Expression of the full-length and alternatively spliced equine luteinizing hormone/chorionic gonadotropin receptor mRNAs in the primary corpus luteum and fetal gonads during pregnancy

Abstract

The full-length equine luteinizing hormone/chorionic gonadotropin (LH/CG) receptor (eLH/CG-R A ) cDNA and two alternatively spliced isoforms (eLH/CG-R B,C ) were isolated from luteal tissue and characterized using a combination of reverse transcription-polymerase chain reaction (RT-PCR) and 5 0 -rapid amplification of cDNA ends. The 680-amino acid full sequence of eLH/CG-R A displayed 87 -92% homology with other mammalian LH/CG-Rs. The eLH/CG-R B and eLH/CG-R C cDNA isoforms were truncated from the 3 0 -end of exon X: eLH/CG-R B spliced out of frame into the last exon whereas eLH/CG-R C contained an in-frame stop codon within a divergent sequence. Consequently, both eLH/CG-R B and eLH/CG-R C cDNA isoforms encoded putative proteins without transmembrane and intracellular domains. In order to study the responsiveness of the primary corpus luteum (CL) and fetal gonads to eCG, the expression of eLH/CG-R mRNAs was examined by RT-PCR and Northern blot analysis during early and mid-pregnancy. All three eLH/CG-R cDNA isoforms (eLH/CG-R A,B,C ) were expressed from day 14 to day 83 of pregnancy in the primary CL and from day 44 to day 222 in fetal gonads. Interestingly, the primary CL at days 89 and 151 expressed only truncated eLH/CG-R cDNA isoforms. The relative values of Northern hybridized major 7, 5.7, 3.9 and 1.8 kb eLH/CG-R mRNA transcripts tended to decrease in the primary CL whereas the unique major 1.8 kb eLH/CG-R mRNA was steadily expressed in fetal gonads during pregnancy. These results show that the expression of eLH/CG-R mRNAs occurs in the fetal gonads before ceasing in the primary CL and suggest that eCG may be involved in the gradual transition from a luteal to a feto-placental output of steroids during equine pregnancy.

Natural and recombinant equine chorionic gonadotropins past and future in animal reproductive technology

“…One of the characteristics of eCG is a long half-life: 5 hours in rats, 21 hours in sheep, and 45.6 hours in cattle (Murphy and Martinuk, 1991). The most important role of eCG in mares is to bind with the LH receptors in corpora lutea to maintain the pregnancy over the first period of gestation (Saint-Dizier et al, 2004). Around week 5 of gestation, accessory corpora lutea appear in mare ovaries.…”

Cloning and functional expression of the equine luteinizing hormone/chorionic gonadotrophin receptor

“…First-strand cDNA was amplified for 30 cycles using Advantage 2 polymerase mix (BD Biosciences) and primer pair P3 and P4 in the PCR conditions described above. Finally, the 5 -end of the eLH/CG-R cDNA (cDNA fragment 3) was isolated using the antisense primer P5 designed from the cDNA fragment 1 (Table 1) and the rapid amplification of cDNA ends technique, as described previously (Saint-Dizier et al 2004). Table 1 Nucleic acid sequences of primers used for the isolation and cloning of the eLH/CG-R cDNA.…”