Saint-Dizier Marie, Chopineau Maryse, Dupont Joëlle, Combarnous Yves
Unité de Physiologie de la Reproduction et des Comportements, UMR 6175 INRA-CNRS-Université F. Rabelais de Tours-Haras Nationaux, 37 380 Nouzilly, France.
Reproduction. 2004 Aug;128(2):219-28. doi: 10.1530/rep.1.00164.
The full-length equine luteinizing hormone/chorionic gonadotropin (LH/CG) receptor (eLH/CG-RA) cDNA and two alternatively spliced isoforms (eLH/CG-RB,C) were isolated from luteal tissue and characterized using a combination of reverse transcription-polymerase chain reaction (RT-PCR) and 5'-rapid amplification of cDNA ends. The 680-amino acid full sequence of eLH/CG-RA displayed 87-92% homology with other mammalian LH/CG-Rs. The eLH/CG-RB and eLH/CG-RC cDNA isoforms were truncated from the 3'-end of exon X: eLH/CG-RB spliced out of frame into the last exon whereas eLH/CG-RC contained an in-frame stop codon within a divergent sequence. Consequently, both eLH/CG-RB and eLH/CG-RC 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-RA,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.
从黄体组织中分离出全长马促黄体生成素/绒毛膜促性腺激素(LH/CG)受体(eLH/CG-RA)cDNA及两种可变剪接异构体(eLH/CG-RB、C),并通过逆转录-聚合酶链反应(RT-PCR)和5'-cDNA末端快速扩增相结合的方法对其进行了表征。eLH/CG-RA的680个氨基酸的完整序列与其他哺乳动物的LH/CG-Rs显示出87%-92%的同源性。eLH/CG-RB和eLH/CG-RC cDNA异构体从外显子X的3'端被截断:eLH/CG-RB框外剪接到最后一个外显子中,而eLH/CG-RC在一个不同序列中包含一个框内终止密码子。因此,eLH/CG-RB和eLH/CG-RC cDNA异构体编码的推定蛋白均无跨膜和细胞内结构域。为了研究初级黄体(CL)和胎儿性腺对eCG的反应性,在妊娠早期和中期通过RT-PCR和Northern印迹分析检测了eLH/CG-R mRNA的表达。在妊娠第14天至第83天,初级CL中表达了所有三种eLH/CG-R cDNA异构体(eLH/CG-RA、B、C),在胎儿性腺中则从第44天至第222天表达。有趣的是,在第89天和第151天的初级CL中仅表达截短的eLH/CG-R cDNA异构体。在初级CL中,Northern杂交的主要7、5.7、3.9和1.8 kb eLH/CG-R mRNA转录本的相对值趋于下降,而在妊娠期间,胎儿性腺中独特的主要1.8 kb eLH/CG-R mRNA稳定表达。这些结果表明,eLH/CG-R mRNA的表达在胎儿性腺中发生,然后在初级CL中停止,这表明eCG可能参与了马妊娠期间从黄体类固醇分泌向胎儿-胎盘类固醇分泌的逐渐转变。