[Frontiers in Bioscience 1, d146-160, August 1, 1996]
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CAVEAT LECTOR



INTERACTION OF HUMAN SPERMATOZOA WITH THE ZONA PELLUCIDA OF OOCYTE: DEVELOPMENT OF THE ACROSOME REACTION

Patricio Morales1 and Miguel Llanos2

1 P. Catholic University of Chile, Faculty of Biological Sciences. P. O. Box 114-D. Santiago, Chile.

2INTA, University of Chile, P. O. Box 138-11. Santiago, Chile.

3. THE ZONA PELLUCIDA, ITS COMPOSITION AND ROLE DURING GAMETE INTERACTION AND FERTILIZATION

All mammalian eggs are surrounded by the ZP, an extracellular coat that is synthesized by the oocyte (1). The ZP is the site of the initial interaction of the spermatozoa with the oocyte. This interaction includes the species-specific spermatozoa-ZP binding and induction of the AR, both of which are prerequisites for successful in-vivo fertilization.

Results of cDNA cloning of the ZP genes and analysis of the composition of ZP from several different species indicate that ZP is constituted of three or four glycoproteins (2). The ZP of the mouse oocyte, one of the best studied ZP, is composed of three sulphated glycoproteins termed ZP1, ZP2 and ZP3 (1, 3). ZP1 is a homodimer (Mr= 185-200 kDa) and its chains are connected by intermolecular disulphide bonds. ZP2 (Mr= 120-140 kDa) and ZP3 (Mr= 83 kDa) form a heterodimer of long filaments with a repeating structure (1, 3). ZP1 provides a structural integrity for the ZP by cross-linking the ZP2/ZP3 filaments. Only ZP2 and ZP3 have been shown to possess biological functions. ZP3 mediates the initial binding of acrosome-intact spermatozoa to the ZP via O-linked side chains (4, 5). Following sperm binding, ZP3 induces the AR in the bound spermatozoa (6). The acrosome-reacted spermatozoa, which can no longer interact with ZP3, bind to ZP2 (7) and penetrate through the ZP. After fertilization, there are molecular changes in ZP2 and ZP3 that constitute a block to polyspermy. ZP3 is converted to a form called ZP3f, which no longer binds acrosome-intact spermatozoa and is incapable to induce the AR (6). Since O-linked carbohydrates are implicated in interaction of ZP3 with spermatozoa and there is no apparent change in the electrophoretic mobility of ZP3f (6, 8), this change in ZP3 is thought to be caused by a cortical granule-released glycosidase (9). ZP2 is converted to a form called ZP2f that no longer interacts with acrosome-reacted spermatozoa (10). ZP2 is cleaved by a protease from the cortical granules (11) and is detected by a shift in its electrophoretic mobility (from Mr= 120 kDa to Mr= 90 kDa) under reducing conditions (12).

In contrast to these findings in the mouse, there are only a few reports regarding the composition of the human ZP (13-17). Shabanowitz and colleagues reported only two components (Mr= 90-110 kDa and Mr= 57-73 kDa) under nonreducing conditions and three components (Mr= 90-110 kDa, Mr= 65-78 kDa, and Mr= 57-73 kDa) under reducing conditions (16, 17). They termed these proteins ZP1, ZP2 and ZP3, respectively. A ZP component corresponding to the mouse ZP1 (Mr= 200 kDa) was not detected. Similarly, under reducing conditions Bercegeay et al. found protein components of 80-92 kDa, 58-66 kDa, and 54-72 kDa (13). Based upon their molecular weight these components could correspond to mouse ZP2 and ZP3, respectively. Naz and Ahmad showed that the human ZP analyzed under non-reducing conditions exhibited 3 major protein bands of 220, 110 and 55 kDa. The ZP protein that reacted strongest with the sperm proteins was the 55 kDa molecular region (ZP3) (15). Recently, Moos et al. using a non-radioactive biotinylation- and a lectin-based detection system found that, under non-reducing conditions, the human ZP of unfertilized eggs is composed of three glycoprotein species designated as ZP1 (Mr~150 kDa), ZP2 (Mr~100 kDa) and ZP3 (Mr~55-65 kDa) (14).

In all the above studies, ZP1 was not detected after fertilization. In contrast, in the mouse ZP1 is present and is apparently unaltered following fertilization (12). Therefore, it has been suggested that in humans, the cortical granule-derived proteases may degrade ZP1 to forms that are not detectable by electrophoresis (14).

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