Reactivation of the Hill reaction of Tris-washed chloroplasts

The Hill reaction of chloroplasts was inhibited by washing themwith 0.8 M Tris buffer. This inhibition was further promotedby adding ferricyanide in the washing medium. When a reducingreagent, such as the 2,6-dichlorophenol indophenol (DCPIP)-ascorbatesystem or the hydroquinone (HQJ-ascorbate system, had been addedto the Tris buffer, Hill reaction activity was unaffected. Hill reaction activity of Tris-washed chloroplasts recoveredup to 70% of the initial level by re-washing the chloroplastswith a preparation medium containing theabove reducing reagents. Photobleaching of carotenoid and chlorophyll is characteristicof Tris-washed chloroplasts. However, reactivated chloroplastsshowed no photobleaching as in the case with intact chloroplasts. (Received April 20, 1970; )     

Function of the <Emphasis Type="Italic">rel</Emphasis> Gene in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Sokawa et al. suggest that rel^- strains of Escherichia coli possess abnormal protein synthesizing machinery, which cannot carry out normal protein synthesis when the supply of amino-acids is limited.     

Ommochrome Deficiency in an Albino Strain of a Terrestrial Isopod,Armadillidium vulgare

In order to clarify the cause of ommochrome deficiency in an albino strain of the terrestrial isopod, Armadillidium vulgare, levels of xanthom-matin, 3-hydroxykynurenine, 3-hydroxyanthranilic acid and tryptophan in whole body extracts of the albino and the wild type individuals were determined together with enzyme activities of kynurenine-3-hydroxylase, kynureninase and tryptophan-2,3-dioxygenase. Xanthommatin could not be detected in the albinos. The levels of 3-hydroxykynurenine and 3-hydroxyanthranilic acid were determined by high-performance liquid chro-matography (HPLC) with electrochemical detection and were markedly low in the albinos compared with the wild type individuals. In contrast to those, the tryptophan levels determined by HPLC with fluorescence detection did not differ significantly between the two phenotypes. In the albino A. vulgare, kynurenine-3-hydroxylase activity was lower and kynureninase activity was higher than in the wild type, although the differences were not statistically significant. Tryptophan-2,3-dioxygenase activity in the albinos was less than 10% that in the wild type. Thus, ommochrome deficiency in the albino A. vulgare is considered to be caused by the extremely low activity of tryptophan-2,3-dioxygenase.     

Permeability of the Nitella internodal cell to organic substances as measured by the double chamber method

Permeability coefficients for the cell membrane of Nitella jlexilishave been determined for some organic substances by measuringtheir transcellular transport with the double chamber method.Benzyl alcohol is rapidly taken up in the vacuole of the cell.In the double chamber method benzyl alcohol taken up in thevacuole at one end of the cell is not as rapidly transportedto the other end as expected from the rate of protoplasmic streamingin the vacuole. This can be explained by assuming the presenceof a "non-streaming part" in the vacuole. The uptake of ureainto the streaming part of the cell is stimulated by directlyilluminating the part where the cell is immersed in urea solution. (Received March 10, 1971; )     

Freeze-Fracture Study of the Bloodstream Form of Trypanosoma brucei gambiense

The ultrastructure of Trypanosoma brucei gambiense was investigated by the freeze-fracture method. Three different regions of the continuous plasma membrane; cell body proper, flagellar pocket, and flagellum were compared in density and distribution of the intramembranous particles (IMP's). The IMP-density was highest in the flagellar pocket membrane and lowest in flagellum. Intra membranous particles of the cell body membrane were distributed uniformly on both the protoplasmic (P) and exoplasmic (E) faces. On the P face of the flagellar membrane, a single row of IMP-clusters was seen along the juncture of the flagllum to the cell body. Since the spacing of the IMP-clusters was almost equal to the spacing of the paired rivet structures observed in thin section, these clusters likely are related to the junction of flagellum and cell body. At the neck of the flagellar pocket, several linear arrays of IMP's were found on the P face of the flagellar membrane, while on the E face rows of depressions were seen. At the flagellar base, the clusters of IMP's were only seen on the P face. On the flagellar pocket membrane, particle-rich depressions and linear particle arrays were also found on the P face, while on the E face such special particle arrangements were not recognized. These particle-rich depressions may correspond to the sites of pinocytosis of the bloodstream forms which have been demonstrated in thin sections.     

DISTRIBUTION OF MATURATION-PROMOTING FACTOR IN STARFISH OOCYTE STRATIFIED BY CENTRIFUGATION

In starfish, cytoplasm taken from maturing oocytes is capable of inducing breakdown of the germinal vesicle and subsequent maturation when injected into immature oocytes. The cytoplasmic factor has been designated as "maturation-promoting factor" (MPF). Ooplasm was stratified by centrifugation of maturing oocytes in density-graded Ficoll-seawater, without disrupting the cell. Three strata were distinguished beginning with the centripetal side: oil cap, hyaline layer and yellow layer. MPF activity was shown to be localized in the hyaline layer. Electron microscopic observation revealed that the hyaline layer contains Golgi complexes, many lucent vesicles and multi-vesicular bodies as distinct organelles, but seldom contains such inclusions as the lipid droplets forming the oil cap, mitochondria, yolk and pigment granules contained in the yellow layer. Based on these observations, a possible cytoplasmic component with MPF activity is discussed.     

BINDING OF CONCANAVALIN A TO THE SURFACE OF STARFISH FOLLICLE CELLS AND PRODUCTION OF 1-METHYLADENINE

Starfish follicle cells, treated with concanavalin A (Con A), continued to produce 1-methyl-adenine (1-MeAde), an inducer of starfish oocyte maturation, after rinsing with artificial seawater (ASW). On the other hand, they ceased to produce the substance if treated with methyl α-Dmannoside (αMM). These cells produced again 1-MeAde when re-stimulated with Con A after removal of αMM. An optical study with fluorescein revealed that Con A bound to the cells was not dissociated by rinsing with ASW, but was removed if the cells were treated with αMM. These results suggest that continuous binding of Con A to the surface of the follicle cells is essential for the production of 1-MeAde.     

PRODUCTION OF 1-METHYLADENINE INDUCED BY CONCANAVALIN A IN STARFISH FOLLICLE CELLS

Concanavalin A (Con A) was found to induce maturation of oocytes with follicular envelopes in the starfish, Asterina pectinifera . Treating a Con A sample with 85% ethanol and heat revealed that the maturation-inducing activity of the sample was not due to possible contamination with 1-methyladenine, but to Con A itself. However, Con A had little maturation inducing effect on isolated oocytes from which the follicular envelope had been removed, suggesting that its effect is indirect and probably mediated by the follicle cells. When follicle cells were incubated in seawater containing Con A, a maturation-inducing substance was found to have been produced in the incubation medium. This was purified and identified as 1-methyladenine. Therefore it is concluded that Con A has the same capacity as GSS, a gonad-stimulating peptide hormone of neural origin, to induce production of the maturation-inducing substance. Other plant lectins such as phytohemagglutinin P and wheat germ agglutinin had little effect in inducing production of 1-methyladenine in follicle cells.