Changes in cytokinins during initiation and development of potato tubers

Cytokinin-like activity was assayed in stolons and tubers of Solanum tuberosum L. ssp. andigena (Juz. et Buk.) Hawkes cv. 165 grown in pots under controlled environment conditions. The plants were allowed to tuberise without the application of environmental or other external stimuli. The soluble sugar and starch contents of stolon tips and tubers were measured. Starch accumulation was a precise indicator of tuber initiation. Cytokinin-like activity began to increase in tubers with a diameter greater than 7.5 mm and, as assessed on a per tuber basis, was greatest in the largest size-category analysed. However, expressed as a function of fresh and dry weight, activity was greatest in tubers of 15–20 mm in diameter. Increases in cytokinin-like activity occurred subsequent to tuber formation, indicating that the tuberisation stimulus is unlikely to be cytokinin-like in nature.     

Dormancy-related changes in cytokinin efficacy and metabolism in potato tubers during postharvest storage

The metabolism of [³H]-zeatin (Z) and[³H]-isopentenyladenosine (IPA) in potato tubers was examined inrelation to changes in cytokinin efficacy during postharvest storage anddormancy progression. Exogenous radiolabeled cytokinins were rapidlymetabolizedby dormant and nondormant tubers. Following injection, [³H]-Z wasmetabolized to zeatin riboside, adenine derivatives andzeatin-riboside-5-monophosphate. Four hours after injection, less than60% of the recovered radioactivity was associated with unmetabolized[³H]-Z. [³H]-IPA was also rapidly metabolized to severalmetabolites including: IPA-5-monophosphate, adenine derivatives andzeatin riboside. Four hours after injection, less than 50% of therecovered radioactivity was associated with [³H]-IPA. Cytokininsensitivity was assessed by determining the effects of exogenous Z or IPA ontuber sprouting. Immediately after harvest and during the initial period ofstorage, tubers were dormant and exogenous Z or IPA were completely ineffectivein breaking tuber dormancy. Thereafter, dormant tubers exhibited a gradualincrease in sensitivity to both cytokinins. Cytokinin sensitivity continued toincrease as postharvest storage was extended and dormancy weakened. The lengthof postharvest storage (hence dormancy status) had no apparent effects on themetabolism of either cytokinin. Neither the rate of metabolism nor the natureofmetabolites detected was affected by the length of postharvest storage. Theseresults suggest that changes in cytokinin efficacy in dormant potato tubersduring postharvest storage are not the result of differential catabolism butrather are due to other cellular processes such as hormone perception and/orsignal transduction.     

Structure of potato tubers formed during spaceflight

Potato (Solanum tuberosum L. cv. Norland) explants, consistingof a leaf, axillary bud, and small stem segment, were used asa model system to study the influence of spaceflight on theformation of sessile tubers from axillary buds. The explantswere flown on the space shuttle Columbia (STS-73, 20 Octoberto 5 November 1995) in the ASTROCULTURE^TM flight package, whichprovided a controlled environment for plant growth. Light andscanning electron microscopy were used to compare the preciselyordered tissues of tubers formed on Earth with those formedduring spaceflight. The structure of tubers produced duringspaceflight was similar to that of tubers produced in a controlexperiment. The size and shape of tubers, the geometry of tubertissues, and the distribution of starch grains and proteinaceouscrystals were comparable In tubers formed in both environments.The shape, surface texture, and size range of starch grainsfrom both environments were similar, but a greater percentageof smaller starch grains formed in spaceflight than on Earth.Since explant leaves must be of given developmental age beforetubers form, instructions regarding the regular shape and orderedtissue geometry of tubers may have been provided in the presenceof gravity. Regardless of when the signalling occurred, gravitywas not required to produce a tuber of typical structure. Key words: Spaceflight, development, potato tuber, microgravity     

Structure of potato tubers formed during spaceflight

Potato (Solanum tuberosum L. cv. Norland) explants, consisting of a leaf, axillary bud, and small stem segment, were used as a model system to study the influence of spaceflight on the formation of sessile tubers from axillary buds. The explants were flown on the space shuttle Columbia (STS-73, 20 October to 5 November 1995) in the ASTROCULTURE (TM) flight package, which provided a controlled environment for plant growth. Light and scanning electron microscopy were used to compare the precisely ordered tissues of tubers formed on Earth with those formed during spaceflight. The structure of tubers produced during spaceflight was similar to that of tubers produced in a control experiment. The size and shape of tubers, the geometry of tuber tissues, and the distribution of starch grains and proteinaceous crystals were comparable in tubers formed in both environments. The shape, surface texture, and size range of starch grains from both environments were similar, but a greater percentage of smaller starch grains formed in spaceflight than on Earth. Since explant leaves must be of given developmental age before tubers form, instructions regarding the regular shape and ordered tissue geometry of tubers may have been provided in the presence of gravity. Regardless of when the signalling occurred, gravity was not required to produce a tuber of typical structure.     

Factors affecting the development of spraing in potato tubers infected with potato mop-top virus

A larger proportion of tubers of Arran Pilot potato growing at the surface of soil infested with potato mop-top virus (PMTV) showed spraing symptoms (brown rings) at harvest than of tubers from below the surface. Infected tubers with or without spraing developed a spraing ring when stored in darkness, first for 1–2 wk at 18 d?C and then for 1–2 wk at any constant temperature between 5 and 13 d?C. Only a faint surface ring developed when either of these periods was decreased to 1 day; 4-day periods were needed to induce distinct symptoms. Internal tuber symptoms developed more slowly than surface symptoms, and their formation was favoured by cutting the tubers in half. Additional pigmented surface rings were produced outside the first ring by successive cycles of treatment at 18 and 9d?. Spraing did not develop when the first stage of treatment was at 22–25d?, when the tubers were kept first at 10d? and then at 5d?, when the treatment at 5–13d? preceded that at 18d?, or when the tubers were kept at constant temperatures ranging from 5 to 25d?. When tubers of six potato varieties were grown in PMTV-infested soil and then stored at temperatures designed to induce symptoms, the varieties known to be the most susceptible in the field were those which had the greatest tendency to develop spraing during storage. When infected tubers were exposed to light, typical spraing symptoms were not induced, but greening of the tuber surface was much delayed in localized ring-shaped areas, so that pale weals appeared. Spraing symptoms were produced, in favourable conditions, by the reaction of cells at the periphery of the PMTV-invaded zone. Internal spraing did not prevent PMTV invading tissue outside the brown arcs; its rate of spread was about 10 μm/h at 14–18d?.     

HRE-type genes are regulated by growth-related changes in internal oxygen concentrations during the normal development of potato (Solanum tuberosum) tubers

The occurrence of hypoxic conditions in plants not only represents a stress condition but is also associated with the normal development and growth of many organs, leading to adaptive changes in metabolism and growth to prevent internal anoxia. Internal oxygen concentrations decrease inside growing potato tubers, due to their active metabolism and increased resistance to gas diffusion as tubers grow. In the present work, we identified three hypoxia-responsive ERF (StHRE) genes whose expression is regulated by the gradual decrease in oxygen tensions that occur when potato tubers grow larger. Increasing the external oxygen concentration counteracted the modification of StHRE expression during tuber growth, supporting the idea that the actual oxygen levels inside the organs, rather than development itself, are responsible for the regulation of StHRE genes. We identified several sugar metabolism-related genes co-regulated with StHRE genes during tuber development and possibly involved in starch accumulation. All together, our data suggest a possible role for low oxygen in the regulation of sugar metabolism in the potato tuber, similar to what happens in storage tissues during seed development.     

Carbohydrate metabolism during cold-induced sweetening of potato tubers

The aim of this work was to discover the effects of lowering the temperature from 25° to 2° on the metabolism of glucose [U-¹⁴C] by tubers of Solanum tuberosum. Isotope was applied to tubers via a 50-μl hole made with a capillary pipette. Tubers were incubated for 2 hr, the pulse; then the glucose- [U-¹⁴C] was replaced with glucose, and incubation was continued for 18 hr, the chase. The detailed distribution of ¹⁴C was determined at the end of the pulse and at the end of the chase at 2°, and compared with those found at 25°. Lowering the temperature reduced the proportion of metabolized ¹⁴C that entered the respiratory pathways. At 2°, but not at 25°, hexose phosphates were the most heavily labelled fraction after the pulse: during the chase at 2° much of this label was metabolized to sucrose. We conclude that lowering the temperature preferentially restricts glycolysis and diverts hexose phosphates to sucrose. We suggest that this is an important cause of cold-inducing sweetening of the tubers and is due to cold-lability of key glycolytic enzymes.     

Lipid metabolism during aging of high-alpha-linolenate-phenotype potato tubers

Previous studies demonstrated that high levels of alpha-linolenate in cell membranes of potato tubers (achieved by overexpressing fatty acid desaturases) enhances lipid peroxidation, oxidative stress, and tuber metabolic rate, effectively accelerating the physiological age of tubers. This study details the changes in lipid molecular species of microsomal and mitochondrial membranes from wild-type (WT) and high-alpha-linolenate tubers during aging. The microsomal and mitochondrial polar lipids of high-alpha-linolenate tubers were dominated by 18:3/18:3 and 16:0/18:3 molecular species. Relative to WT tubers, high-alpha-linolenate tubers had a substantially higher 16:0/18:n to 18:n/18:n molecular species ratio in mitochondria and microsomes, potentially reflecting a compensatory response to maintain membrane biophysical properties in the face of increased unsaturation. Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) accounted for 53 and 37% of polar lipids, respectively, in mitochondria from younger WT and high-alpha-linolenate tubers. The relative proportions of these phospholipids (PL) did not change during aging of WT tubers. In contrast, PE increased to dominate the PL pool of mitochondria during aging of high-alpha-linolenate tubers. While aging effected an increase in mitochondrial 18:3-bearing PCs and PEs in WT tubers, the concentration of 18:3-bearing PCs fell with a concomitant increase in 18:3-bearing PEs during aging of high-alpha-linolenate tubers. These age- and high-alpha-linolenate-induced changes had no effect on the respiration rate and functional integrity of isolated mitochondria. Differential increases in the respiration rates of WT and high-alpha-linolenate tubers during aging were therefore a consequence of unsaturation-dependent alterations in the microenvironments of cells. Microsomal 18:3-bearing PCs, PEs, digalactosyldiacylglycerols (DGDG), and monogalactosyldiacylglycerols all increased in WT tubers during aging. In contrast, a selective loss of 18:3-bearing PCs and DGDGs from microsomes of high-alpha-linolenate tubers likely reflects a greater susceptibility of membranes to peroxidative catabolism during aging. Aging resulted in an increase in sterol/PL ratio in microsomes from WT tubers, due primarily to a decline in PL. In high-alpha-linolenate tubers, the increase in sterol/PL ratio during aging was due to increases in Delta 5-avenasterol and stigmasterol, indicating membrane rigidification and likely contributing to increased membrane permeability. Age-induced changes in 18:3-bearing lipids in membranes of transformed tubers are discussed relative to the development of oxidative stress and accelerated aging.     

Induction and development of potato tubers in a jar fermentor

Potato (Solanum tuberosum L.) tubers were mass propagated in a jar fermentor using a two-step method consisting of a shoot multiplication step (phase 1) followed by a tuber induction and development step (phase 2). Tuberization was observed within 2 weeks of phase 2 and the number of tubers did not increase after this culture period. In contrast, total tuber weight continuously increased for at least 10 weeks. Although the number of tubers and the total tuber weight clearly decreased under the lower temperature (17°C), this weight decrease was partially prevented by changing the temperature from 17°C to 25°C after 2 weeks of phase 2. This result indicates that tuber development can be controlled independently of induction. Lower temperatures influenced the localization and size distribution of tubers in the jar fermentor.     

Lactate metabolism in potato tubers deficient in lactate dehydrogenase activity

The aim of this work was to investigate the effect of decreased activity of lactate dehydrogenase (EC 1.1.1.27; LDH) on lactate metabolism in potato tubers. By expressing a cDNA‐encoding potato tuber LDH in the antisense orientation, we generated transgenic potato plants with a preferential decrease in two of the five isozymes of LDH. Surprisingly, transgenic tubers grown under normoxic conditions did not contain less lactate, but rather instead contained approximately two‐fold more lactate than control tubers. This result is explicable if the decreased isozymes of LDH are responsible for the oxidation of lactate to pyruvate in vivo. This was confirmed by measurements of the rate of metabolism of lactate supplied to tuber discs: the rate in transgenic tubers was approximately half that of control tubers. The decrease in LDH activity had no measurable effect on the accumulation of lactate in cold‐stored tubers under anoxia, nor during the subsequent utilization of this lactate upon return to normoxia. In both control and transgenic tubers, the accumulation of lactate during anoxia was not accompanied by an induction of LDH activity or a change in isozyme distribution. In contrast, the metabolism of lactate after a period of anoxia was accompanied by a two‐fold increase in LDH activity and the induction of two isozymes that were distinct from those which had been decreased in the transgenic plants.     

Light-induced changes in the lipid composition and ultrastructure of plastids from potato tubers

Sandelius, A. S. and Liljenberg, C. 1982. Light-induced changes in the lipid composition and ultrastructure of plastids from potato tubers. – Physiol. Plant. 56: 266–272.
Amyloplasts and starch containing plastids from green tissue – amylochloroplasts – from potato tubers ( Solanum tuberosum L., var. King Edward) were separated from other cell organelles by sedimentation in a discontinuous sucrose gradient. Their lipid composition was analysed with emphasis on galactolipids and phospholipids and the fatty acid compositions of these lipids. Irradiation of the tubers caused increased ratios of monogalactosyl diacylglycerol to digalactosyl diacylglycerol and of total galactolipids to total phospholipids in the plastid membranes. Furthermore, the degree of unsaturation of the fatty acids increased in all lipid classes analysed, this effect being most prominent in the galactolipids. The ultrastructural studies made on tuber tissue revealed that irradiation caused a change in starch grain size distribution concomitant with formation of membrane structures resembling grana within the envelope. In many cases prolamellar bodies and plastoglobuli were present.     

Nitrogen recycling during phenylpropanoid metabolism in sweet potato tubers.

In the first step of the phenylpropanoid metabolic pathway, L-phenylalanine (L-Phe) is deaminated to form E-cinnamate, in a conversion catalyzed by phenylalanine ammonia-lyase (PAL; EC 4.3.1.5). The metabolic fate of the ammonium ion (NH4+) produced in this reaction was investigated in sweet potato (Ipomoea batatas) tuber discs. [15N]-Labeled substrates including L-Phe, in the presence or absence of specific enzyme inhibitors, were administered to sweet potato discs in light under aseptic conditions. 15N-Nuclear magnetic resonance spectroscopic analyses revealed that the 15NH4+ liberated during the PAL reaction is first incorporated into the amide nitrogen of L-glutamine (L-Gln) and then into L-glutamate (L-Glu). These results extend our previous observations in pine and potato that PAL-generated NH4+ is assimilated by the glutamine synthetase (GS; EC 6.3.1.2)/glutamate synthase (GOGAT; EC 1.4.1.13) pathway, with the NH4+ so formed ultimately being recycled back to L-Phe via L-Glu as aminoreceptor and donor.     

Sugar accumulation in chemically debudded potato tubers during cold storage

Potato tuber buds may be excised by immersion of the tubers in a mixture of EtOH-Me₂CO (1:1) for 4 hr. This enabled the study of the effect of tuber aging (at 17°) on the starch-to-sugar conversion during storage at 4°, in the absence of complications due to sprouting. Sugar accumulation during a two-week period of storage at 4° decreased with increasing time of prior storage at 17°.     

Effect of low temperature on the activity of phosphofructokinase from potato tubers

The aim of this work was to compare the coldlability of phosphofructokinase (EC 2.7.1.11) from tubers of potato cultivars (cvs.) known to differ in their propensity to accumulate sugars at low temperature. When stored at 4°C for six weeks, the sugar content of tubers ofSolanum tuberosum L. cv. Record doubled whereas the amount of sugar in tubers of cv. Brodick and an advanced breeding clone (13676) decreased slightly. Tubers from each line contained four forms of phophofructokinase. Over the range 12°–16°C the temperature coefficients of the four forms of phosphofructokinase from cvs. Record and Brodick were similar. In cv. Record the temperature coefficients of three of the enzyme forms were significantly higher at 2°–6°C than at 12°–16°C, whereas those from cv. Brodick were unchanged. These results are consistent with the proposal that inactivation of phosphofructokinase at low temperature results in the accumulation of hexose phosphates leading to increased sucrose synthesis.     

Effects of dithiothreitol on dopa oxidase activity from potato tubers

A filtrate, prepared from potato tuber by grinding in an isotonic medium, has been separated into a particulate and a ‘soluble’ fraction by ultracentrifugation. Following dialysis and lyophilization, both fractions catalysed the oxidation of l-DOPA, with approximately 30% of the l-DOPA: oxygen-oxidoreductase (EC 1.14.18.1; DOPA oxidase) activity being associated with the particulate fraction. When dithiothreitol (DTT, 10^?2M was included in the grinding medium, much lower yields of DOPA oxidase were obtained and 80% appeared to be associated with the particulate fraction. DTT proved to be a powerful inhibitor of DOPA oxidase. With concentrations of DTT causing only partial inhibition, the kinetics of the inhibited rate of dopachrome formation from l-DOPA were complex. When oxygen consumption was measured inhibition was not transient. The degree of inhibition was inversely related to the DOPA oxidase activity, indicating interaction of a product of this activity with DTT. Direct determination of -SH groups in DTT using 5,5′-dithiobis(2-nitrobenzoic acid (DTNB) showed that they were all oxidised during the initial phase of inhibition of dopachrome formation. It is concluded that the first phase of inhibition involves oxidation of DTT by an intermediate between l-DOPA and dopachrome. The second phase of inhibition also appeared to require -SH groups initially, since trans-4,5-dihydroxy-1,2-dithiane (oxidized DTT) caused very little inhibition at all.     

Effect of arachidonic acid on potato tubers during storage]

Potato (Solanum tuberosum L.) tubers were treated with various concentrations (10(-9) to 10(-4) M) of the biogenic elicitor arachidonic acid during the period of storage (from October to July). The data showed that the resistance-inducing concentration of arachidonic acid was 10(-6) M in autumn and 10(-9) M in spring. Possible causes of the change in the immunizing concentration of arachidonic acid during storage of potato tubers are discussed.     

Increased mitochondrial DNA and RNA polymerase activity in ethylene-treated potato tubers

A purified mitochondrial fraction was isolated from potato (Solanum tuberosum L.) tubers respiring normally at 23°C or at an accelerated rate in response to treatment with ethylene (10 microliters per liter).

A pronounced increase in various mitochondrial enzymic activities was observed in response to exposure of the whole tubers to ethylene. Cytochrome c oxidase activity increased more than 50%, DNA polymerase activity increased about 2-fold, and RNA polymerase activity increased 2.5-fold. Moreover, DNA or RNA polymerase activities of mitochondria isolated from tubers not treated with ethylene were not affected by ethylene treatment in vitro. Respiratory control ratios decreased from 2.84 to 1.50 with increasing periods of ethylene treatment from 0 to 15 hours. None of these changes were observed in untreated tubers. It is concluded that the stimulation of respiration by ethylene in potato tubers is accompanied in vivo by an enhancement of mitochondrial enzymic activity of both membrane-associated enzymes which participate in the mitochondrial oxidative electron transport as well as soluble enzymes which are not directly involved in respiration.