Molecular cloning and partial characterization of medaka fish stromelysin-3 and its restricted expression in the oocytes of small growing follicles of the ovary.

A cDNA clone (2755-bp) for stromelysin-3 was isolated by screening the cDNA library and by 3'- and 5'-rapid amplification of cDNA ends using ovary RNA of the medaka fish Oryzias latipes. The clone encodes a protein of 492 amino acids. Stromelysin-3 mRNA was detected only in the ovary. In situ hybridization analysis revealed that stromelysin-3 mRNA was localized in the oocyte cytoplasm of small growing follicles. RT-PCR analysis of total RNAs isolated from various-sized follicles and ovulated oocytes was conducted in order to determine the mRNA levels during oocyte growth. The stromelysin-3 mRNA level was the highest in the small follicles, and the mRNA levels decreased as the follicles grew. No significant stromelysin-3 mRNA was detected in the ovulated oocytes or immature ovaries. The fish stromelysin-3 cDNA was expressed in COS-1 cells in order to characterize the intracellular localization of the protein. A 56 kDa protein was synthesized and secreted into the culture medium. The secreted stromelysin-3 exhibited gelatin-degrading activity.