Presence of LH receptor mRNA in granulosa cells as a potential marker of oocyte developmental competence and characterization of the bovine splicing isoforms.
As the expression of the LH receptor (LH-R) in granulosa cells is thought to be associated with later stages of folliculogenesis, this study was undertaken to evaluate the presence of LH-R mRNA as a suitable marker for developmental competence of oocytes. Granulosa cells and cumulus-oocyte complexes (COCs) were recovered from cows that had received ovarian stimulation. The COCs were subjected to embryo production procedures in vitro to assess the embryonic potential of the oocyte, and the corresponding granulosa cells were used to evaluate the presence of LH-R mRNA by RT-PCR. The presence of LH-R transcripts in granulosa cells is not a key characteristic of a follicle bearing a competent oocyte, although a higher proportion of oocytes reach the blastocyst stage when LH-R mRNA is detected in the granulosa cells. Different LH-R isoforms were cloned and sequence discrepancies among six of the isoforms enabled the design of specific oligonucleotides to study the presence of the isoforms in different follicular cells. All LH-R transcripts studied and the 80 kDa protein product corresponding to the full length receptor were found in granulosa cells of small (< 4 mm) and large (> 5 mm) follicles. When the granulosa cells were cultured, the transcripts were downregulated by the culture conditions; downregulation was more acute in granulosa cells from small follicles. The addition of LH to the culture media enhanced LH-R mRNA downregulation. The presence of several LH-R transcript isoforms was tissue specific and in the theca cells LH-R mRNA was restricted mainly to cells from larger follicles. This finding indicates that the expression and the splicing of LH-R mRNA are regulated in a cell-specific and follicular size-specific manner.
Robert
C
C
Centre de Recherche en Biologie de la Reproduction, Department of Animal Sciences, Laval University, QC, Canada G1K 7P4.
Gagné
D
D
Lussier
J G
JG
Bousquet
D
D
Barnes
F L
FL
Sirard
M-A
MA
D017398:Alternative Splicing; D000818:Animals; D015415:Biomarkers / Q000032:analysis; D001755:Blastocyst / Q000166:cytology; D015153:Blotting, Western / Q000379:methods; D002417:Cattle; D002478:Cells, Cultured; D005260:Female; D005307:Fertilization in Vitro; D006107:Granulosa Cells / Q000737:chemistry*; D009866:Oogenesis; D006080:Ovarian Follicle / Q000502:physiology; D020033:Protein Isoforms / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis*; D011974:Receptors, LH / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015415:Biomarkers; D020033:Protein Isoforms; D012333:RNA, Messenger; D011974:Receptors, LH