Evidence for the participation of two sperm proteases, spermosin and acrosin, in fertilization of the ascidian, Halocynthia roretzi: Inhibitory effects of leupeptin analogs on enzyme activities and fertilization

Sawada, Hitoshi; Yokosawa, Hideyoshi; Someno, Tetsuya; Saino, Tetsushi; Ishii, Shin‐ichi

doi:10.1016/0012-1606(84)90281-1

Cited by 51 publications

(24 citation statements)

References 15 publications

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“…We have previously reported that two trypsin-like sperm proteases, acrosin and spermosin (9)(10)(11)(12)(13)(14), and sperm proteasomes (15,16) play key roles in sperm penetration through the vitelline coat of the ascidian H. roretzi. We provided evidence that the 930-kDa proteasome, which has a 26S-proteasome-like subunit composition, functions as a lysin that directly degrades the vitelline coat (16): the 26S-like proteasome was found in the supernatant obtained by centrifugation of the alkaline seawatertreated sperm and exhibited a vitelline coat-degrading activity.…”

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confidence: 99%

Extracellular ubiquitination and proteasome-mediated degradation of the ascidian sperm receptor

Sawada

Sakai

Abe

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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128

125

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The ubiquitin-proteasome system is essential for intracellular protein degradation, but an extracellular role of this system has not been known until now. We have previously reported that the proteasome is secreted into the surrounding seawater from sperm of the ascidian (Urochordata) Halocynthia roretzi on sperm activation, and that the sperm proteasome plays a key role in fertilization. Here, we show that a 70-kDa component (HrVC70) of the vitelline coat is the physiological substrate for the ubiquitin-proteasome system during fertilization of H. roretzi. A cDNA clone encoding the HrVC70 precursor (HrVC120) was isolated, and a homology search revealed that HrVC120 contains 13 epidermal growth factor-like repeats and a mammalian zona pellucida glycoprotein-homologous domain. HrVC70 functions as a sperm receptor. We demonstrate that HrVC70 is ubiquitinated both in vitro and in vivo. The immunocytochemical localization of multiubiquitin chains in the vitelline coat and the inhibitory effect of monoclonal antibodies against the multiubiquitin chains on fertilization strongly support the role of the ubiquitin-proteasome system in ascidian fertilization. Taken together, these results indicate that the ubiquitin-proteasome system is responsible for extracellular degradation of the sperm receptor HrVC70 and, consequently, for sperm penetration of the vitelline coat during fertilization.fertilization ͉ vitelline coat ͉ ubiquitin F ertilization is a key event in sexual reproduction, creating a new individual with novel genomic information. In animal reproduction, species-specific binding of sperm to the proteinaceous egg coat, called the vitelline coat in marine invertebrates or the zona pellucida in mammals, is particularly important for successful fertilization. Because the egg coat is a potential barrier to sperm-egg fusion, sperm must use a lytic agent (lysin) to penetrate it (1, 2). In mammals, it has long been believed that the sperm acrosomal trypsin-like protease acrosin is a zona lysin, which digests zona pellucida proteins to enable sperm to penetrate through the zona pellucida (3). However, recent studies by using acrosin-knockout mice revealed that acrosin is not essential for fertilization or sperm penetration through the zona pellucida (4). Rather, it is currently thought that acrosin is involved in the dispersal of acrosomal proteins during acrosome reaction (5) and that a sperm protease(s) other than acrosin is the actual zona-lysin in mammalian fertilization (6).Ascidians (Urochordata) occupy a phylogenetic position between invertebrates and segmented vertebrates. They are hermaphrodites that usually release sperm and eggs simultaneously during the spawning season. Several ascidians, including Ciona intestinalis (7) and Halocynthia roretzi (8), strictly prohibit selffertilization. Because self-nonself recognition in fertilization is accomplished by interaction between the sperm and vitelline coat (7,8), the sperm-lysin system seems to be triggered after the sperm recognizes the vitelline coat as nonself...

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confidence: 99%

Extracellular ubiquitination and proteasome-mediated degradation of the ascidian sperm receptor

Sawada

Sakai

Abe

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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128

125

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“…At the chorion, recognition and binding of the gametes take place (Rosati et al, 1978;Honegger, 1986); moreover the participation of an acrosomal reaction is disputed (Woollacott, 1977;Fukumoto, 1988). Sawada et al (1984) discovered that sperm proteases are involved in fertihzation. The modification of the shape of the egg after fertilization has been investigated by Sawada and Osanai in Ciona intestinalis.…”

Section: Discussionmentioning

confidence: 99%

Fertilization and normal development inAscidiella aspersa (Tunicata) studied with Nomarski-optics

Niermann-Kerkenberg

Hofmann

1989

Helgolander Meeresunters

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Normal development of Ascidiella aspersa was studied over a period of approx. 24 h at 20 ~ from egg insemination through metamorphosis of the tadpole larva using Nomarski-optics. Records were made of spermatozoa attaching to and passing through the cellular envelopes and the chorion of the egg. Egg shape alterations upon entry of the fertilizing sperm, which reflect the early phase of ooplasmic segregation, were monitored in intact and dechorionated eggs. The time course of normal development was recorded, and prominent stages were photographed within or deprived of the egg envelopes. The present observations are compared with recent accounts on early development in other solitary ascidian species.

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“…They are indispensable for the sperm to penetrate through the egg envelope (22)(23)(24). FUKUMOTO (25) suggests that the fussy extracellular material at the tip of the sperm head may be the site where these lysins exist.…”

Section: Sectionmentioning

confidence: 99%

Cytological Characterization of Self Incompatibility in Gametes of the Ascidian, Ciona intestinalis

1987

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We have determined how many elements are involved in the regulation of self-fertilization in the solitary ascidian, Ciona intestinalis that is an incompletely self-sterile species. Animals collected in the field were repeatedly induced to spawn in order to examine their selfing ratios. About 20% of them were self-fertile, although the ratios fluctuated considerably among respective spawnings. Naturally or acid-induced self-fertile gametes required much longer time for selfing than that for crossing. Egg-suspending seawater (egg water) as such activated sperm motility, but it lowered conspicuously the self-fertilization ratio. Self-sterile spermatozoa could scarcely bind to the vitelline coat (VC) of glycerinated autologous eggs, or in case they bound well to it the sperm flagella ceased to beat within five min of the 'insemination'. The staining of sectioned gametes with DAPI, a fluorescent dye of DNA, showed that in selfing the spermatozoa could hardly penetrate the VC even though they bound well to it. The results of this study show that the block of self-fertilization can be classified into four elements from a phenomenal viewpoint, such as egg water, low affinity of sperm-VC binding, inactivation of bound sperm and difficulty in sperm penetration through the VC.

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Evidence for the participation of two sperm proteases, spermosin and acrosin, in fertilization of the ascidian, Halocynthia roretzi: Inhibitory effects of leupeptin analogs on enzyme activities and fertilization

Cited by 51 publications

References 15 publications

Extracellular ubiquitination and proteasome-mediated degradation of the ascidian sperm receptor

Extracellular ubiquitination and proteasome-mediated degradation of the ascidian sperm receptor

Fertilization and normal development inAscidiella aspersa (Tunicata) studied with Nomarski-optics

Cytological Characterization of Self Incompatibility in Gametes of the Ascidian, Ciona intestinalis

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