Equine P450 cholesterol side-chain cleavage and 3 beta-hydroxysteroid dehydrogenase/delta(5)-delta(4) isomerase: molecular cloning and regulation of their messenger ribonucleic acids in equine follicles during the ovulatory process.

This research examined the molecular structure and regulation of two enzymes believed to be vital for hormone synthesis in horses, particularly during ovulation. It sheds light on the differences in enzyme activity before and during ovulation, which provides valuable insights into the reproductive process of mares.

Research Purpose and Approach

• The primary aim of the study was to explore the control of follicular progesterone biosynthesis in mares. Progesterone is a crucial hormone involved in the ovulation process.
• The specific enzymes the team was concerned with were cholesterol side-chain cleavage cytochrome P450 (P450scc), and 3 beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase (3 beta-HSD). These enzymes play vital roles in cholesterol and steroid metabolism, respectively, leading to the production of progesterone.
• The researchers cloned and characterized these enzymes, while also tracking their regulation and cellular localization in equine follicles during hCG-induced ovulation. The researchers used Northern blot analyses for this purpose, a common technique for studying gene expression.

Main Findings

• The researchers discovered the full molecular structure (“transcript”) of both P450scc and 3 beta-HSD in horses for the first time.
• They found that the transcript of P450scc consisted of a 5′ untranslated region of 52 nucleotides, an open reading frame of 1560 nucleotides, and a 3′ untranslated region of 225 nucleotides. Whereas 3 beta-HSD’s transcript had a 5′ untranslated region of 61 nucleotides, an open reading frame of 1119 nucleotides, and a 3′ untranslated region of 432 nucleotides.
• The enzyme P450(scc) was predominantly expressed prior to hCG in the theca interna (a layer of the follicle). However, its expression decreased between 30-39 hours after hCG treatment and increased in granulosa cells (another follicle layer) at 39 hours.
• Meanwhile, 3 beta-HSD showed a constant expression level throughout the luteinization (hormone synthesis) process.
• The researchers also found that the expression pattern for the 3 beta-HSD mRNA in equine preovulatory follicles was unique, being localized only in granulosa cells and not in theca interna, unlike in other species.

Significance of the Research

• This study contributes to a better understanding of the equine ovulatory process at the molecular level.
• The findings could potentially provide new ways to manage and optimize equine reproduction.