Acid Phosphatase in Cotyledons of Germinating Vigna mungo Seeds

A marked increase in acid phosphatase activity took place incotyledons of germinating Vigna mungo seeds. The attachmentof axis organs was not required for this development of enzymeactivity in cotyledons. DEAE-cellulose column chromatographyrevealed that the phosphatase is composed of at least threeforms. (Received August 19, 1981; Accepted October 30, 1981)     

Multiple Forms of Acid Phosphatase in Cotyledons of Vigna mungo Seedlings

Acid phosphatase from cotyledons of dark-grown Vigna mungo seedlingswas separated into four forms (la, Ib, Ha and lib) by ion-exchangercolumn chromatographies. Each form of the acid phosphatase wascharacterized for its pH dependency, substrate specificity,thermal stability, activation energy, approximate molecularweight, and the effect of metal ions and other substances onits activity. Each form of the enzyme exhibited high activitytowards ATP and ADP relative to their activity towards p-nitrophenylphosphate,but showed very low activity towards phytate, a major organicphosphate reserve in cotyledons. Among the four forms, lib wasmost distinguishable by its low molecular weight and the thermalenhancement of activity.     

Protein Body Formation in Cotyledons of Developing Cowpea (Vigna unguiculata) Seeds

The sequence of protein body formation in cotyledonary cellsof developing cowpea seeds has been followed by electron microscopy.Some protein bodies are formed by sub-division of the main vacuolesof the cotyledon cell but most are formed by expansion of cytoplasmicvesicles which may originate from Golgi bodies. Protein depositionis accompanied by a massive proliferation of rough endoplasmicreticulum and Golgi body production.     

Organic Acid Metabolism of Sedum praealtum

1. The organic acids present are citric, isocitric, and l-malic,with a small residue of unidentified acids.
2. The diurnal variationin acidity is due chiefly to changes,in malic acid, with aparallel fluctuation shown by citric acid.Under these conditionsisocitric acid shows little change.
3. The importance of carbondioxide during acidification is confirmed,and it is shown thatat room temperatures or higher the CO₂produced in respirationis sufficient to produce maximum acidification.At lower temperaturesthe supply of CO₂ limits acid production.
4. In the absence ofoxygen no acidification occurs, but even smallquantities (approx.1 per cent.) are sufficient to cause someacid production.
5. Completebalance-sheets are presented for acids, carbohydrates,CO₂ andoxygen for leaves maintained in the dark at high andlow temperatures.As acids are produced there is a correspondingloss of carbohydrate(chiefly starch). A scheme of reactionsis suggested to explainthe experimental results.

     

Biochemical and Histochemical Studies on Protease Activity and Reserve Protein Metabolism in the Cotyledons of Germinating Cowpeas (Vigna unguiculata),

Changes in protein and protease distribution in cotyledons ofgerminating cowpea have been followed by histochemical, biochemical,and ultrastructural techniques. A close relationship, in locationand timing, between an enhanced protease activity and proteinmobilization has been demonstrated. The major enzyme(s) involvedin protein breakdown is of the endopeptidase type. Initiationof protein mobilization appears to be associated with the fusionof cytoplasmic vesicles with the protein bodies.     

Multiple Forms of Acid Phosphatase in Cotyledons of Vigna mungo Seedlings: Immunological Detection and Quantitation

Using a combination of column chromatography and gel electrophoresis,we have found that acid phosphatase in cotyledons of Vigna mungoseedlings is composed of at least six forms (Ia₁, Ia₂, Ib₁,Ib₂, IIa and IIb). We purified one of the major forms, Ia₁,as a polypeptide of 53 kDa. Using an antiserum raised againstthe enzyme Ia₁, we examined the immunological relationshipsbetween the multiple forms from cotyledons and the distributionof the enzyme in organs of maturing and germinating seeds. (Received December 25, 1989; Accepted July 11, 1990)     

Imunohistochemical Localization of alpha-Amylase in Cotyledons of Vigna mungo Seedlings

We studied the localization of α-amylase with indirect fluorescence microscopy in transversely sectioned cotyledons of Vigna mungo seedlings. Tissue sections were fixed in periodate-lysine-paraformaldehyde and treated with anti-α-amylase immunoglobulin G followed by fluorescein isothiocyanate labeled goat anti-rabbit immunoglobulin G. α-Amylase appeared in the cells farthest from vascular bundles on the second day of growth and appeared gradually closer to the vascular bundles as growth progressed. The pattern of α-amylase appearance was similar in detached cotyledons, indicating that attachment of the embryonic axis has no effect on this pattern. However, in attached cotyledons, α-amylase disappeared from the regions where starch grains had been digested, but in detached cotyledons there was no disappearance of α-amylase, and digestion was slower than in intact cotyledons.     

Localization of Sorbitol Oxidase in Vacuoles and Other Subcellular Organelles in Apple Cotyledons

To investigate the function and subcellular localization ofsorbitol oxidase, free cells, protoplasts and isolated vacuolesof apple cotyledons (Malus pumila Mill. var. domestica Schneid.)were examined by differential and sucrose density gradient centrifugation.Twenty percent of the activity of sorbitol oxidase in the wholetissue was contained in the subcellular fraction (d=1.06) whichcorresponded closely to the main peaks of activity and proteinafter the recentrifugation of the 150,000?g pellet of rupturedvacuoles with a linear sucrose density gradient. The enzymethus appears to be derived from the tonoplast membrane. Thistonoplast membrane-bound sorbitol oxidase may play an importantrole in the transport of vacuolar sorbitol into the cytoplasm,rather than in the transport of sorbitol into the vacuole. About10% of the enzyme activity also occurred in the subcellularfraction having a density of 1.12–1.16, which coincidedwith the peaks of acid phosphatase and ATPase activities. Thereforesorbitol oxidase may also be associated with the plasma membrane.Furthermore, 30–40% of its activity was located in theinterspace between the cell wall and the plasma membrane, orperhaps attached weakly to them. These results suggest thatsorbitol is transported into the cytoplasm by being convertedto glucose by sorbitol oxidase. ¹ This paper is contribution A-138 of the Fruit Tree ResearchStation. (Received January 20, 1982; Accepted May 18, 1982)     

Functional aspects of Abscisic Acid Metabolism in Cotyledons of Phaseolus vulgaris L. Seedlings

During the early stages of germination and vegetative development,cotyledons of intact bean (Phaseolus vulgaris L.) seedlingsshowed active ABA catabolism causing a low endogenous ABA content.At the end of the substrate mobilizing phase, when the cotyledonsbecame senescent, a drastic increase of the endogenous ABA contentlinked with a decrease of the ABA catabolic activity was observed.This indicates that a close connection exists between senescenceand endogenous ABA content and metabolism in bean cotyledons. Removal of the apical growth region induced in the cotyledonsactivation of the ABA catabolism and the endogenous ABA concentrationdecreased below the detection limit (0.1 ng/g fr wt) at theonset of the outgrowth of the axillary buds. From then, apicaldominance was restored and the cotyledons returned to the senescentstate, which was correlated with a drastic increase of theirendogenous ABA content. ¹ Bevoegd verklaard navorser N. F. W. O. ² Wetenschappelijk medewerker F. K. F. O. (Received November 25, 1980; Accepted February 13, 1981)     

Aminopeptidases Isolated from Cotyledons of Cowpea, Vigna unguiculata

Three aminopeptidases have been separated from cotyledon extractsfrom cowpea, Vigna unguiculata (L.) Walp., and numbered in orderof decreasing affinity for the anion exchange medium DEAE-Sephacel.API showed a wide acceptance of model substrates, with highestactivity under standard conditions against arginyl ß-naphthylamide(NA). AP2 did not act on basic substrates and preferred phenylalanylß-NA. AP3 displayed the narrowest substrate specificity,with strong activity against only alanyl ß-NA andglycyl ß-NA. The chelator 1,10-phenanthroline completelyor almost completely inhibited forms AP1 and AP3, whereas AP2was insensitive to phenanthroline at the same concentration(5 mM). All three aminopeptidases were totally inhibited byAg⁺ or Zn²⁺ ( 0.5 mM). Vigna unguiculata (L.) Walp., aminopeptidase, cotyledon, cowpea, isoenzyme, 1, 10-phenanthroline     

Nucleic Acid Metabolism During Cytokinin Induced Cellular Differentiation

Edstrom's microphoresis technique has been employed to determine the quantitative alterations in nucleic acid content and base composition of individual cells associated with the initiation of bud primordia in Funaria hygrometrica. The filamentous protonema of this moss initiates bud cells which through repeated divisions form the leafy gametophores. The cytokinin, 6-furfurylaminopurine (kinetin), was used to induce the differentiation of bud cells from protonematal cells. The total RNA content of kinetin-induced bud cells (22.0 μμg/cell) was nearly 15 times that of protonematal cells (1.6 μμg/cell). The same dramatic increase in total RNA was apparent in bud cells which developed spontaneously in older cultures. As would be predicted, the adenine (A) to guanine (G) ratio for DNA from bud and protonematal cells was identical (0.7). The A:G ratio for RNA from bud cells (1.0) was much lower than that from protonematal cells (1.7). Thus, kinetin-induced differentiation in this system involves a dramatic increase in total RNA, the base composition of which approaches that of DNA. The base composition (A:G ratio) of DNA remains constant.     

Changes in mRNA of Vigna mungo Cotyledons during Seed Germination

Quantitative and qualitative changes of mRNA in Vigna mungocotyledons during seed germination have been investigated. TotalRNA is higher in dry cotyledons and declines during germination.Poly(A)⁺ RNA also is present at a relatively high level in drycotyledons, increases slightly during the first day of germination,and then decreases. Polysomal RNA is very low in dry cotyledonsbut increases rapidly during the first day of germination, andthen declines. The translational activity of the mRNA in a wheatgerm cell-free system is low on day 0 but increases rapidlyon day 1 of germination. Two-dimensional gel electrophoresisof in vitro translation products reveals that many new peptidesare synthesized on day 1 of germination. Synthesis of most ofthese polypeptides continue throughout 5 days of germination. Change in the mRNA population during germination has been investigatedusing cDNA against poly(A)⁺ RNA from 3-day-old cotyledons. Withtotal RNA of day 3 and 5, the cDNA strongly hybridized withRNA similar in size to 25 S ribosomal RNA, but no specific bandsare detected with samples of day 0 or 1. With poly(A)⁺ RNA ofday 5 or 1, the cDNA tends to hybridize with RNAs of relativelysmall molecular size. Cordycepin and -amanitin prevent the increasein poly (A)⁺ RNA content and the appearance of new mRNAs duringthe first day of germination. ¹Present address: Division of Regulation of Macromolecular Function,Institute for Protein Research, Suita City, Osaka 565, Japan. (Received January 13, 1986; Accepted June 10, 1986)     

The Organic Acid Metabolism of Swede 'Root' Tissue

Data are recorded on the quantitative changes in the contentof succinic, malic and citric acids of discs of washed ‘root’tissue of ‘swede’ (Brassica napus), subjected toexperimental treatments involving the addition of succinate,malate, citrate, -ketoglutarate, pyruvate and malonate. Theresults are interpreted as showing the acids to be associated,in part at least, with the operation of an active Krebs cycle.It is suggested that they are also involved, by providing carbonskeleton material, in the enhanced protein synthesis resultingon washing discs of swede tissue and treating washed discs withphosphate buffer.     

Characterization of Serine Endopeptidases in Cotyledons of Germinated Vigna mungo Seeds

Vigna mungo seeds. SEP activity was separated into two isoforms by CM-cellulose column chromatography. These forms, termed SEP-1 and SEP-II, showed endopeptidase activities even at acidic pH, suggesting that SEPs are unique serine endopeptidases, since most serine proteases are optimum at neutral pH. Received 14 December 1998/ Accepted in revised form 22 February 1999     

Organic Acid Metabolism by Isolated Rhizobium japonicum Bacteroids

Rhizobium japonicum bacteroids isolated from soybean (Glycine max L.) nodules oxidized ¹⁴C-labeled succinate, pyruvate, and acetate in a manner consistent with operation of the tricarboxylic acid cycle and a partial glyoxylate cycle. Substrate carbon was incorporated into all major cellular components (cell wall + membrane, nucleic acids, and protein).