The activity for the induction of the sperm acrosome reaction localises in the outer layers and exists in the high-molecular-weight components of the egg-jelly of the newt, Cynops pyrrhogaster

Abstract

Localisation of the acrosome reaction inducing activity in egg-jelly was examined in the newt, Cynops pyrrhogaster. The jelly has six layers: the J0, J1, J2, J3, J4 and st layers. Jelly was mechanically dissected and placed on a Millipore filter. When sperm were added from the outer surface side of the jelly, most of them exhibited the acrosome reaction after passing through the jelly. When egg-jelly was divided into four layers, strong activity for the induction of acrosome reaction was detected in the outer layers, J4+st. These findings suggest that the acrosome reaction is induced by a substance in the outer layers of the egg-jelly. Among jelly components separated by SDS-PAGE, a fraction of more than 500 kDa in molecular weight induced the acrosome reaction. Wheat germ agglutinin (WGA), Griffonia simplicifoliar agglutinin 1 (GS-1), Maclura pomifera agglutinin (MPA) and Arachis hypogaea agglutinin (PNA) inhibited the induction of the acrosome reaction by jelly extract, and WGA did so in a dose-dependent manner. Those lectins precipitated some molecules of over 500 kDa. These results suggest that the acrosome reaction is induced by the high molecular-weight components of egg-jelly in C. pyrrhogaster.

Identification of the sperm motility-initiating substance in the newt, Cynops pyrrhogaster, and its possible relationship with the acrosome reaction during internal fertilization

“…This layer harbors the acrosome reaction-inducing activity (Sasaki et al, 2002), and is crucial for the success of internal fertilization (Takahashi et al, 2006). Based upon our present data, the SMIS-localizing granular structures are covered with a sheet-like structure in which ARIS localizes (Figs.…”

“…Acrosome reaction was induced according to the method described previously (Sasaki et al, 2002;Watanabe et al, 2009). JE was pretreated with 100 μg/ml α34 mAb for 30 min.…”

“…The pars convoluta is divided into several compartments, each of which secretes specific substances to form several morphologically and biochemically different layers in egg-jelly Okimura et al, 2001). The egg-jelly plays a pivotal role in amphibian fertilization through a variety of active substances such as sperm chemoattractant in Xenopus (Al-Anzi and Chandler, 1998;Olson et al, 2001), an inducer for capacitation-like change in Bufo sperm (Krapf et al, 2007), a mediator for sperm binding to the vitelline envelope in Cynops (Hiyoshi et al, 2007), a sperm acrosome-stabilizing factor in Bufo (Krapf et al, 2006), and an acrosome reaction-inducing substance in Cynops (Sasaki et al, 2002;Watanabe et al, 2009).…”

“…The outermost layer of the egg-jelly is crucial for internal fertilization in the Japanese newt, Cynops pyrrhogaster (Takahashi et al, 2006) through both sperm motility-initiating (Ukita et al, 1999; and acrosome reactioninducing activities (Sasaki et al, 2002;Watanabe et al, 2009). The former of these activities is induced by the sperm motility-initiating substance (SMIS), a heat-stable proteinaceous factor (Mizuno et al, 1999;Onitake et al, 2000).…”

Characteristics of Sperm Motility Induced on the Egg-Jelly in the Internal Fertilization of the Newt, Cynops pyrrhogaster

“…The amphibian eggjelly is composed of several layers with a stable fibrous-network containing diffusible globular-molecules (Bonnell et al , 1996;Watanabe and Onitake, 2002). As in other species, it plays an important role in the sperm-egg interaction (Ishihara et al , 1984;Al-Anzi and Chandler, 1998;Ukita et al , 1999;Onitake et al , 2000;Sasaki et al , 2002). In the newt, Cynops pyrrhogaster , the egg-jelly is composed of six sublayers, each of which contains unique carbohydrate components (Okimura et al ., 2001;Watanabe and Onitake, 2002).…”