Using combinations of reverse transcription-polymerase chain reaction (RT-PCR) and 5′- and 3′-rapid amplification of cDNA ends, three different, alternatively spliced, partial turkey LH receptor (tLH-R) cDNA isoforms were characterized from ovarian mRNA. The first cDNA (tLH-Rintact) showed 98% and 72–75% similarity with chicken and mammalian LH-R sequences, respectively. The second cloned cDNA isoform (tLH-Rinsert) contained an in-frame TGA stop codon within an 86-base pair insertion that was located in the extracellular domain of the seven-transmembrane region. The tLH-Rinsert isoform could encode a truncated soluble protein isoform that lacked the transmembrane region. The third cDNA isoform truncated the transmembrane region (tLH-Rtrunc) and was derived by the deletion of the last exon by incomplete splicing. Generation of multiple transcripts by alternative splicing was elucidated by partial characterization of tLH-R genomic sequences.
The differentially regulated expression of the tLH-R mRNA isoforms in nongonadal tissues and ovarian stromal tissues during various reproductive stages was quantified and analyzed by Northern blot and/or RT-PCR. Alternatively spliced tLH-R isoforms were differentially expressed in a tissue-specific manner in most of the tissues examined. The steady-state levels of tLH-R mRNA isoforms were relatively high in the hypothalamus and optic nerve and relatively low in the cortex, pituitary, and cerebellum when compared to levels in ovarian follicles. In nongonadal reproductive tissues, the steady-state levels of tLH-R mRNA isoforms were relatively high in the uterus and infundibulum and relatively low in the isthmus, oviduct, and magnum. In addition, in the nongonadal peripheral tissues, the steady-state levels of tLH-R isoforms were relatively high in the thyroid gland and relatively low in the spleen, adrenal gland, kidney, skin, bursa, and muscle.
The present study suggests that the alternative splicing of LH-R transcripts occurs in a tissue-specific manner and has been evolutionarily conserved (similar results were obtained in chicken and swine). These results raise fundamental questions as to the function of LH-R isoforms in nongonadal tissues.
