Insulin stimulates placental leucine aminopeptidase/oxytocinase/insulin-regulated membrane aminopeptidase expression in BeWo choriocarcinoma cells.

Abstract

Placental leucine aminopeptidase (P-LAP), a cystine aminopeptidase that is identical to insulin-regulated membrane aminopeptidase, hydrolyzes oxytocin, which results in the loss of oxytocin activity. We previously isolated genomic clones containing the human P-LAP promoter region, which included two sites homologous to the 10-bp-insulin responsive element (IRE) that was identified on the phosphoenolpyruvate carboxinase gene. We therefore postulated that insulin regulates P-LAP expression via these IREs and investigated this notion using BeWo choriocarcinoma trophoblastic cells cultured in the presence of insulin. Insulin increased P-LAP activity in a time- and dose-dependent manner. Physiological concentrations of insulin at 10(-7) M exhibited the most potent effect on P-LAP activity. Western blotting demonstrated that 10(-7) M insulin increased P-LAP protein levels. Semi-quantitative RT-PCR and Southern blotting showed that insulin also increased P-LAP mRNA, which was abrogated by prior exposure to cycloheximide. Luciferase assay did not reveal any regulatory regions within 1.1 kb upstream of the P-LAP gene that could explain the insulin-induced P-LAP mRNA accumulation. These findings indicate that insulin induces P-LAP expression in trophoblasts, and that it acts via de novo synthesis of other proteins, which partially contradicts our initial hypothesis.

• Authors: Masayuki Nakata, Seiji Nomura, Yoko Ikoma, Seiji Sumigama, Fumi Shido, Tomomi Ito, Mayumi Okada, Fumitaka Kikkawa, Masafumi Tsujimoto, Shigehiko Mizutani
• Journal: Regulatory peptides (Regul Pept) Vol. 117 Issue 3 Pg. 187-93 (Mar 15 2004) ISSN: 0167-0115 [Print] Netherlands
• PMID: 14749039 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Insulin
• RNA, Messenger
• Cycloheximide
• Luciferases
• Cystinyl Aminopeptidase
• leucyl-cystinyl aminopeptidase

Topics

• Base Sequence
• Cell Membrane (enzymology, metabolism)
• Choriocarcinoma
• Cycloheximide (pharmacology)
• Cystinyl Aminopeptidase (genetics, metabolism)
• Dose-Response Relationship, Drug
• Female
• Humans
• Insulin (pharmacology, physiology)
• Luciferases (metabolism)
• Molecular Sequence Data
• Promoter Regions, Genetic
• RNA, Messenger (metabolism)
• Time Factors
• Trophoblasts (enzymology)
• Uterine Neoplasms