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Involvement of proteases in sperm-egg interaction in ciona intestinals
522
523
YOLK-ORGANELLE MEMBRANES OF X E N O P U S O O C Y T E S H.-P. R i c h t e r a n d H. Erempel, FR 3.2 Physiologie, D-6650 Homburg/S. F.R.G. Y o l k - p l a t e l e t m e m b r a n e s of X e n o p u s have b e e n s t u d i e d by t h i n - s e c t i o n and freeze-fracture electron microscopy and by gel e l e c t r o p h o r e s i s . In sectioned vitellogenic oocytes, the large yolk platelets (YP) are in c]ose membrane contact with each o t h e r or w i t h the two t y p e s of primo r d i a l y o l k (pYP I and pYP II). A f t e r membrane fusion these precursors are c o m p l e t e l y i n c o r p o r a t e d into the YPs. Lipoid d r o p l e t s are a t t a c h e d to the YPs i n d i c a t i n g lipid uptake across the m e m b r a n e . F r e e z e - e t c h e d y o l k m e m branes represent lntra-membraneous particles of v a r i o u s size and of a h e t e r o g e n e o u s d i s t r i b u t i o n of 2 0 0 - 6 0 0 I M P s / ~ m = at the E-face, and 1 2 0 0 - 2 1 0 0 I M P s / u m = at the P-face, T h e s e p r e s e n t a t i o n s of the m e m b r a n e n e i t h e r e x h i bit a n a s t o m o s e s to ER or d i c t y o s o m e s , excluding a contribution of these c o m p a r t m e n t s to YP f o r m a t i o n . An isol a t i o n p r o c e d u r e leads to fully g r o w n YPs and from that to p u r i f i e d platelet m e m b r a n e s . T h e s e have been subjected to EM and S D S - P A G E to detect intrinsic transport orot~in~
MORPHOLOGICAL EVINDENCE FOR GERM PLASM IN PRIMODIAL GERM CELLS (PGCs) OF CHICKEMBRYO L. Rongxiu. Chinese Society of Dev. Biologist, Shenyang Agriculture University, Shenyang Liaoning Province, People's Rep. of China. The ultrastructural features of the PGCs in the germinal crescent area were observed in the chick embryos. The PGCs were characterized by: i) round or oval shape, 13-16 m., oval and dented nucleus; 2) the presence of pseudopodia and microvilli; 3) more abundant and mature oncocells as compared with other embryonic development; 4) the presence of desmosome-like structure connecting adjacent PGCs and abundant micro filaments. The author has observed also the special granular material (electron-dense bodies) in the PGCs. These electrorr-dense bodies were demonstrated to be derived from the nucleus, and transferred via the inter-nuclear and extranuclear membrane into the cytoplasm forming the cytoplasm in the form of gathering electrondense bodies. The special granular material would be germ granules and the special cytoplasm would be the germinal plasm.
524
525
OOCA~E MATURATION (MEIOSIS) AND FERTILIZATION IN GYNOGENETIC TELEOSTS
INVOLVEMENT OF PROTEASES IN SPERM-EGG INTERACTION IN CIONA INTESTINALIS. R. De Santis and M.R. Pinto. Stazione Zoologica di Napoli, Italy and Institute of Protein Biochemistry and Enzymology, Arco Felice, Napoli, Italy. In the ascidian Ciona intestinalis,the vitelline coat of the egg is the site of the species-specific binding between spermatozoa and the egg. Following binding the spermatozoon passes through the coat. In other ascidians spermatozoa utilize t ryspin-like and chymotrypsin-like proteinases as lysin(s) to facilitate passage through the investments. Using warlous inhibitors and substrates of proteinases we have exa1~med fertilization of C~ona intestinalis for involvement of lyelns in sperm penetration. Among all the inhibitors tested only chymostatin, a chymotrypsin inhibitor, completely blocks fertilization. Fertilization is not affected by trypsin inhibitors. A pr elimir~ry screening with specif ic substrates of chymotrypsin parallels the results obtained with chymostatin. Since binding seems not to be affected by these substrates, the results point to the involvement of sperm chymot rypsin-like enzymes in sperm penetration through the vitelline coat.
Saat, P. Tartu State University, Department of Zoology, 18 Ulikooli St., Tartu, Estonian SSR, 202400 USSR Morphology and chronology of oocyte maturation in vitro was investigated in triploid forms of Carassius auratus gibello and Cobitis-complex. In Carassius, the reduction division is omitted. In Cobitis, two meiotic divisions occurred and therefore a supplementary endomitosis in oogerresis is presumed. In Carassius, the formation of male pronucleus was never observed either in normal or in pelyspermic (after dechorionization) eggs. Nevertheless, the cytoplasm of Eynogenetic eggs possesses factors responsible for male pronucleus formation since injected demembranized sperm heads transformed into male pronuclei. According to preliminary TEM data, the action of these factors is prevented by surrounding incorporated sperm heads with membranous structures. In all triploid Cobitis females, only about 50% of eggs were gynogenetie.
S159
523
YOLK-ORGANELLE MEMBRANES OF X E N O P U S O O C Y T E S H.-P. R i c h t e r a n d H. Erempel, FR 3.2 Physiologie, D-6650 Homburg/S. F.R.G. Y o l k - p l a t e l e t m e m b r a n e s of X e n o p u s have b e e n s t u d i e d by t h i n - s e c t i o n and freeze-fracture electron microscopy and by gel e l e c t r o p h o r e s i s . In sectioned vitellogenic oocytes, the large yolk platelets (YP) are in c]ose membrane contact with each o t h e r or w i t h the two t y p e s of primo r d i a l y o l k (pYP I and pYP II). A f t e r membrane fusion these precursors are c o m p l e t e l y i n c o r p o r a t e d into the YPs. Lipoid d r o p l e t s are a t t a c h e d to the YPs i n d i c a t i n g lipid uptake across the m e m b r a n e . F r e e z e - e t c h e d y o l k m e m branes represent lntra-membraneous particles of v a r i o u s size and of a h e t e r o g e n e o u s d i s t r i b u t i o n of 2 0 0 - 6 0 0 I M P s / ~ m = at the E-face, and 1 2 0 0 - 2 1 0 0 I M P s / u m = at the P-face, T h e s e p r e s e n t a t i o n s of the m e m b r a n e n e i t h e r e x h i bit a n a s t o m o s e s to ER or d i c t y o s o m e s , excluding a contribution of these c o m p a r t m e n t s to YP f o r m a t i o n . An isol a t i o n p r o c e d u r e leads to fully g r o w n YPs and from that to p u r i f i e d platelet m e m b r a n e s . T h e s e have been subjected to EM and S D S - P A G E to detect intrinsic transport orot~in~
MORPHOLOGICAL EVINDENCE FOR GERM PLASM IN PRIMODIAL GERM CELLS (PGCs) OF CHICKEMBRYO L. Rongxiu. Chinese Society of Dev. Biologist, Shenyang Agriculture University, Shenyang Liaoning Province, People's Rep. of China. The ultrastructural features of the PGCs in the germinal crescent area were observed in the chick embryos. The PGCs were characterized by: i) round or oval shape, 13-16 m., oval and dented nucleus; 2) the presence of pseudopodia and microvilli; 3) more abundant and mature oncocells as compared with other embryonic development; 4) the presence of desmosome-like structure connecting adjacent PGCs and abundant micro filaments. The author has observed also the special granular material (electron-dense bodies) in the PGCs. These electrorr-dense bodies were demonstrated to be derived from the nucleus, and transferred via the inter-nuclear and extranuclear membrane into the cytoplasm forming the cytoplasm in the form of gathering electrondense bodies. The special granular material would be germ granules and the special cytoplasm would be the germinal plasm.
524
525
OOCA~E MATURATION (MEIOSIS) AND FERTILIZATION IN GYNOGENETIC TELEOSTS
INVOLVEMENT OF PROTEASES IN SPERM-EGG INTERACTION IN CIONA INTESTINALIS. R. De Santis and M.R. Pinto. Stazione Zoologica di Napoli, Italy and Institute of Protein Biochemistry and Enzymology, Arco Felice, Napoli, Italy. In the ascidian Ciona intestinalis,the vitelline coat of the egg is the site of the species-specific binding between spermatozoa and the egg. Following binding the spermatozoon passes through the coat. In other ascidians spermatozoa utilize t ryspin-like and chymotrypsin-like proteinases as lysin(s) to facilitate passage through the investments. Using warlous inhibitors and substrates of proteinases we have exa1~med fertilization of C~ona intestinalis for involvement of lyelns in sperm penetration. Among all the inhibitors tested only chymostatin, a chymotrypsin inhibitor, completely blocks fertilization. Fertilization is not affected by trypsin inhibitors. A pr elimir~ry screening with specif ic substrates of chymotrypsin parallels the results obtained with chymostatin. Since binding seems not to be affected by these substrates, the results point to the involvement of sperm chymot rypsin-like enzymes in sperm penetration through the vitelline coat.
Saat, P. Tartu State University, Department of Zoology, 18 Ulikooli St., Tartu, Estonian SSR, 202400 USSR Morphology and chronology of oocyte maturation in vitro was investigated in triploid forms of Carassius auratus gibello and Cobitis-complex. In Carassius, the reduction division is omitted. In Cobitis, two meiotic divisions occurred and therefore a supplementary endomitosis in oogerresis is presumed. In Carassius, the formation of male pronucleus was never observed either in normal or in pelyspermic (after dechorionization) eggs. Nevertheless, the cytoplasm of Eynogenetic eggs possesses factors responsible for male pronucleus formation since injected demembranized sperm heads transformed into male pronuclei. According to preliminary TEM data, the action of these factors is prevented by surrounding incorporated sperm heads with membranous structures. In all triploid Cobitis females, only about 50% of eggs were gynogenetie.
S159