MORPHOLOGY OF GAMETE MEMBRANE FUSION AND OF SPERM ENTRY INTO OOCYTES OF THE SEA URCHIN

Sea urchin gametes predominate in molecular studies of fertilization, yet relatively little is known of the subcellular aspects of sperm entry in this group. Accordingly, it seemed desirable to make a detailed examination of sperm entry phenomena in sea urchins with the electron microscope. Gametes of the sea urchins Arbacia punctulata and Lytechinus variegatus were used in this study. Samples of eggs containing 2 to 8 per cent oocytes were selected and fixed with osmium tetroxide in sea water at various intervals after insemination. Fixed specimens were embedded in Epon 812, sectioned, and examined with an electron microscope. An apical vesicle was observed at the anterior end of the acrosome. The presence of this structure, together with other observations, suggested that initiation of the acrosome reaction in sea urchin sperm involves dehiscence of the acrosomal region with the subsequent release of the acrosomal granule. Contact and initial fusion of gamete membranes was observed in mature eggs and oocytes and invariably involved the extended acrosomal tubule of the spermatozoon. Only one spermatozoon normally enters the mature egg. The probability of locating such a sperm in ultrathin sections is exceedingly low. Several sperm do normally enter oocytes. Consequently, observations of sperm entry were primarily restricted to the latter. The manner of sperm entry into oocytes did not resemble phagocytosis. Organelles of the spermatozoon were progressively divested of their plasma membrane as they entered the ground cytoplasm of the oocyte fertilization cone. Initiation of the acrosome reaction, contact and initial fusion of gamete membranes, and sperm entry into oocytes of sea urchins conform to the Hydroides-Saccoglossus pattern of early fertilization events as described by Colwin and Colwin (13).     

Zur Normalen Histologie des Nasenscheidewandgerüstes

Zusammenfassung An der Hand von neun F?llen wurde die kn?cherne, und knorpelige Nasenscheidewand einer genaueren histologischen Untersuchung unterzogen. Dabei wurde nicht nur der Gewebsaufbau des Vomer und der Lamina perpendicularis einerseits und der der Cartilago quadrangularis andererseits genauer beschrieben, sondern die überaus verwickelten Verh?ltnisse der Nahtverbindungen zwischen allen diesen Bestandteilen der Nasenscheidewand einer genaueren Bearbeitung unterzogen. Das Verwickelte dieser Verh?ltnisse beruht darauf, da? die drei Hauptbestandteile der Scheidewand einen sehr verschiedenen Werdegang hinter sich haben; so stellt die Cartilago quadrangularis den ursprünglichen embryonalen Zustand der Nasenscheidewand dar, die Lamina perpendicularis entsteht durch enchondrale Verkn?cherung der knorpeligen Scheidewand und der Vomer entsteht als perichondraler Belegknochen der letzteren. Jeder einzelne dieser drei verschiedenartigen Bestandteile der Nasenscheidewand hat eine Verbindung mit den beiden anderen und daraus ergibt sich eine alle M?glichkeiten ersch?pfende Mannigfaltigkeit der Verbindungsart dieser Bestandteile. Dazu kommt noch, da? an der enchondralen Verbindungsgrenze zwischen Lamina perpendicularis und Cartilago quadrangularis der Zustand, wie er nach abgeschlossener enchondraler Vérkn?cherung bestanden hat, dem ganz anders gearteten, einer Nahtverbindung Platz macht und endlich sind auch die Verh?ltnisse im Bereiche des Processus sphenoidalis besonders verwickelter Art. Auch derHuschkesche Knorpel wird besprochen und überdies ein offenbar bisher noch nicht bekannter, bereits im Gebiet der Choanen gelegener knorpeliger Bestandteil der Nasenscheidewand. Wegen der Einzelheiten sei auf das Original verwiesen. Mit 19 Textabbildungen.     

Denaturation and Renaturation of Viral Ribonucleic Acid I. Annealing R17 Ribonucleic Acid with Denatured Replicative Form or with Denatured Replicative Intermediate

Purified replicative form (RF) and replicative intermediate (RI) prepared from Escherichia coli infected with R17 were denatured in 0.15 m NaCl, 0.015 m sodium citrate containing 65% dimethylsulfoxide. Denaturation of RF or RI was demonstrated spectrophotometrically, chromatographically, and by sedimentation analysis. Denatured RF or RI was annealed by carefully decreasing the temperature from 62 to 20 C. Annealing was accompanied by a decreased absorbance at 260 mmu. The decrease in absorbance during annealing appeared to be dependent upon the rate of cooling and the concentration of ribonucleic acid (RNA). Denatured RF or RI was annealed with R17 RNA which was labeled with (3)H-uridine. The annealed product was 73 to 82% resistant to 0.1 mug/ml of ribonuclease. Annealing R17 RNA with either denatured RF or RI resulted in the formation of a ribonuclease-resistant product with a sedimentation profile resembling that of native RI. Melting the annealed products in 85.7% dimethyl sulfoxide produced 27S single-stranded R17 RNA and a heterogeneous population of more slowly sedimenting RNA.     

Purification and Properties of Reovirus Ribonucleic Acid

NaClO(4) was employed in a technique for the rapid extraction of reovirus ribonucleic acid (RNA). The extracted RNA, which was purified in a Cs(2)SO(4) equilibrium density gradient, had a buoyant density of 1.61 g/cm(3) and a sedimentation coefficient of 15S in a 7 to 20% sucrose gradient. It was 90% resistant to ribonuclease in a solution of high ionic strength (0.1 m NaCl). The sensitivity of reovirus RNA to ribonuclease increased with decreasing ionic strength. The thermal denaturation transition of the RNA began at 78 C and was complete at 85 C. The T(m) of the transition was 81 C in 0.01 m tris(hydroxymethyl)aminomethane buffer (pH 7.2) containing 0.001 m ethylenediaminetetraacetate. Thermal denaturation of reovirus RNA resulted in the formation of three ribonuclease-sensitive fractions. Denaturation at 25 C in the presence of dimethyl sulfoxide resulted in the formation of two ribonuclease-sensitive fractions.     

Referate

Ohne Zusammenfassung