Identification of hepsin and protein disulfide isomerase A3 as targets of gelatinolytic action in rat ovarian granulosa cells during the periovulatory period

Katherine Rosewell, Linah Al-Alem, Feixue Li, Brian Kelty, Thomas E. Curry

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

The matrix metalloproteinase (MMP) family is believed to play a role in the ovulatory process because MMP inhibitors block oocyte release. However, little is known about the mechanisms by which the MMPs affect ovulation. The present study investigated the degradomic actions of the gelatinases, MMP2 and MMP9, by identifying gelatinolytic targets in periovulatory granulosa cells. Granulosa cells were collected from immature rats 48 h after equine chorionic gonadotropin treatment and were cultured with human chorionic gonadotropin (hCG) in the absence or presence of a specific MMP2/9 inhibitor ((2R)-2-[(4-biphenylylsulfonyl)a- mino]-3-phenylpropionic acid) for an additional 24 h. The conditioned media was analyzed for gelatinolytic activity, progesterone, and peptide profiles. Gelatinolytic activity and progesterone were induced in response to hCG; however, there was no difference in progesterone between cells treated with or without the inhibitor. Peptide fragments of proteins altered in the presence of the gelatinase inhibitor were identified by two- dimensional gel electrophoresis and mass spectrometry. Protein disulfide isomerase A3 (PDIA3), which plays a role in protein folding, was identified as a peptide that decreased in the presence of inhibitor while the serine protease hepsin, was found to increase with inhibitor treatment. Subsequent experiments established that PDIA3 and hepsin were targets of MMP2/9 action by cleavage with MMP2 and Western blot analysis, respectively. Additionally, hepsin was identified as a gelatinolytic target in ovarian cancer cells. In the present study, proteomics has identified proteins that may be involved in novel ways in the complex cascades that are mediated by gelatinolytic MMPs during the periovulatory period.

Original languageEnglish
Pages (from-to)858-866
Number of pages9
JournalBiology of Reproduction
Volume85
Issue number4
DOIs
StatePublished - Oct 1 2011

Keywords

  • Corpus luteum
  • Hepsin
  • Matrix metalloproteinase
  • Ovulation
  • Protein disulfide isomerase A3

ASJC Scopus subject areas

  • Reproductive Medicine
  • Cell Biology

Fingerprint

Dive into the research topics of 'Identification of hepsin and protein disulfide isomerase A3 as targets of gelatinolytic action in rat ovarian granulosa cells during the periovulatory period'. Together they form a unique fingerprint.

Cite this