Effect of Adenine Nucleotides on Levels of Glycolytic Intermediates during the Development of Induced Respiration in Carrot Root Slices

Incubation of freshly cut carrot tissue in Na(3)ADP and Na(3)ATP promotes a marked intracellular increase in both ADP and ATP. The rapid increase in ATP during an ADP incubation and in ADP during an ATP incubation results from the activity of cytoplasmic enzyme systems upon the nucleotide absorbed into the cell from the incubation medium. There is a crossover at the pyruvate kinase reaction, but not at phosphofructokinase, when either ADP or ATP is added to freshly cut tissue. In tissue slices washed in distilled water, pyruvate kinase exhibits a negative crossover in the first 2 hours and a positive crossover between 2 and 10 hours after cutting. Cutting induces large changes in levels of nucleotides and glycolytic intermediates. There is an immediate depletion of these compounds upon cutting, so that nucleotides are added to a system where respiration rate is limited by endogenous nucleotide level.Variation in respiratory values for fresh cut tissue can be explained in terms of a range of endogenous ADP levels in different tissue batches. Nucleotide incubation experiments are discussed in relation to the provision of ADP to rate-limiting pyruvate kinase during the first phase in development of the induced respiration.     

Effects of Cyanide on Respiration, Growth, and Potassium Absorption by Carrot Root Tissue 4

Slowly and rapidly growing carrot root tissue cultures havebeen found to be similarly sensitive to inhibition by cyanideat 10^-4 M in respect of respiration, growth, and potassium absorption.These observations conflict with those of Steward and Millar(1954), who reported that the actively proliferating cultureswere strikingly insensitive to the inhibitor at this level.It is suggested that the failure of these authors to detectany effects of cyanide on their cultures was due to the dilutionof the inhibitor by evaporation.     

Ethylene-induced Isocoumarin Formation in Carrot Root Tissue

The concentrations of 3-methyl-6-methoxy-8-hydroxy-3,4-dihydroisocoumarin (MMHD) formed in carrot roots inoculated with certain fungi or treated with indole-3-acetic acid, 2,4-dichlorophenoxyacetic acid, or 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), were related to the amount of ethylene produced by the root tissue. Ethylene applied exogenously in concentrations above 0.3 ppm induced the formation of MMHD in carrot root discs. Continued production of MMHD required the continued presence of ethylene. The amounts of MMHD in the discs were reduced by CO₂, an inhibitor of ethylene action, and by reduction of the partial pressure of ethylene in fungus-inoculated or 2,4,5-T-treated carrot root discs. The results indicate that ethylene is required for the induction of MMHD formation by carrot root tissue.     

Effect of Adenine Nucleotides on the Respiration of Carrot Root Slices

Sodium pyruvate and dinitrophenol stimulated O(2) uptake of freshly cut phloem parenchyma from carrot roots by 63 and 120% at optimal concentrations, indicating that production of pyruvate by glycolysis regulates over-all respiratory rate. Adding 0.5 to 6.7 mm Na(3)ADP and Na(3)ATP to slices rapidly stimulates respiration rate by 20 to 85%. The effect is greater at the lower end of this concentration range and is not due to change in pH or active cation uptake. It is suggested that treating tissue with both nucleotides stimulates pyruvate kinase, the rate-limiting step in respiration of freshly cut slices, by increasing the concentration of endogenous ADP. Adenosine diphosphate continued to stimulate O(2) uptake until the peak of induced respiration, but ATP inhibited respiration during development and decline of this peak. Absence of respiratory stimulation by NaH(2)PO(4) and of respiratory inhibition by added nucleosides confirms that inorganic phosphate is not a limiting factor of respiration in freshly cut slices. The stimulation of respiration rate of these slices by dinitrophenol is consistent with results from experiments in which ADP and ATP were applied to the tissue.     

Derepression and Repression of Lysine-sensitive Aspartokinase during in Vitro Culture of Carrot Root Tissue

The increment of lysine-sensitive aspartokinase (EC 2.7.2.4) activity during in vitro culture of carrot (Daucus carota, cv. Oogata sanzun) root tissue was explained in terms of derepression caused by an earlier decrease in the endogenous level of lysine, a possible end product repressor. Tissue content of free lysine decreased to about one-third of the initial level after 1 day of culture and no lysine was detected in the 2nd day. Inclusion of lysine (0.1 to 1.0 millimolar) in the culture medium resulted in a specific suppression of increase in lysine-sensitive aspartokinase activity without affecting the increase in threonine-sensitive aspartokinase activity.     

Soluble Sugars, Respiration, and Energy Charge during Aging of Excised Maize Root Tips

Oxygen uptake and energy charge were monitored during aging of excised maize root tips and related to the soluble sugar content and exogenous sugar supply.     

Absorption of Potassium and Sodium Ions by Carrot Root Tissue Cultures

The absorption of potassium and sodium ions by cultured explantsof young secondary phloem of carrot has been examined. Muchsmaller differences were observed between slowly growing andrapidly growing tissues than in the work of Steward and Millar(1954). This discrepancy is attributed to the use by Stewardand Millar of media of dissimilar mineral salts compositionto grow the two types of tissue. At first, potassium uptake per unit of fresh weight proceededmore rapidly in the actively proliferating cultures than theslowly growing ones, but these ultimately had the greater capacityfor potassium absorption. The rapidly growing cultures showeda higher selectivity towards potassium vis-à-vis sodiumions than did those growing slowly. The effect was evident throughouta culture period of 5 weeks, but was most marked during theearly stages of growth. Some possible explanations of theseobservations are discussed.     

Aging and the Development of Enhanced Respiration in Potato Tuber Tissue

Disks of potato tissue were aged by two separate methods. The resulting enhaiicetl respirations differed both qualitatively and quantitativety. It is suggested that the diverse results previously reported indicating changes in the terminal oxidative steps and/or in the relative contributions by competing metabolic pathways to the total respiration reflect variations in the methods employed to age the tissues. The nature of the stimulated respiration is discussed in terms of removal or retention of metabolites within the tissue during aging.     

Effects of Chloramphenicol on the Uptake and Incorporation of Amino-acids by Carrot Root Tissue

The uptake of uniformly labelled L-glutamic acid-¹⁴C, glycine-¹⁴C,and L-proline-¹⁴C by carrot root tissue is inhibited by chloramphenicolat a concentration of 2 g./l. L-glutamic acid absorption isaffected to a much greater extent than is the uptake of theother two amino-acids. It is shown that the ¹⁴C from the labelledamino-acids is incorporated into protein of the carrot slices,and that the amount of incorporation from glycine and L-prolineis about six times as large as that from L-glutamic acid undercomparable conditions. Almost all the ¹⁴C detected in the proteinafter feeding labelled L-proline or glycine remains in the amino-acidsupplied; in the case of L-glutamic acid some of the label istransferred to other amino-acid residues, but most of it remainsin glutamic acid. Chloramphenicol is found to inhibit ¹⁴C-incorporationfrom L-proline and glycine in short-term, but not in long-term,experiments whereas incorporation from L-glutamic acid is inhibitedin both long- and short-term experiments. Chloramphenicol alsoinhibits incorporation of ¹⁴C from L-glutamic acid into proteinsof preparations of isolated subcellular particles from carrotroot tissue. Under some conditions, ¹⁴C incorporation into proteinproceeds for some time after the slices have been removed fromthe radioactive solution. Such incorporation is not inhibitedsignificantly by chloramphenicol when ¹⁴C-labelled L-prolineis supplied, but it is reduced by about 50 per cent. when glycineand L-glutamic acid are used. It is shown that chloramphenicolinhibits net protein synthesis in carrot root slices suspendedin aerated solutions. It is concluded that amino-acid incorporationinto the protein of carrot slices proceeds by at least two mechanisms,one of which is related to protein synthesis and is sensitiveto chloramphenicol and the other is unrelated to protein synthesisand insensitive to the drug. Most of the L-proline and glycineincorporation into carrot root tissue protein seems to occurby the chloramphenicol-insensitive mechanism while much of theL-glutamic acid-¹⁴C incorporation seems to take place by thechloramphenicol-sensitive one.     

The Influence of External Concentration on Sodium Ion Absorption by Carrot Root Tissue 4

The influence of external concentration on sodium ion absorptionby slowly and rapidly growing carrot root tissue cultures hasbeen examined. In conflict with the observations of Stewardand Millar (1954) it is observed that absorption by the twokinds of cultures is affected similarly by external concentration.The discrepancy between the two investigations is discussedand an alternative interpretation of the results of Stewardand Millar is proposed.     

Endogenous Elicitor Activity from Damaged Carrot Root Tissue and Induced Resistance to Botrytis cinerea

A peptide-containing fraction (mol. wt. c 5000 Da) extractedfrom carrot root tissues damaged by slicing or freeze-thawing,induced active defence mechanisms in carrot slices against Botrytiscinerea. Endogenous elicitor activity was present in homogenatesof fresh and freeze-thawed tissues and those treated with germinationfluid, but was absent in autoclaved tissue. The detection ofthe elicitor in homogenates within 2h of treatment, suggeststhat it is released or activated during the early stages ofcell damage. Botrytis cinerea, carrot, induced resistance, endogenous elicitor     

Activation of Protein Synthesis and Biogenesis of Mitochondrial Particles during Aging of Discs of Sweet Potato Root Tissue

Protein synthesis in mitochondrial and supernatant fractions of sweet potato root tissue was found to be activated up to about 3 and 1.5 times in response to wounding, respectively. The activation seemed to take place within 8 hr after slicing. Sucrose density gradient centrifugation of mitochondrial fraction prepared from the aged sliced tissue showed the presence of new fraction of mitochondrial particles which were not seen in the case of fresh tissue. The particles in the fraction were labeled by radioactive leucine in vivo more rapidly than the other particles. Chloramphenicol treatment of tissue before aging blocked the development of these particles. These results suggest that the particles were newly formed during aging.     

Suppression by Exogenous Phospholipid of Cyanide-Insensitive Respiration of Submitochondrial Particles from Sweet Potato Root Tissue

Submitochondrial particles from sweet potato root tissue retainedthe respiratory characteristics of the intact mitochondria withrespect to the sensitivity to cyanide and salicylhydroxamicacid. The activities of total, cyanide-insensitive, and salicylhydroxamate-sensitiverespiration of the submitochondrial particles yielded from adefinite weight of tissue slices incubated under aerobic conditions,particularly in ethylenecontaining air, were higher than thosefrom the same weight of intact tissue. The less phospholipidthe submitochondrial particles contained relative to protein,the higher the activities of cyanide-insensitive and salicylhydroxamate-sensitiverespiration tended to be relative to total respiratory activity.When the submitochondrial particles were incubated with phospholipidliposomes, the activities of cyanide-insensitive and salicylhydroxamate-sensitive,but not cyanide-sensitive, respiration became extremely low.All phospholipids showed this effect. Such incubation of thesubmitochondrial particles with phospholipid liposomes yieldedlighter particles, indicating close association of exogenouslyadded phospholipid with the particles. Phospholipid moleculesseemed to enter the membrane of the particles. We propose thatphospholipid deficiency in the mitochondrial inner membranefacilitates operation of the cyanide-insensitive electron transportpath. (Received March 30, 1984; Accepted June 15, 1984)     

Glycolytic Enzymes in Adipose Tissue of Adult Diabetics

The maximal activities of hexokinase and phosphofructokinase were measured in human adipose tissue. Though the mean activity of hexokinase was similar between a group of adult diabetics (n=24) and a group of non-diabetic controls (n=25), in the diabetic group the activity of phosphofructokinase was definitely reduced. Possibly this enzyme deficiency might contribute to impaired glucose utilization by the adipose cell in diabetes.     

The Influence of Internal Ion Concentration on Potassium Accumulation and Salt Respiration of Red Beet Root Tissue

It is shown that during a period of washing in serated distilledwater disks of Red Beet root tissue acquire the capacity toabsorb potassium rapidly. Subsequently the rate of accumulationof this ion is closely related to the internal salt-contentof the material. On the other hand, whilst the level of saltrespiration increases during the preliminary washing, it isnot influenced significantly by the internal potassium concentration.The implications of these observations are discussed in relationto a possible mechanism of mineral salt absorption.     

芍药组织培养中污染现象的克服

针对芍药在组织培养过程中外植体污染难于控制的问题进行了研究。试验分别于不同的季节取材后对外植体进行不同的表面消毒处理。结果表明：以芍药带芽茎段、顶芽、侧芽为外植体,采用0.1%~0.2% HgCl2消毒8 min,结合母株黄化处理、4℃消毒液+搅拌的表面消毒处理,可降低外植体初期培养污染率;在培养基中添加抗菌素,可明显控制内生菌的污染。