Characterization of a chicken luteinizing hormone receptor (cLH-R) complementary deoxyribonucleic acid, and expression of cLH-R messenger ribonucleic acid in the ovary.

Abstract

Studies were conducted to characterize the chicken ovarian LH receptor (cLH-R) cDNA and to evaluate expression of cLH-R mRNA in follicles at different stages of development. A total of 1.89 kb of nucleic acid sequence corresponding to the cLH-R (1.79 kb of the predicted coding region) was isolated by a combination of reverse transcription-polymerase chain reaction and cDNA library screening techniques. Also of interest was the finding that two of three positive clones isolated from the hen ovarian cDNA library contained an 86-bp insert located in the extracellular domain within 69 bp of the putative transmembrane domain. This insert contains an inframe TGA stop codon, suggesting that an alternatively spliced transcript results in translation of a truncated protein corresponding to the extracellular domain of the cLH-R. Considering all protein domains thus far characterized, the deduced amino acid sequence of the cLH-R shares 73.2% and 74.2% identity with the rat and porcine LH-R sequences, respectively, with highest homology occurring within the seven transmembrane spanning regions (86-88% identity vs. mammalian sequences). Northern blot analysis determined that cLH-R mRNA levels in the theca layer tend to increase through follicle development to the second largest (F2) preovulatory follicle (p = 0.084), and to decrease in the largest preovulatory (F1) follicle (p < 0.02 vs. F2). By comparison, cLH-R mRNA levels are nondetectable (by Northern blot analysis) in granulosa cells from prehierarchal (3-8-mm diameter) follicles. Constitutive expression of cLH-R mRNA in granulosa cells is first detectable at the 9-12-mm diameter stage of follicle development, and levels are further increased in cells from large preovulatory (F1, F2, and F3) follicles (p < 0.01 vs. 9-12-mm stage). Collectively, these results are consistent with previous observations that granulosa cells from prehierarchal follicles fail to produce cAMP or steroids in response to short-term incubation with ovine LH, in vitro, and that granulosa cells acquire LH responsiveness only subsequent to follicle selection into the rapid growth phase.