Age-related changes in oxidative stress markers and abscisic acid levels in a drought-tolerant shrub, Cistus clusii grown under Mediterranean field conditions

Compared with our knowledge of senescence in annuals and biennials, little is known about age-related changes in perennials. To get new insights into the mechanisms underlying aging in perennials, we measured oxidative stress markers in leaves and organelles, together with abscisic acid levels in leaves of 2- and 7-year-old Cistus clusii dunal plants grown under Mediterranean field conditions. Recently emerged leaves, which either appeared during autumn or spring, were compared to evaluate the effects of environmental constraints on oxidative stress and abscisic acid accumulation as plants aged. Plant aging led to an enhanced oxidation of α-tocopherol and ascorbate, increased lipid peroxidation and reduced PSII efficiency in leaves during the more stressful conditions of spring and summer, but not during autumn. Analyses of lipid peroxidation in organelles isolated from the same leaves revealed that oxidative stress occurred both in chloroplasts and mitochondria. Although both plant groups showed similar leaf water and nitrogen contents throughout the study, abscisic acid levels were markedly higher (up to 75%) in 7-year-old plants compared to 2-year-old plants throughout the study. It is concluded that (a) meristematic tissues of C. clusii maintain the capacity to make new leaves with no symptoms of oxidative stress for several years, unless these leaves are exposed to environmental constraints, (b) leaves of oldest plants show higher oxidative stress than those of young plants when exposed to adverse climatic conditions, thus supporting the idea that the oxidative stress associated with aging is due at least partly to extrinsic factors, (c) at the subcellular level, age-induced oxidative stress occurs both in chloroplasts and mitochondria, and (d) even in the absence of environmental stress, newly emerged leaves accumulate higher amounts of ABA as plants age.     

Meristem aging is not responsible for age-related changes in growth and abscisic acid levels in the Mediterranean shrub, Cistus clusii

To obtain new insights into the mechanisms underlying aging in perennials, we measured abscisic acid levels, growth and other stress indicators in leaves of Cistus clusii Dunal plants of different ages grown under Mediterranean field conditions. Recently emerged leaves from 9-year-old plants were compared to those of 1-year-old plants (obtained from cuttings from 9-year-old plants) to evaluate the effects of meristem aging on plant aging. Rooting and successful establishment of the cuttings allowed us to compare the physiology of plants with old meristems, but of different size. Plants obtained from cuttings were rejuvenated, with new leaves displaying a higher leaf area and chlorophyll content, but smaller leaf mass per unit area ratios and endogenous abscisic acid levels than those of 9-year-old plants. A comparative study in 1-, 4- and 9-year-old plants revealed that abscisic acid levels increase during the early stages of plant life (with increases of 90% between 1- and 4-year-old plants), but then remain constant at advanced developmental stages (between 4- and 9-year-old plants). Although leaf biomass was 53% smaller in 9-year-old compared to 4-year-old plants, the dry matter produced per shoot apical meristem was equivalent in both plant groups due to an increased number of leaves per shoot in the former. It is concluded that (i) C. clusii plants maintain the capacity to rejuvenate for several years; (ii) newly emerged leaves accumulate higher amounts of abscisic acid during early stages of plant life, but the levels of this phytohormone later remain constant; and (iii) although plant aging leads to the production of smaller leaves, the amount of biomass produced per shoot apical meristem remains constant at advanced developmental stages.     

Plant aging and excess light enhance flavan-3-ol content in Cistus clusii

Physiological studies on aging in perennials are mainly focused either on the primary metabolism or the hormonal regulation of the process. However, to our knowledge, the involvement of the secondary metabolism in this process has not yet been explored. Cistus clusii, a Mediterranean sclerophyllous evergreen bush, shows considerable amounts of flavan-3-ols in leaves. In the present study, we aimed at determining the impact of environmental conditions and plant aging in the flavan-3-ol content in C. clusii plants grown in field conditions, which included summer drought and recovery periods. Six-year-old plants suffered more from photo-oxidative stress, especially during excess light periods, and showed lower maximum photosynthetic rates than 1-year-old plants. C. clusii leaves accumulated (−)-epigallocatechin gallate in early summer, in a strong positive correlation with both the photon flux density and the photoperiod, but not with the plant water status. Moreover, C. clusii plants accumulated proanthocyanidins (polymeric flavan-3-ols) in leaves during summer. Older plants showed higher levels of proanthocyanidins and (−)-epicatechin, but only during late spring and summer. From the result of the present study, we conclude that excess light enhances flavan-3-ol content in C. clusii, a process enhanced as plants age due to increased excess light stress.     

Axillary meristem development in Arabidopsis thaliana

Axillary shoot apical meristems initiate post-embryonically in the axils of leaves. Their developmental fate is a main determinant of the final plant body plan. In Arabidopsis, usually a single axillary meristem initiates in the leaf axil even though there is developmental potential for formation of multiple branches. While the wild-type plants rarely form multiple branches in the leaf axil, tfl1-2 plants regularly develop two or more branches in the axils of the rosette leaves. Axillary meristem formation in Arabidopsis occurs in two waves: an acropetal wave forms during plant vegetative development, and a basipetal wave forms during plant reproductive development. We report here the morphological and anatomical changes, and the STM expression pattern associated with the formation of axillary and accessory meristems during Arabidopsis vegetative development.     

Effects of microgravity on the structure of leaves cells involved in water transport

The oppression of plant cells growth and accelerated of their aging and cell structure changes take place during growth of plants at conditions of long time space flight or clinorotation. Such changes reflect the alterations of gravity-sensitive links in plant cell metabolism, of plant polarity and possible of water balance of plants at weightlessness. Because the study of plant cells ultrastructure changes that took part in water transport under microgravity influence was the aim of our investigation.     

PRIMORDIAL LEAF AND PHYTOHORMONE EFFECTS ON EXCISED SHOOT APICAL MERISTEMS OF COLEUS BLUMEI BENTH.

Coleus blumei Benth. apical meristems and apical meristems +1, +2, +3 primordial leaf pairs were cultured to examine phytohormone influences on development and correlative effects of developing primordial leaves on in vitro responses. The meristem with no phytohormones or low levels of IAA could not develop in vitro. At least 0.1 mg/l IAA and optimumly 1-2 mg/l IAA were required for development into complete plants. IAA from 0.1 to 3 mg/l also resulted in root development with no apparent leaf or shoot formation. Levels of IAA higher than 3 mg/l were inhibitory to development. Kinetin, as a substitute for naturally occurring cytokinins, alone (0.0003 to 3 mg/l) resulted in development of rosettes of leaves. In the presence of IAA (***1 mg/l) and kinetin (0.003 mg/l) plants, rosettes, individual leaves with roots, and roots developed from isolated meristems. Glutamine and adenine sulfate both appeared inhibitory to meristem development. With +1, +2, +3 developing primordial leaf pairs left attached to the apical dome, three pairs were required for plant formation in the absence of phytohormones. In the presence of IAA, two pairs of primordial leaves resulted in plant formation; whereas, with IAA and low levels of kinetin one pair of primordial leaves was enough. Higher levels of kinetin were inhibitory to plant development with primordial leaves present. ABA appeared to be inhibitory to development of meristems and meristems +1, +3 primordial leaves at low concentrations and resulted in death at ***1 mg/l. Developing primordial leaves appear to supply the apical meristem with a balance of phytohormones during growth. Meristem development into a plant first involved formation of leaf primordia. Establishment of a bipolar axis with root formation followed.     

Reproductive competence from an annual and a perennial perspective

Plants at early stages of development undergo a juvenile phase during which they are not competent to flower in response to environmental stimuli. The length of this phase varies among species and is extended in perennial plants particularly. In annuals, temporal changes in expression of microR156 (miR156), miR172, and their targets are correlated with the transition from the juvenile to the adult phase and flowering. This developmental transition in perennials is probably more complex than in other plants and the molecular mechanisms are less well understood. In addition, once perennials become adult and capable of reproduction they still keep some meristems in the vegetative state that contribute to their polycarpic growth habit. Juvenility and polycarpy, although considered as two different processes in perennials, might be related.     

Loss of flower bud vigour in the Mediterranean shrub,Cistus albidus L. at advanced developmental stages

To better understand aging in perennials, age‐related changes in the physiology of leaves and flower buds of the Mediterranean shrub, Cistus albidus L. were evaluated. Two groups of different ages (5 and 10 years old), both at advanced developmental stages but of similar size, were compared. Total plant biomass, biomass produced per apical meristem and levels of cytokinins, abscisic acid and jasmonic acid in leaves and flower buds, as well as flower production, were measured. No differences in plant size, vegetative growth rates and levels of phytohormones in leaves were observed between 5‐ and 10‐year‐old plants. However, they showed significant differences in flower bud development; the older plants having reduced vigour, with 29.6% of flowers reaching anthesis compared to 52.5% in the younger plants. Furthermore, endogenous concentrations of zeatin and abscisic acid in flower buds at stage I (start of flower organ formation) were 61% and 41%, respectively, smaller in 10‐ than in 5‐year‐old plants. At stage II (with all flower organs formed), zeatin and abscisic acid concentrations decreased by ca. 90% and 80%, respectively, but differences between age groups were still evident (60% and 29% for zeatin and abscisic acid, respectively). Jasmonic acid levels in flower buds decreased by 80% from stage I to II, but did not differ between age groups. Despite reductions in flower bud vigour, total number of flowers per individual was not significantly different between age groups, so that an age‐related loss in reproductive vigour at the organ level did not lead to a decrease in flower production at the whole plant level.     

Genetics of flower initiation and development in annual and perennial plants

Flowering is an integral developmental process in angiosperms, crucial to reproductive success and continuity of the species through time. Some angiosperms complete their life cycle within a year (annual plants), and others have a longer reproductive life, which is characterized by the generation of new flowering and vegetative shoots every year (perennial plants). Despite the differences in their lifespan, the underlying genetics of flower induction and floral organ formation appears to be similar among these plants. Hence, the knowledge gained from the study of flowering mechanism in Arabidopsis thaliana can be used to better understand similar processes in other plant species, especially the perennials, which usually have a long generation time and are not amenable to genetic analysis. Using Arabidopsis as a model, we briefly discuss the current understanding of the transition from vegetative to reproductive growth and the subsequent formation of individual floral organs, and how this knowledge has been successfully applied to the identification of homologous genes from perennial crops. Although annuals appear to share many similarities with perennials in terms of gene function, they differ in their commitment to flowering. Once an annual reaches the reproductive phase, all meristems are typically converted into either floral or inflorescence meristems. In contrast, each year, each meristem of a mature perennial has the choice to produce either a vegetative or a reproductive shoot. The physiology and genetics of flowering in Citrus are used to highlight the complexity of reproductive development in perennials, and to discus possible future research directions.     

Do perennials really senesce?

Although senescence remains less studied in perennials than in monocarpic plants, major advances in understanding senescence in perennials have been achieved recently. This success is due not only to the use of genetic tools in woody plants but also to a renaissance of research on how perennials can live for centuries or even millennia. The particularities of perennial life are considered here, with an emphasis on how these affect senescence at different levels of organization. I conclude that although cellular and leaf senescence do share common features in monocarpic and perennial plants, the indeterminacy of meristems found in perennials begs the question of whether senescence really occurs in these organisms at the whole-plant level.     

Effects of soil water level on the development of adult plant resistance to powdery mildew in barley

Barley grown in dry soil developed greater adult plant resistance (APR) to powdery mildew (Erysiphe graminis DC. f. sp. hordei Mérat) than barley grown in wet soil. Conidial germination and appressorium formation were less, and fungal development between formation of appressoria and elongating secondary hyphae on upper leaves was inhibited, when adult plants were grown in dry soil. Mildew colonies expanded more slowly on leaves of adult plants than on leaves of seedlings, especially if adult plants had grown in dry soil. APR was reduced if plants, previously grown in dry soil, were well watered more than 32 h before inoculation. Conidia originating from plants grown in dry soil had a lower solute potential and greater ability to infect plants grown in dry but not wet soil than conidia originating from plants grown in wet soil. APR could not be attributed simply to increased cell wall or cuticle thickness, nor to lowered leaf solute potentials, as has sometimes been suggested for powdery mildew diseases. Increasing plant age and water stress induced increases in cell wall and cuticle thickness, but these changes did not always coincide with changes in disease resistance. Increasing plant age and water stress also lowered leaf solute potentials but fungal solute potentials were lower than leaf solute potentials and, more importantly, were lower than leaf water potentials. Thus, fungal growth was not limited by the availability of water from the host during penetration and hyphal establishment. It is suggested that resistance levels may be determined not by the thickness of epidermal structures, nor by lowering of solute potential per se, but by specific substances harmful to the fungus which accumulate in either cell wall, cuticle or sap, and whose concentration is dependent on the age and water stress of leaves.     

Ectopic expression of soybean GmKNT1 in Arabidopsis results in altered leaf morphology and flower identity

Plant morphology is specified by leaves and flowers, and the shoot apical meristem (SAM) defines the architecture of plant leaves and flowers. Here, we reported the characterization of a soybean KNOX gene GmKNT1, which was highly homologous to Arabidopsis STM. The GmKNT1 was strongly expressed in roots, flowers and developing seeds. Its expression could be induced by IAA, ABA and JA, but inhibited by GA or cytokinin. Staining of the transgenic plants overexpressing GmKNT1-GUS fusion protein revealed that the GmKNT1 was mainly expressed at lobe region, SAM of young leaves, sepal and carpel, not in seed and mature leaves. Scanning electron micros- copy (SEM) disclosed multiple changes in morphology of the epidermal cells and stigma. The transgenic Arabidopsis plants overexpress- ing the GmKNT1 showed small and lobed leaves, shortened internodes and small clustered inflorescence. The lobed leaves might result from the function of the meristems located at the boundary of the leaf. Compared with wild type plants, transgenic plants had higher ex- pression of the SAM-related genes including the CUP, WUS, CUC1, KNAT2 and KNAT6. These results indicated that the GmKNT1 could affect multiple aspects of plant growth and development by regulation of downstream genes expression.     

Node-specific branching and heterochronic changes underlie population-level differences in Mimulus guttatus (Phrymaceae) shoot architecture

? Premise of the study: Shoot architecture is a fundamentally developmental aspect of plant biology with implications for plant form, function, reproduction, and life history evolution. Mimulus guttatus is morphologically diverse and becoming a model for evolutionary biology. Shoot architecture, however, has never been studied from a developmental perspective in M. guttatus. ? Methods: We examined the development of branches and flowers in plants from two locally adapted populations of M. guttatus with contrasting flowering times, life histories, and branch numbers. We planted second-generation seed in growth chambers to control for maternal and environmental effects. ? Key results: Most branches occurred at nodes one and two of the main axis. Onset of branching occurred earlier and at a greater frequency in perennials than in annuals. In perennials, almost all flowers occurred at the fifth or more distal nodes. In annuals, most flowers occurred at the third and more distal nodes. Accessory axillary meristems and higher-order branching did not influence shoot architecture. ? Conclusions: We found no evidence for trade-offs between flowers and branches because axillary meristem number was not limiting: a large number of meristems remained quiescent. If, however, quiescence is a component of meristem allocation strategy, then meristems may be limited despite presence of quiescent meristems. At the two basalmost nodes, branch number was determined by mechanisms governing either meristem initiation or outgrowth, rather than flowering vs. branching. At the third and more distal nodes, heterochronic processes contributed to flowering time and branch number differences between populations.     

THE MORPHOLOGICAL EFFECTS OF PROTEIN SYNTHESIS INHIBITION IN MARSILEA

Marsilea vestita and M. drummondii were grown in sterile cultures to which concentrations of the protein synthesis inhibitors, 2-thiouracil (10 mg/liter) and 5-fluorouracil (1 mg/liter) had been added. When young sporelings are grown in a solution of thiouracil at optimum concentration, there is an inhibition of the rate of leaf formation, a retardation of the leaf heteroblastic series, and all leaves develop as land forms. When thiouracil is added to plants which are already producing typical adult, quadrifid leaves, the effects depend on whether the treated plants are water or land forms. Plants which are typically water forms convert to land forms. After treatment successive leaves develop typical sunken stomata on both leaf surfaces. The tissues of the rhizome, root and petiole are more compact and, in general, the cells of the plant have thicker walls. Vascular patterns are not changed, though the size of the rhizome, root and petiole may be reduced. Plants which are typically land forms are less affected than the water forms, but they show a small reduction in apex volume and an apparent reversion of the leaflet number from the typical quadrifid leaf to a trifid, bifid, or single lamina condition. In both land and water forms apical dominance may be broken by treatment with 10 mg/liter thiouracil or 1 mg/liter fluorouracil and numerous lateral branches develop. Higher concentrations (15–25 mg/liter of thiouracil) may result in abnormal development of lateral axillary buds, petiole bases and leaflets. The meristems of the plant are differentially sensitive to thiouracil; leaflet meristems are most sensitive, the root meristems are the least sensitive. It appears that a true reversion to juvenile leaf development need not occur even though protein synthesis and the volume of the apex are reduced. The development of the land or water form in Marsilea appears to depend on rate of growth. Hence inhibition of the growth of typical water forms, through inhibition of protein synthesis, causes a shift in development toward the morphology typical of land forms.     

Influence of arbuscular mycorrhiza on growth and reproductive response of plants under water deficit: a meta-analysis

Despite a large body of literature that describes the effects of arbuscular mycorrhizal colonization on plant response to water deficit, reviews of these works have been mainly in narrative form, and it is therefore difficult to quantify the magnitude of the effect. We performed a meta-analysis to examine the effect of mycorrhizal colonization on growth and yield of plants exposed to water deficit stress. Data were compared in the context of annual vs. perennial plants, herbaceous vs. woody plants, field vs. greenhouse conditions, degree of stress, functional group, regions of plant growth, and mycorrhizal and host species. We found that, in terms of biomass measurements, mycorrhizal plants have better growth and reproductive response under water stress compared to non-mycorrhizal plants. When variables such as habit, life cycle, or water stress level are considered, differences in mycorrhizal effect on plant growth between variables are observed. While growth of both annual and perennial plants is improved by symbiosis, perennials respond more favorably to colonization than annuals. Overall, our meta-analysis reveals a quantifiable corroboration of the commonly held view that, under water-deficit conditions, plants colonized by mycorrhizal fungi have better growth and reproductive response than those that are not.     

Phosphorus Accretion in Old Leaves of a Mediterranean Shrub Growing at a Phosphorus-Rich Site

Studies dealing with changes in the plant internal nutrient cycling in response to natural, long-term P-fertility gradients are scarce. In this short report, we show some evidence on how leaf P dynamics can be drastically altered when plants typical from nutrient-poor sites grow in long-term P-enriched soils. The study was conducted in two natural populations of the Mediterranean evergreen shrub Pistacia lentiscus L.: one in a P-poor site and the other in a P-rich site. Soil texture and N, P, and organic matter contents were measured at each site. Leaf N and P concentrations were also measured in current-year, 1-year-old, and 2-year-old leaves, and in the senesced leaves. In the P-poor site, leaf P and N decreased as the leaves aged. This occurs because of nutrient reabsorption to other plant organs and/or dilution of nutrients by carbon compounds. In contrast, the leaves from the P-rich site acummulated P (but not N) during leaf lifespan. Consequently, P concentration in senesced leaves was very high in the P-rich site. These results show that, in long-lived perennials living in the field, long-term P enrichment can switch the normal process of P resorption during leaf aging to P accretion in the leaf. P accumulation in the leaves, which are periodically shed, might constitute a simple P excretion mechanism for plants typical from P-poor soils.     

A greenprint for growth: signalling the pattern of proliferation

The shoot and root apical meristems (SAM and RAM, respectively) of plants serve both as sites of cell division and as stem cell niches. The SAM is also responsible for the initiation of new leaves, whereas the analogous process of lateral root initiation occurs in the pericycle, a specialized layer of cells that retains organogenic potential within an otherwise non-dividing region of the root. A picture is emerging of how cell division, growth, and differentiation are coordinated in the meristems and lateral organ primordia of plants. This is starting to reveal striking parallels between the control of stem cell maintenance in both shoots and roots, and to provide information on how signalling from developmental processes and the environment impact on cell behaviour within meristems.     

MORPHOLOGY OF MARSILEA VESTITA. III. MORPHOGENESIS OF THE LEAVES OF ETIOLATED PLANTS

The laminae of etiolated Marsilea vestita leaves develop by means of marginal meristems. Unlike light-grown plants, the form of the etiolated plant is not affected by growth on a solid medium. All of the young leaves isolated from light-grown submerged plants will elongate in darkness. The smallest, etiolated, uncoiled leaves develop into land leaves when they are placed in light, and this development occurs regardless of whether the leaves remain on the plant or are isolated on nutrient agar. Only these smallest leaves (2.3 mm average length) are actually capable of being converted from a submerged leaf form to a land leaf form by darkness.     

Plant aging increases oxidative stress in chloroplasts

Aging has received considerable attention in biomedicine, but little is known about the regulatory mechanisms responsible for the aging not associated with senescence in plants. This study provides new insights into the relationship between oxidative stress and plant aging, and points out chloroplasts as one of the target organelles of age-associated oxidative stress in plants. We simultaneously analyzed lipid oxidation, photosynthesis, chlorophyll content, de-epoxidation state of the xanthophyll cycle, and levels of chloroplastic antioxidant defenses such as beta-carotene and alpha-tocopherol in leaves of the same age in 1-, 3- and 7-year-old Cistus clusii Dunal plants growing under Mediterranean field conditions. Enhanced formation of malondialdehyde in leaves (2.7-fold) and chloroplasts (2.8-fold), decreased photosynthetic activity (25%), and lower chlorophyll (ca. 20%) and chloroplastic antioxidant defense levels (ca. 25%-85%) were observed in 7-year-old plants, when compared with 1- and 3-year-old plants. The differences observed, which were associated with plant aging, were only noticeable in mature nonsenescing plants (7-year-old plants). No differences were observed between pre-reproductive (1-year-old plants) and young plants (3-year-old plants). This study shows that from a certain age, oxidative stress increases progressively in chloroplasts as plants age, whereas photosynthesis is reduced. The results indicate that the oxidative stress associated with the aging in plants accumulates progressively in chloroplasts, and that the contribution of oxidative stress to aging increases as plants age.     

Defoliation and Regrowth in the Graminaceous Plant: The Role of Current Assimilate

Infra-red gas analysis and a quantitative radiocarbon tracertechnique were used to measure photosynthesis, and the export,distribution and utilization of current assimilate in the regrowthof leaf tissue and the growth of stem and root of partially-defoliateduniculm barley plants. After defoliation, which removed allleaf tissue above the ligule of leaf 3, the rate of photosynthesisof the remaining two older leaves fell to 90–95 per centof that of control leaves, but they exported more of their assimilatedcarbon to meristems elsewhere in the plant during the first48 h after the defoliation. The level of export from the twoolder leaves began to decline when new leaf tissue regrew fromthe shoot apex, and fell below that of the control leaves 4days after defoliation. The two older leaves supplied the assimilateused in the regrowth of new leaf tissue immediately after defoliation:previously they had exported most of their assimilate to root.There was no evidence that ‘reserves’ were mobilizedto meet the needs of regrowth at leaf meristems or, indeed,of the growth in stem and root; current photosynthesis suppliedsufficient assimilate to account for all observed growth. Ingeneral, the plants responded to defoliation with a rapid andmarked re-allocation of assimilate from root to leaf meristems,with the result that root growth was severely retarded but newleaf tissue grew at 70–100 per cent of the rate observedin control plants.