Equine P450 Cholesterol Side-Chain Cleavage and 3β-Hydroxysteroid Dehydrogenase/Δ5-Δ4 Isomerase: Molecular Cloning and Regulation of Their Messenger Ribonucleic Acids in Equine Follicles During the Ovulatory Process1
Bibliographic record
Abstract
The preovulatory LH rise is the physiological trigger of follicular luteinization, a process during which the synthesis of progesterone is markedly increased. To study the control of follicular progesterone biosynthesis in mares, the objectives of this study were to clone and characterize the equine cholesterol side-chain cleavage cytochrome P450 (P450(scc)) and 3 beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase (3 beta-HSD), and describe the regulation and cellular localization of their transcripts in equine follicles during hCG-induced ovulation. Complementary DNA cloning and primer extension analyses revealed that the equine P450(scc) transcript is composed of a 5'-untranslated region (UTR) of 52 nucleotides, an open reading frame (ORF) of 1560 nucleotides, and a 3'-UTR of 225 nucleotides, whereas the equine 3 beta-HSD mRNA consists of a 5'-UTR of 61 nucleotides, an ORF of 1119 nucleotides, and a 3'-UTR of 432 nucleotides. The equine P450(scc) and 3 beta-HSD ORF encode 520 and 373 amino acid proteins, respectively, that are highly conserved (68-79% identity) when compared to homologs of other mammalian species. Northern blot analyses were performed with preovulatory follicles isolated 0, 12, 24, 30, 33, 36, and 39 h post-hCG, and corpora lutea obtained on day 8 of the cycle. Results showed that levels of P450(scc) mRNA in follicular wall (theca interna with attached granulosa cells) decreased after hCG treatment (30-39 h versus 0 h post-hCG, P: < 0.05), and increased again after ovulation to reach their highest levels in corpora lutea (P: < 0.05). Northern blots on isolated cellular preparations revealed that theca interna was the predominant site of P450(scc) expression in follicles prior to hCG (P: < 0.05). However, transcript levels decreased in theca interna between 30-39 h (P: < 0.05) and increased in granulosa cells at 39 h (P: < 0.05), making the granulosa cell layer the predominant site of P450(scc) expression at the end of the ovulatory process. A different pattern of regulation was observed for 3 beta-HSD, as transcript levels remained constant throughout the luteinization process (P: > 0.05). Also, in contrast to other species, expression of 3 beta-HSD mRNA in equine preovulatory follicles was localized only in granulosa cells and not in theca interna. Thus, this study characterizes for the first time the complete structure of equine P450(scc) and 3 beta-HSD mRNA and identifies novel patterns of expression and regulation of these transcripts in equine follicles prior to ovulation.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".