IDENTIFICATION OF CYSTATIN AS A COMPONENT OF CARP CHORION

An ovary-specific cystatin is immunocytochemically demonstrated to be localized in the chorions, cortical granules, and yolk granules of car p oocytes, as well as in the follicle cells surrounding oocytes. Durin g cortical reaction, cystatin is exocytosed from cortical granules int o the perivitelline space. In situ hybridization confirms that cystati n is synthesized by oocytes and follicle cells. Western blotting revea ls that chorion cystatin appears in multiple bands of high molecular w eight (from 65 kDa to larger than 200 kDa). No cystatin monomer of 14 kDa is found. These results indicate that chorion cystatin is conjugat ed with other chorion components. Two forms of conjugates are found. I n one form, cystatin, ZP2, fibroin-like substance (FLS], and cathepsin -like substance (CLS) ave conjugated, which is extracted by sodium dod ecyl sulfate. In the other form, cystatin, FLS, and CLS are conjugated , which is extracted by guanidine thiocyanate (GTC). Most chorion cyst atin of oocytes and ovulated eggs is solubilized by GTC, while a large amount of cystatin remains in the fertilization envelope of cortical reacted eggs after extraction by GTC. Mel. Reprod. Dev. 51:430-435, 19 98. (C) 1998 Wiley-Liss, Inc.