Juan J. Guiamet

The Photosynthetic Role of Ears in C3 Cereals : Metabolism, Water Use Efficiency and Contribution to Grain Yield

This review concerns ear photosynthesis and its contribution to grain filling in C3 cereals. Ear photosynthesis is quantitatively important to grain filling, particularly in dry areas where source (i.e., assimilate) limitations can occur. Compared to the flag leaf, ear photosynthesis exhibits higher water stress tolerance. Several factors could be involved in the ears “drought tolerance.” First, although degree of C4 metabolism in ear parts has been reported, current evidence supports only typical C3 metabolism. Second, recycling of respired CO2 (i.e., refixation) could have considerable impact on final crop yield by preventing loss of CO2. Because refixation of CO2 is independent of atmospheric conditions, water use efficiency (measured as total ear photosynthesis divided by transpiration) could be higher in the ear than in the flag leaf. Moreover, ear parts (in particular awns) show higher relative water content and better osmotic adjustment under water stress compared to the flag leaf. This capacity, in addition to persistence of photosynthetic components under drought (delayed senescence), might help the ear to continue to fix CO2 late in the grain filling period.

Senescence‐associated degradation of chloroplast proteins inside and outside the organelle

Leaf proteins, and in particular the photosynthetic proteins of plastids, are extensively degraded during senescence. Although this involves massive amounts of protein, the mechanisms responsible for chloroplast protein degradation are largely unknown. Degradation within the plastid itself is supported by the observation that chloroplasts contain active proteases, and that chloroplasts isolated from senescing leaves can cleave Rubisco to release partially digested fragments. It is less clear whether chloroplasts can complete Rubisco degradation. Chloroplastic proteases are likely involved in the breakdown of the D1 and LHCII proteins of photosystem II. Small senescence-associated vacuoles (SAVs) with high-proteolytic activity develop in senescing leaf cells, and there is evidence that SAVs contain chloroplast proteins. Thus, an extra-plastidic pathway involving SAVs might participate in the degradation of some chloroplast proteins. Plastidic and extra-plastidic pathways might cooperate in the degradation of chloroplast proteins, or they might represent alternative, redundant pathways for photosynthetic protein degradation.

Thylakoid-Bound Ascorbate Peroxidase Mutant Exhibits Impaired Electron Transport and Photosynthetic Activity

In chloroplasts, stromal and thylakoid-bound ascorbate peroxidases (tAPX) play a major role in the removal of H2O2 produced during photosynthesis. Here, we report that hexaploid wheat (Triticum aestivum) expresses three homeologous tAPX genes (TaAPX-6A, TaAPX-6B, and TaAPX-6D) mapping on group-6 chromosomes. The tAPX activity of a mutant line lacking TaAPX-6B was 40% lower than that of the wild type. When grown at high-light intensity photosystem II electron transfer, photosynthetic activity and biomass accumulation were significantly reduced in this mutant, suggesting that tAPX activity is essential for photosynthesis. Despite the reduced tAPX activity, mutant plants did not exhibit oxidative damage probably due to the reduced photochemical activity. This might be the result of a compensating mechanism to prevent oxidative damage having as a consequence a decrease in growth of the tAPX mutant plants.

Ethylene as promoter of wheat grain maturation and ear senescence

This work was aimed at testing the involvement of ethylene in the maturation of grain and senescence of the foliar structures of the wheat inflorescence. Whole wheat ears emitted ethylene to the atmosphere. From pre-anthesis, ethylene emission progressively increased from 0.76 nl g−1FW h−1 to a peak 1.53 nl g−1FW h−1 at the hard dough stage of the grains, to fall to a minimum of 0.10 nl g−1FW h−1 at the dormant seed stage. Ethephon increased the ethylene release, hastened the process of grain maturation and senescence of the ears. Aminoethoxyvinylglycine and silver thiosulfate produced the opposite effects. It is concluded that ethylene plays a role in grain maturation and in the senescence of the green bracts of the inflorescence.

Antioxidant enzymes and lipid peroxidation during aging of Chrysanthemum morifolium RAM petals

Abstract To characterize the physiological status of petals over the senescence period, the rate of leakage of electrolytes and weight were measured in Chrysanthemum morifolium RAM petals. Both a significant increase in electrolyte leakage to the external medium and a concomitant decrease of flower weight were observed. Lipid peroxidation was evaluated by measurements of thiobarbituric acid reactive substance (TBARS) in Chrysanthemum morifolium RAM petals during senescence, that was arbitrarily divided into five stages. TBARS content was higher in petals from stage 5 (complete wilting) than in stage 1 (blooming). Non-significant differences were detected in ethylene production between petals classified in stages 1–3 (50 ± 4 pmol/g fresh weight/h). The activity of the enzymes involved in hydroperoxide metabolism was determined. The activities of superoxide dismutase (SOD), catalase, peroxidases and ascorbate peroxidase (AP) were measured. SOD activity showed a maximum value at stage 3 (245 ± 30 U/mg protein), followed by a decline. A progressive increase in peroxidase (5-fold increase) and AP (6-fold increase) activities was measured as a function of time. Petals in stage 1 showed lower catalase activity (20.8 ± 0.5 μmol/min/mg protein) than petals in stages 2–5 (43 ± 3 μmol/min/mg protein). The data presented here suggest that lipid peroxidation and membrane damage are involved in deterioration of Chrysanthemum morifolium RAM petals. The significant increase in the activity of SOD, catalase, peroxidases and AP in the initial stages of senescence, indicates that antioxidant defenses are triggered by coordinated mechanisms to control damage by aging in petals.

In vivo inhibition of cysteine proteases provides evidence for the involvement of ‘senescence-associated vacuoles’ in chloroplast protein degradation during dark-induced senescence of tobacco leaves

Breakdown of leaf proteins, particularly chloroplast proteins, is a massive process in senescing leaves. In spite of its importance in internal N recycling, the mechanism(s) and the enzymes involved are largely unknown. Senescence-associated vacuoles (SAVs) are small, acidic vacuoles with high cysteine peptidase activity. Chloroplast-targeted proteins re-localize to SAVs during senescence, suggesting that SAVs might be involved in chloroplast protein degradation. SAVs were undetectable in mature, non-senescent tobacco leaves. Their abundance, visualized either with the acidotropic marker Lysotracker Red or by green fluorescent protein (GFP) fluorescence in a line expressing the senescence-associated cysteine protease SAG12 fused to GFP, increased during senescence induction in darkness, and peaked after 2-4 d, when chloroplast dismantling was most intense. Increased abundance of SAVs correlated with higher levels of SAG12 mRNA. Activity labelling with a biotinylated derivative of the cysteine protease inhibitor E-64 was used to detect active cysteine proteases. The two apparently most abundant cysteine proteases of senescing leaves, of 40kDa and 33kDa were detected in isolated SAVs. Rubisco degradation in isolated SAVs was completely blocked by E-64. Treatment of leaf disks with E-64 in vivo substantially reduced degradation of Rubisco and leaf proteins. Overall, these results indicate that SAVs contain most of the cysteine protease activity of senescing cells, and that SAV cysteine proteases are at least partly responsible for the degradation of stromal proteins of the chloroplast.

Modulation of Progressive Leaf Senescence by the Red:Far-Red Ratio of Incident Light

Unifoliate soybean leaves were irradiated with different red:far-red ratios (R:FR) while the shoot above the leaves was kept under a constant R:FR. The R:FR was changed by lowering the red photon fluence rate. An R:FR of 0.07 accelerated chlorophyll and protein loss and enhanced the rate of exudation of amino acids from the detached irradiated leaves. Different photon fluence rates of far-red were added to a background of cool white fluorescent light, to lower R:FR without significantly changing photosynthetically active radiation. R:FR of 0.09-0.18 accelerated chlorophyll loss. These results are consistent with the view that the R:FR may modulate progressive senescence and that low R:FR occurring in shadelight may promote senescence.

Ethylene production and responses to exogenous ethylene in senescing petals of Chrysanthemum morifolium RAM cv. Unsei

Abstract Senescence of chrysanthemum (Chrysanthemum morifolium RAM cv. Unsei) petals was associated with a marked deterioration of the plasma membrane and loss of water. The rates of ethylene synthesis declined after full bloom, and became non-detectable by the time the first symptoms of senescence were apparent. The drop in ethylene production correlated with a reciprocal increase in 1-aminocyclopropane-1-carboxylic acid (ACC) content. At full bloom, pre-treatment of petals with 1 mM ACC increased their rates of ethylene production several fold, but the capacity to convert exogenous ACC into ethylene decline thereafter. Inhibitors of ethylene synthesis (i.e. aminoxyacetic acid) or ethylene action (i.e. silver thiosulfate) had no effect on senescence, implying that the minute amounts of ethylene synthesized by petals are below the threshold to cause any acceleration of this process. However, petal senescence was significantly accelerated by a 24 h treatment with 3 ppm of ethylene at full bloom, or by continuous treatment with ethephon and ACC at concentrations that produce a significant increase of ethylene synthesis. Thus, our results show that petal senescence in some varieties of chrysanthemum is sensitive to exogenous ethylene, and that the extended vase life that is characteristic of these flowers may be due to their low rates of ethylene synthesis.

Analysis of early vigour in twenty modern cultivars of bread wheat (Triticum aestivum L.)

Abstract. Fast development of seedling leaf area is a relevant trait in order to increase early resource acquisition. The use of semi-dwarf genotypes of wheat has decreased early vigour of modern cultivars. We studied early vigour of 20 cultivars cropped in Argentina, and our main objectives were: (i) to analyse the genotypic variability in early vigour; (ii) to study morphological traits that can be good indicators of early vigour, such as seed mass, leaf width, and specific leaf area; and (iii) to determine whether increased dry mass allocation to roots impacts negatively on early vigour. Experiments with non-size-selected and size-selected seeds were carried out in a greenhouse. A field trial was also conducted in order to test the reliability of the greenhouse results. Seeds mass was the main parameter related to early vigour. However, results from the experiment with seeds selected by size (45–50 mg seed–1) showed that seed mass per se only partially explains early vigour, since a significant coefficient of determination was observed between the seedling leaf area of each cultivar in both experiments (i.e. with randomly chosen or size-selected seeds). We observed a high coefficient of determination between net assimilation rate and changes in the ranking of early vigour of the cultivars with time after transplant. Root biomass was positively correlated with leaf area, indicating that the traits were not mutually exclusive. We built simple models by multiple regression to predict early vigour, including some parameters that were easy to measure. Seed mass and leaf width taken together showed better fit than seed mass or leaf width alone. We found a significant coefficient of determination between early vigour in greenhouse and field experiments; thus, screening for early vigour under semi-controlled conditions may be feasible.

Senescence-Associated Vacuoles, a Specific Lytic Compartment for Degradation of Chloroplast Proteins?

Degradation of chloroplasts and chloroplast components is a distinctive feature of leaf senescence. In spite of its importance in the nutrient economy of plants, knowledge about the mechanism(s) involved in the breakdown of chloroplast proteins is incomplete. A novel class of vacuoles, “senescence-associated vacuoles” (SAVs), characterized by intense proteolytic activity appear during senescence in chloroplast-containing cells of leaves. Since SAVs contain some chloroplast proteins, they are candidate organelles to participate in chloroplast breakdown. In this review we discuss the characteristics of SAVs, and their possible involvement in the degradation of Rubisco, the most abundant chloroplast protein. Finally, SAVs are compared with other extra-plastidial protein degradation pathways operating in senescing leaves.