Bradykinin-induced changes in inositol trisphosphate mass in MDCK cells

Bradykinin produces increases in cytosolic calcium in MDCK cells. We have extracted and separated Inositol 1,4,5 trisphosphate by HPLC and after-acid hydrolysis and conversion to the hexatrifluoro-acetyl derivative quantitated by negative ion chemical ionization mass spectrometry the mass of inositol trisphosphate in MDCK cells. Bradykinin causes an increase in the mass of Inositol trisphosphate from basal levels of 152 pmoles/mg cell protein to 537 pmoles/mg cell protein by 10 secs of stimulation. We conclude that bradykinin stimulates PLC hydrolysis of PIP2 with rapid release of IP3 in sufficient amount to account for the increase in cytosolic Ca++.     

钙对不同成熟期番茄果实的PG活性及其合成的影响

本文研究了钙处理不同成熟期番茄果实对果壁组织中钙含量与转化、多聚半乳糖醛酸酶(PG)活性与 PG 合成的影响。结果表明,钙处理绿熟期的番茄果实可使总钙和可溶性钙含量明显增加,并较多转化为结合钙;后期处理,进入和转化的钙都减少。同样,钙处理愈早,对果实 PG 活性的抑制愈强,绿熟期处理可完全抑制 PG 活性。凝胶电泳结合钌红染色,证明绿熟期果实无 PG,PG 是在果实成熟过程中新合成的。钙处理愈早,对 PG 合成的抑制愈强,绿熟期钙处理可完全抑制 PG 合成。     

PRO－LONG涂膜处理对后熟香蕉果实乙烯形成以及其它有关生理变化的影响

香蕉（ＭｕｓａａｃｕｍｉｎａｔａＣｏｌｌａｃｖ．ＤｗａｒｆＣａｖｅｎｄｉｓｈ）果实采后以商业上推荐使用的１．５％Ｐｒｏ－ｌｏｎｇ溶液处理,贮藏于２０℃和７５％相对湿度下,分别测定果实的ＡＣＣ含量、ＭＡＣＣ含量、ＥＦＥ酶活性、乙烯释放、叶绿素含量的变化和果实的硬度变化．结果表明,ＰＲＯ－ＬＯＮＧ处理延缓了香蕉果实果皮的叶绿素降解、硬度的下降以及乙烯释放的增加．在后熟过程中,处理果实的ＡＣＣ含量发生积累．ＡＣＣ含量的高峰在乙烯释放高峰和ＥＦＥ酶活性高峰之前出现．与对照比较,处理果实的ＡＣＣ含量和ＥＦＥ酶活性的高峰延迟了５ｄ出现．在后熟过程中,以Ｐｒｏ－ｌｏｎｇ处理果肉四片,其ＥＦＥ酶活性受部分抑制（抑制率为１９．４５％至４０．５１％）．果实ＭＡＣＣ含量在贮藏起初处于一个较显著水平,随着后熟的发展而逐步增加,但与ＡＣＣ含量的明显增加相比变化是微小的．我们的研究进一步阐明了ＰＲＯ－ＬＯＮＧ涂膜对香蕉果实后熟的影响主要是通过减少氧的供给,部分地抑制了ＥＦＥ酶活性,延缓了乙烯的形成和释放,从而延长了后熟过程．     

SYNCHRONOUS CULTURE OF CHLORELLA: II. CHANGES IN CONTENT OF VARIOUS VITAMINS DURING THE COURSE OF THE ALGAL LIFE CYCLE

1. Chlorella ellipsoidea was grown synchronously and the changesin content of various vitamins during the algal life cycle werefollowed either by chemical or microbiological assay methods.
2. In terms of µg per gram of cell dry weight, the contentof some vitamins (niacin, biotin, inositol and choline) remainedalmost constant throughout the algal life cycle, while thatof others (vitamin B₆-complex, pantothenic acid, folic acid,thiamine and riboflavin) was found to decrease more or lessmarkedly during the "growing phase" and increase at later phasesof "ripening". The content of p-aminobenzoic acid increasedonly at an early stage of "ripening", and that of ascorbic acidincreased only at the stages in which photosynthesis occurredmost actively.
3. These results were discussed in an attemptto interprete theirrelationship with the previously reportedobservations pertainingto the physiological and biochemicalevents occurring in thelife cycle of the alga.

(Received November 7, 1959; )     

Two phases of inositol polyphosphate and diacylglycerol production at fertilisation

[3H]Inositol and [3H]arachidonic acid were used to label polyphosphoinositide phospholipids in sea urchin eggs. Both [3H]inositol polyphosphate (InsP3) and [3H]diacylglycerol (DAG) increase at fertilisation. An early increase in InsP3 occurs as the sperm-induced calcium transient crosses the egg and exocytosis occurs; a later increase in InsP3 as calcium declines and the protein kinase C-dependent Na/H antiporter causes the cytoplasmic pH in increase. These results support suggestions that a calcium-induced hydrolysis of phosphatidylinositol bisphosphate occurs at fertilisation, that the production of diacylglycerol may be essential for exocytosis and that diacylglycerol production at fertilisation stimulates the Na/H antiporter. The increase in [3H]inositol polyphosphate as calcium declines indicates that this second messenger may have some function later in the cell cycle.     

Lack of Involvement of Nutrients in the Latency of Alternaria alternata in Unripe Mango Fruits

The possible relation between changes in mango fruit composition during ripening and the latency of Alternaria alternata infections was investigated. The main increase in total soluble solids and the decrease in acid content in ripening mangofruits occurred during the initial 13 days after harvest, while symptoms of A. alternata appeared only 5 days later. Susceptibility of resistant peeled mango fruits flesh could not be increased by infiltration with sucrose, at a concentration that sustains maximal growth in vitro.     

Effect of the Calcium on Polygalacturonase (PG) Activity and Synthesis at Different Ripening Stages of Tomato Fruits

It has been reported that PG is a key enzyme related to the tomato fruit ripening and that the application of calcium can dramatically decrease the PG activity and delay the ripening of fruits. In this paper the effects of calcium treament at various ripening stages on the transformation of absorbed calcium, PG activity and PG synthesis in tomato fruits were studicd. According to the analysis of calcium by atomic absorption spectroscopy, it was shown that the soluble and total calcium contents in pericarp of fruits treated with calcium at mature-green stage were increased significantly, and that more soluble calcium was transformed into bound calcium. Both the absorption and transformation of calcium decreased in fruits treated with calcium at later stage of ripening. The inhibition of calcium on PG activity was most effective by treatment at mature-green stage, but less effective at later stage of ripening. One reason for the decrease of calcium inhibition was probably due to the decline of calcium absorption as fruit ripening. The polyacrylamide gel electrophoresis of PG showed that PG with a molecular weight of 46.7 kD was absent in mature-green fruits, and PG synthesis occurred only at the later stage of ripening. It seems that the earlier the treatment was done the more effective of the calcium inhibition of PG synthesis. Based on the above results, it was concluded that the PG plays a major role in ripening and senescence of tomato fruits, and both PG synthesis and its activity were inhibited by calcium. In order to delay the ripening and senescence of tomato fruits, the treatment with calcium should be done at mature-green stage.     

Composition of ethanol-insoluble polysaccharides in water extracts of ripening tomatoes

The carbohydrate composition of the 80% ethanol-insoluble polysaccharides (EIP) from water extracts of ‘Rutgers,’ rin (ripening inhibitor) and nor (non-ripening) tomatoes has been determined. The amount of EIP extracted from ‘Rutgers’ fruit increased from 0.34 to 0.61 mg/g fr. wt during ripening little change occurred in rin or nor fruit. The carbohydrate composition (μg/g fr. wt) of EIP from mature green fruit was: galacturonic acid (48); rhamnose (3); arabinose (20); xylose (48); mannose (31); glucose (139); galactose (51). The most obvious changes that accompanied ripening were a 7.4-fold and 4-fold increase in galacturonic acid and rhamnose content, respectively. These changes were attenuated in the ripening mutants. EIP was fractionated into three major peaks by using DEAE-cellulose ion exchange chromatography. The first peak, which was not retained by the column, contained predominantly glucose and mannose, with lower amounts of galacturonic acid and galactose. The two retained peaks which eluted at 0.1 and 0.2 M sodium chloride contained primarily galacturonic acid, xylose, galactose and arabinose. The galacturonic acid content of these two fractions increased substantially during ripening, whereas the other components decreased. No changes were evident in the ripening mutants. No increase in water-soluble polysaccharides high in galactose content was observed during ripening.     

Maturation and Ripening of Fruit ofAmelanchier alnifoliaNutt. are Accompanied by Increasing Oxidative Stress

The extent of oxidative stress during ripening of saskatoon(AmelanchieralnifoliaNutt.) fruit was examined. Lipid peroxidation duringfruit development from the mature green to the fully ripe (purple)stage was evidenced by the accumulation of ethane and 2-thiobarbituricacid reactive substances. Fruit polar lipid and free fatty acidconcentrations also declined during ripening. Moreover, thedouble bond index of fatty acids in the polar lipid fractionfell during ripening, reflecting a progressive increase in thesaturation of membrane lipids. This increase in saturation waspartly due to a 65% decline in the concentration of linolenicacid. Activities of superoxide dismutase (SOD) and catalase(CAT) fell about 4-fold and 18-fold, respectively, during development,indicating higher potential for the accumulation of cytotoxicH₂O₂. Peroxidase activity remained relatively low and constantfrom the mature green to the dark red stage of development,then increased towards the end of ripening as fruits turnedpurple. Lipoxygenase (LOX) activity increased 2.5-fold fromthe mature green to the fully ripe stage. Tissue prints showedLOX to be present throughout fruit development and Western analysisrevealed that the increase in activity during ripening was dueto increased synthesis of the enzyme. Collectively, these resultsprovide evidence that ripening of this climacteric fruit isaccompanied by a substantial increase in free-radical-mediatedperoxidation of membrane lipids, probably as a direct consequenceof a progressive decline in the enzymatic systems responsiblefor catabolism of active oxygen species. The role of glutathione-mediatedfree-radical scavenging was also examined as a potential systemfor coping with this increased oxidative stress. Concentrationsof reduced and oxidized glutathione (GSSG) increased 2-foldand GSSG increased as a percentage of total glutathione, reflectingthe increase in oxidative status of fruits during ripening.Tissue prints of glutathione reductase (GRase) and transferase(GTase) showed these enzymes to be distributed throughout thepericarp at all stages of fruit development. GRase and GTaseactivities rose sharply during the later stages of fruit ripening,correlating well with substantial increases in the levels ofboth enzymes. Hence, the glutathione-mediated free-radical scavengingsystem was up-regulated towards the end of ripening, perhapsin response to the increasing oxidative stress resulting fromthe accumulation of lipid hydroperoxides from increased LOXactivity, in conjunction with a decline in SOD/CAT activities.Copyright1998 Annals of Botany Company Amelanchier alnifoliaNutt.; saskatoon fruit; ripening; oxidative stress.     

Phenological resistance of grapes to the green June beetle,an obligate fruit-eating scarab

Changes in fruit characteristics associated with ripening increase the vulnerability of crops to insect depredation, making it difficult for growers to protect cultivated fruits from pest injury close to harvest. This study evaluated phenological resistance, the use of cultivars that ripen before or after peak pest activity, for reducing injury to grapes (Vitis spp.) by the green June beetle (GJB) (Cotinis nitida), an obligate feeder on soft, ripe fruits. Accumulation of sugars, softening of berry skins and recruitment of GJB feeding aggregations were monitored on replicated vines of early-, mid- and late-season ripening cultivars that require from 85 to 125 growing days from bloom to harvest. GJB flight peaked in late July and early August coinciding with later stages of veraison of early-season ripening cultivars which recruited numerous GJB feeding aggregations resulting in >95% crop loss. Small (1–2 weeks) phenological differences between mid-season ripening cultivars and peak GJB flight translated to marked differences in injury, whereas cultivars that ripened in mid-August or later, after GJB flight had waned, sustained little or no damage. Trapping experiments confirmed that the tougher berries and low sugar content of less-ripe fruit clusters inhibited beetle feeding and induction of yeast-mediated volatiles responsible for GJB host-location. Implications of these findings for sustainable or organic management of GJB and other near-harvest fruit pests are discussed.     

Analysis of the Expression of Anthocyanin Pathway Genes in Developing Vitis vinifera L. cv Shiraz Grape Berries and the Implications for Pathway Regulation

Anthocyanin synthesis in Vitis vinifera L. cv Shiraz grape berries began 10 weeks postflowering and continued throughout berry ripening. Expression of seven genes of the anthocyanin biosynthetic pathway (phenylalanine ammonia lyase [PAL], chalcone synthase [CHS], chalcone isomerase [CHI], flavanone-3-hydroxylase [F3H], dihydroflavonol 4-reductase [DFR], leucoanthocyanidin dioxygen-ase [LDOX], and UDP glucose-flavonoid 3-o-glucosyl transferase [UFGT]) was determined. In flowers and grape berry skins, expression of all of the genes, except UFGT, was detected up to 4 weeks postflowering, followed by a reduction in this expression 6 to 8 weeks postflowering. Expression of CHS, CHI, F3H, DFR, LDOX, and UFGT then increased 10 weeks postflowering, coinciding with the onset of anthocyanin synthesis. In grape berry flesh, no PAL or UFGT expression was detected at any stage of development, but CHS, CHI, F3H, DFR, and LDOX were expressed up to 4 weeks postflowering. These results indicate that the onset of anthocyanin synthesis in ripening grape berry skins coincides with a coordinated increase in expression of a number of genes in the anthocyanin biosynthetic pathway, suggesting the involvement of regulatory genes. UFGT is regulated independently of the other genes, suggesting that in grapes the major control point in this pathway is later than that observed in maize, petunia, and snapdragon.     

Phosphorus metabolism during ripening ofGlycine max. (L.)Merril

The seeds, seed covers and leaves, taken after 17, 27, 37, and 47 days after tagging of flowers of soybean, were analysed quantitatively for their contents of different phosphorus fractions. Total phosphorus content increased in seed cover and leaves, there was a gradual decrease during ripening. All the phosphorus fractions i.e. acid soluble—P, lipid—P, nucleic acid—P, and protein—P were found to increase with maturity in seeds whereas in case of seed covers the content of acid soluble—P, nucleic acid—P and protein—P decreased but a marked increase was observed in lipid—P. In leaves during ripening, all the phosphorus fractions decreased except protein—P which was found to be almost constant. In lipid—P an increase was observed during later stages of maturity.     

The regulation of the phosphorylation of inositol 1,3,4-trisphosphate in cell-free preparations and its relevance to the formation of inositol 1,3,4,6-tetrakisphosphate in agonist-stimulated rat parotid acinar cells

High performance liquid chromatography analysis of supernatants from acid-quenched [3H]inositol-labeled parotid acinar cells revealed an inositol pentakisphosphate and three inositol tetrakisphosphates. Two of the latter were identified as the 1,3,4,5 and 1,3,4,6 isomers, whereas the third was probably a mixture of unknown proportions of the 3,4,5,6/1,4,5,6 enantiomeric pair. Methacholine (100 microM) produced a 40-50-fold increase in the levels of inositol trisphosphate (mainly the 1,3,4 isomer) and inositol 1,3,4,5-tetrakisphosphate, but inositol 1,3,4,6-tetrakisphosphate only increased 5-fold. Levels of inositol 3,4,5,6/1,4,5,6-tetrakisphosphate and inositol pentakisphosphate were unaffected by agonist stimulation. Thus, in parotid cells, an agonist-induced increase in both inositol trisphosphate and inositol 1,3,4,6-tetrakisphosphate formation does not result in an increase in the rate of formation of inositol pentakisphosphate. Following the addition of 100 microM atropine to methacholine-stimulated parotid cells, the levels of [3H]inositol 1,3,4,5-tetrakisphosphate fell rapidly, returning to basal levels within 5 min. Inositol trisphosphate was metabolized more slowly and was still elevated 20-fold above basal 5 min after the addition of atropine. Inositol 1,3,4,6-tetrakisphosphate was metabolized much more slowly (t1/2 approximately 15 min). Inositol 1,3,4-trisphosphate metabolism was examined in parotid homogenates as well as in 100,000 x g cytosolic and particulate fractions. Inositol 1,3,4-trisphosphate was both dephosphorylated and phosphorylated. Two inositol tetrakisphosphate products were formed, namely the 1,3,4,6 and 1,3,4,5 isomers. Over 90% of both kinase and phosphatase activities were found in the cytosolic fractions. The ratio of activities of kinase to phosphatase decreased as the levels of inositol 1,3,4-trisphosphate substrate were increased from 1 nM to 10 microM. These data led to the conclusion that the kinetic parameters of the inositol 1,3,4-trisphosphate kinases and phosphatases are such that in stimulated cells, dephosphorylation of inositol 1,3,4-trisphosphate is greatly favored. Inositol 1,3,4-trisphosphate kinase activity was potently inhibited by inositol 3,4,5,6-tetrakisphosphate (IC50 = 0.1-0.2 microM), which leads us to propose that inositol 3,4,5,6-tetrakisphosphate is an endogenous inhibitor of the kinase.     

几个气候区木本植物的开花结果物候

分析了我国海南和广东、秦岭、东北等不同森林气候区木本植物开花、结果物候以及果实和种子大小分布的规律。三个区系的开花、结果物候和种子、果实大小分布,都有类似的格局。但随着纬度的升高,一年中植物开花和结果的时间更加集中,海南和广东整年都有木本植物开花,秦岭有10个月左右,而东北仅有7个月。并且随着纬度升高,开花高峰的时间较迟,而结果高峰的时间较早。在海南和广东,热带区系成分和温带区系成分的木本植物,一年中开花和结果物候格局是很一致的。三个区系木本植物的果实和种子大小分布的格局也是很相似的,但海南和广东植物果实和种子大小范围较大,较多样,随着纬度升高,果实和种子大小范围变小,较单调。三个区系木本植物最小的果实的大小都差不多,为0．1cm,但最小的种子的大小却很不相同,随着纬度的升高而增大,开花和结果物候与月均气温及降水量的相关性因不同的区系而不同．鼎湖山常见木本植物果熟期和气候因子的相关性比结果期更显著。     

Increase in cytoskeletal actin induced by inositol 1,4-bisphosphate in saponin-permeated pig platelets

Inositol 1,4-bisphosphate (IP₂), which rapidly accumulates during cell activation, strongly stimulates an increase in cytoskeletal actin in saponin-permeated platelets, and the effect is insensitive to 5′-Chloro-5′-deoxyadenosine. Within 10 s, the amount of cytoskeletal actin in platelets rapidly increases by 41%, and then slowly increases further. IP₂ induces the increase in cytoskeletal actin in a dose-dependent manner. The half-maximal effect requires approximately 2 μM of IP₂ Inositol 1,4,5- triphosphate, the messenger for Ca²⁺ release, causes the increase in cytoskeletal actin, but is less effective than IP₂. Inositol 1-monophosphate and inositol 2-monophosphate have no effect on cytoskeletal actin. Phorbol 12-myristate 13-acetate, which has been shown to activate IP₃ 5′-phosphatase through protein kinase C, stimulates the increase in cytoskeletal actin. Spermine, an inhibitor of IP₃ 5′-phosphatase, inhibits the thrombin stimulated increase in cytoskeletal actin. These results suggest that IP₂ may be a messenger that controls the organization of actin filaments during cell activation. This study presents the first evidence for IP₂ as a messenger during cell activation.     

SELECTION OF ATTRIBUTES FOR THE SENSORY EVALUATION OF ANCHOVIES DURING THE RIPENING PROCESS

Anchovy semipreserves are elaborated in Mediterranean countries with ripened Engraulis encrasicolus obtained by a traditional process. In order to identify those sensory attributes that are most useful as indicators of the ripening stage of Engraulis encrasicolus we performed sensory analyses of anchovy samples obtained at intervals throughout the ripening process. The resulting 90 × 44 sample-by-attribute matrix was analyzed by principal components analysis, leading to the selection of 11 attributes that are reliable indicators of ripening stage and spoilage. The attributes chosen were gray color of the external side; red color of the external side, internal side and backbone mark; the odors Iberian ham and fish meal; the flavors anchovy and bitter; and the texture attributes firmness and juiciness.