular development. To gain a better understanding of the physiological significance of the three different tLH-R isoforms, total RNA from the theca layer through follicular development after prolactin (PRL) treatment was analyzed by RT-PCR. PRL treatment for 8-14 days significantly increased the steady-state levels of total tLH-R mRNAs, including tLH-R(insert) and tLH-R(trunc) mRNAs, compared to those in nontreated controls. In contrast, the steady-state levels of tLH-R(intact) mRNA during the same period was not significantly changed when compared to that in nontreated controls. The present study shows that tLH-R transcripts are alternatively spliced in a tissue-specific manner in the turkey and that the mechanism may, in part, be controlled hormonally.