MAGNESIUM REQUIREMENT IN FERTILIZATION PROCESS OF SEA URCHINS

The Mg²⁺ requirement in fertilization was investigated in sea urchins. It was found that when sea urchin eggs were inseminated in sea water free of Mg²⁺, little fertilization took place. Even when spermatozoa pre-treated with dissolved egg-jelly to induce the acrosome reaction, which needs Ca²⁺, were used, the fertilization rate remained quite low in the absence of Mg²⁺. In Strongylo-centrotus intermedius , the lowest concentration of Mg²⁺ required for 50% fertilization was 0.05 mM in the presence of 10 mM Ca²⁺, whereas that of calcium was 3 mM in the presence of 49 mM Mg²⁺. These critical concentrations increased when the concentration of the other ion decreased. Removal of Mg²⁺ or Ca²⁺ or both from the suspending medium had little adverse effect on sperm motility. The elevation of the fertilization membrane was also induced by butyric acid independent of the presence or absence of Mg²⁺ and/or Ca²⁺. These results indicate that Mg²⁺ are required at least in some process(es) between acrosome reaction and fertilization membrane elevation, such as sperm penetration or membrane fusion.     

SPECIES-SPECIFIC ADHESION OF SPERMATOZOA TO THE SURFACE OF FIXED EGGS IN SEA URCHINS

When the spermatozoa of sea urchins are added to eggs which have been fixed with glutaraldehyde and washed thoroughly, the spermatozoa swarm around the eggs and adhere to the egg surface. The mode of sperm adhesion to the fixed egg is assumed, on the evidence of electron-microscopical studies, to be the same as that of adhesion to the intact egg at the initial stage of normal fertilization. The spermatozoa and fixed eggs of five species of sea urchins were combined and heterologous crosses were studied. Species-specific adhesion of sperm to fixed eggs was clearly demonstrated. There is a direct relationship between the cross-fertilization of living gametes and the binding capacity of spermatozoa and fixed eggs in so far as the employed five species are concerned.     

Accumulation of Trehalose as a Compatible Solute under Osmotic Stress in Euglena gracilis Z

ABSTRACT. The photosynthetic protozoon Euglena gracilis, accumulated a large amount of trehalose in the cells under salt or osmotic stresses. Radioactivity of [14C] paramylon, a β-1,3-polyglucan which was stored in the cells of E. gracilis. was degraded rapidly and this radioactivity was almost stoichiometrically incorporated into trehalose. The interconversion of trehalose from paramylon by salt or osmotic stresses was dependent on the concentrations or osmotic pressures, suggesting that E. gracilis accumulate trehalose as an osmoprotectant. After the removal of salt or osmotic stresses, trehalose was gradually degraded, however, it was not converted into paramylon.     

Phylogenetic analysis of the Brazilian microendemic Paepalanthus subgenus Xeractis (Eriocaulaceae) inferred from morphology

Paepalanthus subgenus Xeractis (Eriocaulaceae) comprises 28 recognized species endemic to the Espinhaço Range, in Minas Gerais state, Brazil. Most species of the subgenus are restricted to small localities and critically endangered, but still in need of systematic study. The monophyly of the subgenus has already been tested, but only with a few species. Our study presents the first phylogenetic hypothesis within the group, based on morphology. A maximum parsimony analysis was conducted on a matrix of 30 characters for 30 terminal taxa, including all species of the subgenus and two outgroups. The biogeographical hypotheses for the subgenus were inferred based on dispersal–vicariance analysis (DIVA). The analysis provided one most‐parsimonious hypothesis that supports most of the latest published subdivisions (sections and series). However, some conflicts remain concerning the position of a few species and the relationships between sections. The distribution and origin(s) of microendemism are also discussed, providing the ground for conservation strategies to be developed in the region. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 167 , 137–152.     

Calcium Deposition in Idioblasts of Mulberry Leaves

Large, rounded idioblasts were observed in adaxial leaves ofmulberry plants; they were clearly distinguishable from epidermal,trichome and parenchyma cells. The size and density of idioblastsvaried according to leaf age. Cytological features of idioblastswere as follows: the outermost region (‘cap’) ofidioblasts was situated on the adaxial surface as a dome-likeprotrusion; a cylindrical protuberance extended from the capregion to the inner part of the idioblast; in idioblasts frommature leaves a crystal mass was suspended from the lower tipof the cylindrical protuberance. Elemental analysis of idioblastsdemonstrated that silicon (Si) was localized in both the capregion and the cylindrical protuberance but calcium (Ca) waspresent in the large crystal, indicating site-specific cellularlocalization of Ca and Si within an idioblast. Histochemicalassays showed that a distinct Ca crystal filled the vacuolesof idioblasts in mature leaves, while immature leaves had manyidioblasts without Ca deposition. The increase in the Ca contentof leaves was directly proportional to the increase in leafage and appeared to be closely related to the Ca sink capacityof the developing idioblast vacuoles. The maximum sink capacitywas quantified to be approximately 40 ng per idioblast whenmulberry plants were grown hydroponically with excess Ca.Copyright1999 Annals of Botany Company Morus alba, idioblast, Ca deposition, Ca sink capacity, silicon, X-ray microanalysis, histochemistry, scanning electron microscopy.     

蓖麻蚕卵黄原蛋白cDNA的克隆及序列分析

本文论述了蓖麻蚕卵黄原蛋白cDNA迅速克隆化的方法,SDS－PAGE鉴定的蓖麻蚕卵黄原蛋白由大小二个亚基构成,求出的分子量大亚基为180kDa,小亚基为45kDa,从解析的蓖麻蚕卵黄原蛋白氨基酸序列推算的分子量大亚基为161．06kDa,小亚基为40．53kDa,如果考虑到翻译后的修饰,这与SDS－PAGE求出的分子量是吻合的。蓖麻蚕卵黄原蛋白cDNA是由5788pb构成,一个ORF为5337个碱基,编码了1779个氨基酸。在信号肽的15个氨基酸残基中,有12个是硫水性氨基酸残基。这与其他昆虫卵黄原蛋白信号肽区域的硫水性分析是一致的。蓖麻蚕卵黄原蛋白N-linked glycosylation site的分布与家蚕、柞蚕和天蚕不同,3处N-linked glycosylation site存在于大亚基里,2处地多聚丝氨酸区域上流的小亚基里。另外,我们发现在所解析的柞蚕、天蚕的蓖麻蚕卵黄原蛋白氨基酸序列的C－末端区域里DGQR、GICG功能部位及其后的6个半胱氨酸都完好地保存。     

天蚕卵黄原蛋白cDNA的克隆及序列分析

天蚕卵黄原蛋白cDNA的全碱基序列由5 720个碱基构成,一个开读框有5 334个碱基序列,编码了包括由15个氨基酸组成的信号肽在内的共1 778个氨基酸的蛋白质前体。天蚕和柞蚕卵黄原蛋白的同源性非常高,氨基酸序列达到92.6%,碱基序列达到94%。天蚕、柞蚕和家蚕的卵黄原蛋白的糖链附加位点(N-linkedg-lycosylationsite):柞蚕有4个,家蚕有5个,天蚕和家蚕相同保存着5个。在N-末端区域,天蚕和家蚕有多聚丝氨酸区域和可被胰蛋白酶识别的RSRR部位;柞蚕虽然也具有多聚丝氨酸区域,但没有可被胰蛋白酶识别的RSRR部位。在C-末端区域里,大多数昆虫从G ICG功能部位至C-末端结尾具有7个~10个半胱氨酸(Cys)。鳞翅目的4种昆虫柞蚕、天蚕、家蚕和舞毒蛾(A.pernyi,A.yam am ai,B.m ori,L.dispar)都是7个半胱氨酸。     

Calcium- and Cyclic AMP-independent,Labile Protein Kinase Appearing during Starfish Oocyte Maturation: its Extraction and Partial Characterization*

A burst of protein phosphorylation and an appearance of maturation-promoting factor have been reported to occur shortly before germinal vesicle (nucleus) breakdown (GVBD) in 1-methyladenine-induced oocyte maturation of starfish. To detect if a protein kinase is activated before GVBD, protein kinase activity was compared in maturing oocytes which were just undergoing GVBD and immature oocytes of Asterina pectinifera. The oocytes were homogenized in a buffer modified from that used for extracting amphibian maturation-promoting factor. When the supernatant protein of homogenized immature oocytes was used as a substrate, protein kinase activity in the supernatant of the maturing oocytes was 7-fold higher than that of immature oocytes. The protein kinase in the supernatant of the maturing oocytes showed a high substrate specificity for histone H1 among the exogenous substrates examined, and the activity of the maturing oocytes for histone H1 was 6- to 7-fold higher than that of immature oocytes. The protein kinase detected in the maturing oocytes was very labile and was inhibited neither by ethylene glycol bis(β-aminoethyl ether)N, N, N′, N′-tetraacetic acid nor by the heat-stable inhibitor protein of cyclic AMP-dependent protein kinase. These results indicate that a calcium- and cyclic AMP-independent, labile “maturation-specific protein kinase” appeared before GVBD in maturing oocytes, and suggest its participation in the phosphorylation burst in vivo. The possible correlation of this kinase with maturation-promoting factor and chromosome condensation was discussed.