Sugar uptake into strawberry fruit is stimulated by abscisic acid and indoleacetic acid

Changes in sugar uptake into strawberry fruits with maturation and the hormonal effect on uptake mechanisms, though important to fruit development, are not known. Therefore, the kinetics of sugar uptake into strawberry ( Fragaria x ananassa Duch cv. Nyoho) fruit tissue and the effects of abscisic acid (ABA) and indoleacetic acid (LAA) on the mechanism of uptake were investigated at 25 and 35 days after pollination (DAP). Uptake of ¹⁴C-sugar was measured over the concentration range of 2 to 30 m M. Uptake kinetics showed a biphasic response to increasing external concentration of ¹⁴C-sugars, and indicated the presence of P -chlorormercuribenzenesulfonic acid (PCMBS)-sensitive and PCMBS-insensitive uptake. The K_m value for each sugar was in the range of 10 to 20 m M. Stage of development had no effect on K_m. but V_max for glucose decreased with maturation. Further, sucrose was not taken up through a PC-MBS-sensitive transport at 35 DAP. ABA, especially 10 μ M , at 25 DAP stimulated uptake of all sugars, mostly through enhanced PCMBS-insensitive uptake but not PC-MBS-sensitive uptake. In contrast to ABA, stimulation of sugar uptake by IAA was most effective at 1 μ M . The PCMBS-insensitive uptake of each sugar was also stimulated by IAA. Further, the PCMBS-sensitive uptake of glucose was enhanced. The developmental change of PCMBS-sensitive sugar uptake and the effect of ABA and IAA on uptake mechanism in this study are considered to be important in influencing the development and enlargement of fruits.     

Gastric mucosal cell homeostatic physiome. Critical role of ER-initiated membranes restitution in the fidelity of cell function renewal.

Homeostatic cell physiology is preserved through the fidelity of the cell membranes restitution. The task is accomplished through the assembly of the precisely duplicated segments of the cell membranes, and transport to the site of their function. Here we examined the mechanism that initiates and directs the restitution of the intra- and extracellular membranes of gastric mucosal cell. The homeostatic restitution of gastrointestinal epithelial cell membrane components was investigated by studying the lipidomic processes in endoplasmic reticulum (ER) and Golgi. The biomembrane lipid synthesis during the formation of transport vesicles in the systems containing isolated organelle and the cell-specific cytosol (Cyt) from rat gastric mucosal epithelial cells was assessed. The results revealed that lipids of ER transport vesicle and the transmembrane and intravesicular cargo are delivered en bloc to the point of destination. En bloc delivery of proteins, incorporated into predetermined in ER lipid environment, ensures fidelity of the membrane modification in Golgi and the restitution of the lipid and protein elements that are consistent with the organelle and the cell function. The mechanism that maintains apical membrane restitution is mediated through the synthesis of membrane segments containing ceramide (Cer). The Cer-containing membranes and protein cargo are further specialized in Golgi. The portion of the vesicles destined for apical membrane renewal contains glycosphingolipids and phosphatidylinositol 3-phosphate. The vesicles containing phosphatidylinositol 4-phosphate are directed to endosomes. Our findings revealed that the preservation of the physiological equilibrium in cell structure and function is attributed to (1) a complete membrane segment synthesis in ER, (2) its transport in the form of ER-transport vesicle to Golgi, (3) the membrane components-defined maturation of lipids and proteins in Golgi, and (4) en bloc transfer of the new segment of the membrane to the cell apical membrane or intracellular organelle.     

Non-lysosomal Degradation of Singly Phosphorylated Oligosaccharides Initiated by the Action of a Cytosolic Endo-β-N-acetylglucosaminidase

Phosphorylated oligosaccharides (POSs) are produced by the degradation of dolichol-linked oligosaccharides (DLOs) by an unclarified mechanism in mammalian cells. Although POSs are exclusively found in the cytosol, their intracellular fates remain unclear. Our findings indicate that POSs are catabolized via a non-lysosomal glycan degradation pathway that involves a cytosolic endo-β-N-acetylglucosaminidase (ENGase). Quantitative and structural analyses of POSs revealed that ablation of the ENGase results in the significant accumulation of POSs with a hexasaccharide structure composed of Manα1,2Manα1,3(Manα1,6)Manβ1,4GlcNAcβ1,4GlcNAc. In vitro ENGase assays revealed that the presence of an α1,2-linked mannose residue facilitates the hydrolysis of POSs by the ENGase. Liquid chromatography-mass spectrometric analyses and fluorescent labeling experiments show that such POSs contain one phosphate group at the reducing end. These results indicate that ENGase efficiently hydrolyzes POSs that are larger than Man₄GlcNAc₂-P, generating GlcNAc-1-P and neutral Gn1-type free oligosaccharides. These results provide insight into important aspects of the generation and degradation of POSs.     

Assignment of the S-antigen gene (SAG) to human chromosome 2q24-q37

We report the mapping of the gene coding for the S-antigen (48-kDa protein) to human chromosome 2 using somatic cell hybrids. In situ hybridization further confirms this assignment and regionally maps the gene to 2q24-q37.     

Mg++ -activated and -inhibited ATPases from mung bean hypocotyls

Mg⁺⁺-activated and inhibited ATPases were isolated from dark-grownmung bean hypocotyls. The enzymes hydrolyzed nucleoside tri-,di- and monophosphates and ß-glycerophosphate. Theeffect of Mg⁺⁺ was most marked when ATP and other nucleosidetriphosphates were used as substrates. Mg⁺⁺-activated ATPases: The activity of enzyme-I was localizedin the membranes and was not released by treatment with 0.1%deoxycholate. Enzyme-II was released and separated by CM-cellulosecolumn chromatography. Enzyme-V was separated from the solublefraction of the cell homogenate by DEAE-cellulose column chromatography.The rates of activivation by Mg⁺⁺ of enzyme-II and enzyme-Vwere very small compared to that of enzyme-I. Mg⁺⁺-inhibited ATPases: Enzyme-II and -IV were precipitatedwith 50–80% ammonium sulfate from the soluble fractionof the cell homogenate and were separated by successive columnchromatographies on Sepharose 6B and DEAE-cellulose. The activitiesof enzyme-III and -IV were inhibited by Mg⁺⁺, when ATP, UTPand GTP were used as substrates. Enzyme-III was purified approximately38-fold, and was more remarkably inhibited by Mg⁺⁺ than wasenzyme-IV. ¹Present address: Institute for Plant Virus Research, 959 Aobacho,Chiba 280, Japan. (Received January 7, 1974; )     

Effects of arildone on the immunogenicity of formalin-inactivated polioviruses

Concentrated and purified Sabin and virulent strains of poliovirus types 1, 2 and 3 were inactivated with formalin at 37 C. By addition of 5.4 microM arildone, an antiviral agent, to the virus suspension, the stability of D antigen increased in both Sabin and virulent strains of all types, especially in virulent type 1 Mahoney strain. The drug had neither any inhibitory nor enhancing effect on the formalin inactivation. When antibody response was compared in guinea pigs, Sabin strains inactivated in the absence of arildone were less immunogenic against homotypic virulent strains than inactivated vaccine prepared from virulent strains. On the other hand, Sabin strains inactivated in the presence of arildone were equally immunogenic. These results indicate that it is possible to prepare from Sabin strains a potent and safe inactivated vaccine having an immunogenicity comparable to that prepared from virulent strains.     

Mechanism of Chilling Injury in Sweet Potato: X. Change in Lipid-Protein Interaction in Mitochondria from Cold-stored Tissue

Seventy per cent of the phospholipid in mitochondria from sweet potato roots was removed by aqueous acetone treatment. The amount of phospholipid that could be rebound to these lipid-depleted mitochondria roughly corresponded to the amount of phospholipid in untreated mitochondria. The activities of NADH-cytochrome c oxidoreductase, succinate-cytochrome c oxidoreductase, cytochrome oxidase, and succinoxidase in lipid-depleted mitochondria were restored by addition of mitochondrial phospholipid to about 60, 50, 15, and 35%, respectively, in comparison to untreated mitochondria. The capacity of lipid-depleted mitochondria from 14-day cold-stored tissue to bind mitochondrial phospholipid from healthy tissue was lower than that from healthy tissue. However, there was no large difference in activities of NADH-cytochrome c oxidoreductase and succinate-cytochrome c oxidoreductase between both phospholipid rebound lipid-depleted mitochondria from healthy and 14-day cold-stored tissues. On the other hand, activity of succinoxidase in phospholipid rebound lipid-depleted mitochondria from 14-day cold-stored tissue was decreased by about 50% of that from healthy tissue. Furthermore, the capacity of lipid-depleted mitochondria from 2-day cold-stored tissue to bind mitochondrial phospholipid from healthy tissue was higher than that from healthy tissue.     

The Relationship between Growth and Proline Incorporation after Auxin Treatment of Mung Bean Hypocotyls

Indoleacetic acid (IAA) stimulates the incorporation of ¹⁴C-proline into both the cyloplasmic and the cell wall fractions of the hypocotyl of mung bean (Phaseolus aureus Roxb. cv. Black). It neither stimulates the transfer of ¹⁴C-proline from the cyloplasmic fraction into the cell wall fraction, nor the retention of ¹⁴C-proline in the wall or cytoplasmic fractions. Moreover, the stimulation of growth caused by IAA parallels the stimulation of the incorporation of proline into the cytoplasmic fraction, but does not parallel the stimulation into the cell wall fractions. The stimulation of the incorporation into the cyloplasmic fraction seems to appear within 30 minutes after auxin treatment, at about the same time the increase in the growth is observed in response to IAA, suggesting a connection between these effects. On the other hand, the stimulation of the proline incorporation into the cell wall fraction seems to require more than 90 minutes after auxin treatment, suggesting no close connection between growth and proline incorporation into the cell wall fraction.     

Intracellular processes associated with glycoprotein transport and processing.

The intracellular transport of mucus glycoprotein precursor (apomucin) from endoplasmic reticulum (ER) to Golgi was quantitated by the immunoprecipitation with 3G12 antimucin monoclonal antibody and by estimation of the apomucin glycosylation using UDP-[3H]galactose. The assembly of the entities carrying apomucin to Golgi was assessed by electron microscopy and by quantitation of the incorporation of [14C]choline, [14C]ethanolamine, and [14C]oleic acid into their lipids. The microscopic image of the isolated transport components revealed a population of 80- to 100-nm vesicles with occasional membranes of the ER used for their synthesis. On the average, the vesicles contained 82 ng apomucin/microgram of protein and 80-90% of the total incorporated lipid precursors. From that, 91% of [14C]choline was detected in phosphatidylcholine, and 9% in phosphatidylethanolamine, lysophosphatidylcholine, and sphingomyelin. With [14C]oleate, 54% of the label was incorporated into ceramide, diglyceride, and phosphatidic acid, 35% to phosphatidylcholine, 7% in phosphatidylethanolamine, and 2% in sphingomyelin. After incubation of the vesicles with Golgi, the apomucin was found glycosylated and the lipids of the transport vesicles incorporated into Golgi membranes. The fusion of the vesicular membranes was accompanied by the synthesis of sphingomyelin. In the Golgi, 39-55% of the radiolabeled phosphatidylcholine of transport vesicles was converted to sphingomyelin. The results indicate that the newly synthesized membranes of apomucin transporting vesicles are enriched in phosphoglycerides and ceramides. Upon fusion with the Golgi, the membranes of the vesicles are replenished with sphingomyelin by exchange reaction between phosphatidylcholine and ceramide.