Concanavalin A-Induced Morphological Changes and Its Binding Sites in Starfish Follicle Cells as Revealed by Electron Microscopy

Concanavalin A (Con A) stimulates the production in starfish follicle cells of 1-methyladenine, a hormone which induces oocyte maturation. We have therefore investigated Con A-induced morphological changes and Con A-binding sites in the follicle cell using native Con A and horseradish peroxidase- or ferritin-labeled Con A (HRP-Con A, Fer-Con A). After isolated follicle cells were incubated with Con A (1 mg/ml), vacuoles, the Golgi complex and multivesicular body-like organelles (MVBs) became prominent in most of the cells. After follicle cells were prefixed and then incubated with Fer-Con A for 60 min, tagged ferritin was diffusely and randomly distributed as single or small clustered particles on the cell surface. The incubation of isolated follicle cells with Fer-Con A for 10 min before fixation resulted in numerous ferritin particles localized along the internalized membrane, and also in vacuoles, MVBs and small lysosome-like structures. After 60 min incubation with Fer-Con A, ferritin was further located in large lysosome-like structures and in vesicles near and in the Golgi area as well as in the organelles described above. HRP-Con A binding sites were also observed in vacuoles and MVBs of the intact cells.
These results suggest that Con A binds at first to the cell surface and causes rapid internalization and that membrane-bound Con A is easily endocytosed into vacuoles, MVBs and lysosome-like structures, and is later incorporated in some vesicles in the Golgi area.     

MECHANISM OF STARFISH SPAWNING. III. PROPERTIES AND ACTION OF MEIOSIS-INDUCING SUBSTANCE PRODUCED IN GONAD UNDER INFLUENCE OF GONAD-STIMULATING SUBSTANCE

The gonad-stimulating substance (GSS) contained in nerve extract of the starfish, Asterina pectinifera, acts on the ovary to produce an active substance responsible for oocyte maturation, meiosis-inducing substance (MIS). MIS was successfully separated from GSS by gel-filtration on Sephadex. The MIS fraction had also spawning-inducing activity. Starfish testis also produced MIS under the influence of GSS. MIS production was shown in six starfish species. Although some species specificity characterizes GSS, this was not observed in MIS. Production of MIS in the ovary was found to begin immediately after the addition of GSS. The amount of MIS produced increased as the concentration of GSS was raised, and the longer the time of treatment with GSS, the greater was the amount of MIS produced. MIS was rather heatstable, insoluble in ether, benzene and petroleum ether but soluble in 95% ethanol. Its activity was not destroyed by pronase. Injection of MIS into the coelomic cavity induced gamete-shedding in both male and female. However, MIS failed to induce spawning when applied from the outside of the body. Both GSS and MIS were found in the coelomic fluid only when the starfish were undergoing natural spawning. A mechanism of starfish spawning, based on the action of GSS and MIS is discussed.     

Uptake and Metabolism of Sugars by Suspension cultured Catharanthus roseus Cells

p. 186, right column line 11 ‘27.2 KBq’ change to‘37.2 MBq’ p. 187, left column line 8 ‘27.2 KBq’ change to‘37.2 MBq’ line 10 ‘13.6 KBq’ change to ‘18.6 MBq’ line 11 ‘13.6 KBq’ change to ‘18.6 MBq’ line 21 ‘89%’ change to ‘80%’ right column line 24 ‘CLC-NH₄’ change to ‘CLC-NH₂’ P. 189, Table 1 appeared incorrectly: it should appear as indicated.     

Phylogeny of ground beetles subgenus Nialoe (s. lat.) Tanaka (Coleoptera: Carabidae; genus Pterostichus): A molecular phylogenetic approach

We constructed a phylogeny of the ground beetle subgenus Nialoe ( s. lat. ), genus Pterostichus (Coleoptera: Carabidae) based on two mitochondrial (cytochrome oxidase I and 16S ribosomal DNA) and one nuclear (28S ribosomal DNA) gene sequences. Thirty-three representative species of the group and three outgroup species were analyzed. The resultant trees (maximum parsimonious, maximum likelihood and Bayesian trees of the combined data of the three gene sequences) indicated that there are two large and three small lineages in the group, some of which were supported by a previous morphology-based phylogeny. In all the analyses, the small lineage composed of two Korean species is sister to the rest of the subgenus, but relationships of other four lineages differed among the analyses and remained unresolved. The implications of the present results are discussed in terms of taxonomy and biogeography of the group.     

EFFECTS OF TAURODEOXYCHOLATE AND CALCIUM ON CONTRACTILITY OF NEWT EGGS

Injection of taurodeoxycholate (TDOC) alone or in combination with 10⁻⁵ M Ca²⁺(Ca-EGTA buffer) into newt eggs induced a cup- or furrow-like depression on the egg surface. Reduction of the Ca²⁺ concentration inhibited the response. These findings imply that TDOC induces a cytoplasmic contraction associated with the membrane and that a micromolar Ca²⁺ concentration is required for this process. Injection of 10⁻⁵ M Ca²⁺(Ca-EGTA buffer) alone had no effect. On the basis of these findings, the roles of TDOC and Ca²⁺ in induction of contraction were discussed.     

Gene flow between two closely related species in the acuticollis species group (Coleoptera: Lucanidae: genus Platycerus) near their distribution border based on morphology and mitochondrial gene

The male endophallic morphology and mitochondrial cytochrome oxidase subunit I (COI) gene of two parapatric species of the acuticollis species group, Platycerus acuticollis and P. albisomni, were examined at the border between the two species. No morphologically inferred hybrid was found. The endophallic and mitochondrial types of each individual were significantly linked, and the two species inferred by both types were essentially parapatric even at a very local level. We inferred that the reproductive isolation between them is high, although introgressive hybridization occurred rarely. The taxonomical assignment of P. acuticollis and P. albisomni as two distinct species is considered appropriate.     

PRESENCE OF 1-METHYLADENINE IN SEA URCHIN GONAD AND ITS RELATION TO OOCYTE MATURATION

A water extract of sea urchin ovary was found to induce maturation of starfish oocytes in vitro . The presence of the active substance was demonstrated in ovaries of the sea urchins, Pseudocentrotus depressus, Anthocidaris crassispina and Hemicentrotus pulcherrimus , and the sand dollars, Clypeaster japonicus and Peronella japonica . The active substance was also contained in the testes of these echinoids. That the content of this substance increases during the reproductive season was demonstrated with Anthocidaris gonads. The active substance present in ovary or testis of the sea urchins was successfully extracted with 85% ethanol and purified with gel-filtrations on Sephadex G-15 columns after washing with chloroform and ether. The purified active substances were the same and were identified as 1-1-methyladenine by thin layer chromatography. 1-Methyladenine was found to be effective in inducing oocyte maturation in Anthocidaris crassispina in vitro . Therefore, 1-methyladenine seems to play an important role in oocyte maturation in echinoids as well as in asteroids.     

Uptake and Metabolism of Sugars by Suspension Cultured Catharanthus roseus Cells

The uptake and metabolism of sugars by suspension-cultured Catharanthusroseus cells were investigated. Substantially all the sucrosein the culture medium was hydrolyzed to glucose and fructosebefore being taken up by the cells. The activity of invertasebound to cell walls, determined in situ, was high at the earlystage of culture. Glucose was more easily taken up by the cellsthan was fructose. Tracer experiments using [U-¹⁴C]glucose and[U-¹⁴C]fructose indicated that glucose is a better precursorfor respiration than fructose, while fructose is preferentiallyutilized for the synthesis of sucrose, especially in the earlyphase of cell growth. Possible metabolic routes of sugar insuspension-cultured Catharanthus roseus cells are discussedin the context of these results. Catharanthus roseus, Madagascar periwinkle, suspension culture, sucrose, glucose, fructose, metabolism, glycolysis     

Effect of 17β-Estradiol on Growth of Oocytes in Cultured Ovarian Fragments of the Starfish, Asterina Pectinifera

The development of growing oocytes of the starfish, Asterina pectinifera , was divided into five stages according to their histological features. Specimens collected monthly from Rishiri Island throughout a year were used. The effect of 17 β-estradiol on oocyte growth was investigated. When ovarian fragments taken from April females were cultured in artificial seawater containing Medium 199 and streptomycin sulfate for 3 days, oocytes within such fragments did not show any changes in diameter. However, when similar ovarian fragments were cultured in the presence of 10⁻⁵M 17 β-estradiol, the oocyte diameters increased significantly. It is concluded that 17 β-estradiol enhances the growth of starfish oocytes in cultured ovarian pieces.