Chloroplast Responses Research Articles

Differential Protein Phosphorylation Regulates Chloroplast Movement in Response to Strong Light and Darkness in Arabidopsis thaliana

Phototropin-dependent chloroplast movement is essential to the photosynthetic acclimation of mesophyll cells to incident light. Chloroplast movement involves many cellular actors, such as chloroplast-associated actin filaments and proteins that mediate signalling between phototropins and chloroplast motion. In the past few years, genetic approaches have identified several key proteins but the intrinsic mechanisms of the signalling cascade, such as phosphorylation events, remain undefined. Here, we took advantage of phosphoproteomics to examine the involvement of protein phosphorylation in chloroplast movement in darkness or under high light, at different CO2 mole fractions (100, 380 or 1,000 ppm) to vary photosynthetic activity. Amongst the 100 relevant identified phosphopeptides, 19 (corresponding to 8 proteins) were differentially phosphorylated in darkness vs. high light. There was no significant CO2 effect on the observed phosphorylation patterns. We further characterized the phosphorylation sites in THRUMIN1, which is believed to be crucial for the attachment of chloroplast-associated actin filaments to the plasma membrane and thus for chloroplast movements. The mutant thrumin1 was complemented with a mutated protein in which phospho-sites were substituted to a phosphomimetic (Asp) or a non-phosphorylatable (Ala) residue. While the phosphomimetic substitution altered the chloroplast response in the light only, both light and dark responses were altered with the non-phosphorylatable substitution. Our data suggest a key role of protein phosphorylation, including that of THRUMIN1, in the light/dark control of chloroplast movements.

A Thylakoid-Located Two-Pore K + Channel Controls Photosynthetic Light Utilization in Plants

The size of the light-induced proton motive force (pmf) across the thylakoid membrane of chloroplasts is regulated in response to environmental stimuli. Here, we describe a component of the thylakoid membrane, the two-pore potassium (K(+)) channel TPK3, which modulates the composition of the pmf through ion counterbalancing. Recombinant TPK3 exhibited potassium-selective channel activity sensitive to Ca(2+) and H(+). In Arabidopsis plants, the channel is found in the thylakoid stromal lamellae. Arabidopsis plants silenced for the TPK3 gene display reduced growth and altered thylakoid membrane organization. This phenotype reflects an impaired capacity to generate a normal pmf, which results in reduced CO2 assimilation and deficient nonphotochemical dissipation of excess absorbed light. Thus, the TPK3 channel manages the pmf necessary to convert photochemical energy into physiological functions.

Decoding the role of phosphoinositides in phototropin signaling involved in chloroplast movements

In angiosperms, light-dependent chloroplast movements are exclusively mediated by UVA/blue light receptors - phototropins. The two photoreceptors of Arabidopsis thaliana, phot1 and phot2, have overlapping roles in the control of these movements. Experiments performed in different plant species point to the participation of phosphoinositides in blue light-controlled chloroplast relocations. Here, we report a summary of recent findings presenting the involvement of phosphatidylinositol 4,5-bisphosphate as well as phosphatidylinositol 3- and 4-phosphates in weak blue light-mediated (accumulation) and strong blue light-mediated (avoidance) responses of chloroplasts. The blue light-activated alterations in phosphoinositide concentration are partly responsible for cytosolic Ca2+ changes. Ca2+ influx from apoplast does not seem to be involved in the mechanism of movement responses. In summary, interplay between phosphoinositides and intracellular Ca2+ regulates chloroplast redistribution in response to blue light in higher plants.

The chloroplast. Basics and applications

The chloroplast. Basics and applications

Erratum to: External K+ Deficiency Inhibits Photosynthetic Activity Through Superoxide Anion Production in Protoplasts Isolated from the Thallus of Ulva pertusa

To investigate the effects of potassium on photosynthetic activity in the green alga Ulva pertusa, we enzymatically isolated protoplasts from thallus samples. Photochemical quenching showed that protoplasts were capable of electron transport by photosystem II (PS II) during illumination. This quenching was dependent on external pH, with a reduced electron transport rate at pH >6.8 and less ability to use HCO 3 − under alkaline conditions. In the presence of external Na+, K+ enhanced PS II quantum yield, indicating a functional role for K+ during photosynthesis. That yield was enhanced in a [K+]-dependent manner, with maximum activity at 100 mM. However, potassium alone did not maintain photochemical activity, and its addition supported photosynthetic O2 evolution only in the presence of Na+. A deficiency of K+ led to the production of superoxide anions. Because of that generation, activities of superoxide dismutase and ascorbate peroxidase, two key enzymes involved in scavenging reactive oxygen species in the water–water cycle, also increased during such stress. These results strongly suggest that a series of ROS-scavenging systems are initiated in Ulva chloroplasts in response to K+ deficiency and that enzyme activities might protect algal cell photosynthesis.

Genotypic differences in post-storage photosynthesis and leaf chloroplasts in response to ethylene and 1-methylcyclopropene in Aglaonema

This study investigated the effects of ethylene in storage and 1-methylcyclopropene (1-MCP) pretreatment on post-storage leaf senescence as measured by changes in photosynthesis and chloroplast degradation of two Aglaonema cultivars. Potted plants of ‘Chalit's Fantasy’ and ‘White Tip’ with or without 1-MCP treatment (600nLL−1 1-MCP for 6h) were exposed to 3.0μLL−1 ethylene, while being stored for 5d at 16°C in the dark, and then placed under an indoor environment for further observation. Plants that did not receive 1-MCP and ethylene served as controls. Ethylene did not affect the stomatal conductance in either cultivar. Ethylene reduced the net CO2 assimilation rate and Fv/Fm (potential photochemical efficiency of photosystem II) in ‘White Tip’, but not in ‘Chalit's Fantasy’. Chloroplast number in a palisade or spongy mesophyll cell did not differ among treatments in ‘Chalit's Fantasy’. However, ethylene-treated ‘White Tip’ had fewer chloroplasts in the mesophyll cells, had more and larger plastoglobules in the chloroplasts, and had looser granal stacking with enlarged thylakoid lumens. ‘Chalit's Fantasy’ plants that were treated with 1-MCP before exposure to ethylene had higher net CO2 assimilation rates and stomatal conductance than the control or plants that were exposed to ethylene without 1-MCP pretreatment. 1-MCP pretreatment mitigated the injurious effect of ethylene on ‘White Tip’ by increasing net CO2 assimilation rate and Fv/Fm, and maintaining the quantity and structural integrity of chloroplasts.

Blue light signalling in chloroplast movements

Chloroplast movements are among the mechanisms allowing plants to cope with changes in their environment. Chloroplasts accumulate at illuminated cell areas under weak light while they avoid areas exposed to strong light. These directional responses may be controlled by blue and/or red light, depending on the plant group. In terrestrial angiosperms only the blue light perceived by phototropins is active. The last decade has seen a rapid development of studies on the mechanism of directional chloroplast movements, which started with an identification of the photoreceptors. A forward genetic approach has been used to identify the components which control chloroplast movements. This review summarizes the current state of research into the signalling pathways which lead to chloroplast responses. First, the molecular properties of phototropins are presented, followed by a characterization both of proteins which are active downstream of phototropins and of secondary messengers. Finally, cross-talk between light signalling involved in chloroplast movements and other signalling pathways is discussed.

Coarse-Grained Model of Cooperative Chloroplast Transport in Moss

Organelle motility is essential for the functioning of the eukaryotic cell. Actively modifying intracellular structures allows cells to change and adapt to different conditions. One of these cellular structures is the microtubule cytoskeleton, which is comprised of polarized filaments that function as tracks to transport cargo via molecular motors. Recent studies have revealed the importance of cooperative transport in living cells. In the moss Physcomitrella patens, reorganization of the chloroplasts is critical to adapt to changes in light quality and intensity. In this work, we performed detailed analysis of the transport of chloroplasts in protonemal moss cells, and showed that their transport is facilitated by molecular motors and the microtubule cytoskeleton. Our findings were recapitulated using coarse-grained modeling of this cargo transport over different microtubule network topologies. Our simulations include a detailed description of motor and microtubule dynamics, in the presence of thermal fluctuations. Our modeling sheds light into the molecular mechanisms at play in the photo-relocation response of chloroplasts in moss.

Soil metals, chloroplasts, and secure crop production: a review

An increasing number of soils of poor quality with unbalanced metal concentrations are used worldwide for crop cultivation. Even though plants are able to develop strategies to cope with metal stress, either metal deficiency or excess, this unbalance affects the whole plant. Chloroplasts are key organelles for organic matter synthesis and biomass production. Under metal stress, chloroplasts suffer severe alterations of their ultrastructure, associated with profound molecular and metabolic damages. These alterations are accompanied by unbalanced metal distribution in plants, in particular in edible crop organs. Toxic metals get either accumulated or there is a deficiency of nutrients, resulting in a weak nutritional value. Nonetheless, there is more and more knowledge on the functioning and regulation of metal transporters in plants. Such knowledge will allow growing crops with well-balanced metal concentrations in edible parts even on metal unbalanced soils. This review shows that almost all vital functions of chloroplasts, such as photosynthesis, CO2 fixation, nitrogen and sulfur assimilation, and protein and nucleic acid metabolism, require metals. Therefore, the uptake of essential metals is necessary for the proper functioning of chloroplasts and, in turn, for crop productivity. We describe nutrient uptake mechanisms of plants and processes that influence essential and non-essential metal concentrations in different plant organs. We present an overview of metal transporters in chloroplasts. Several questions still need to be elucidated about the uptake and the trafficking of essential and non-essential metals into and within chloroplasts. Similarly to transporters present in other cellular compartments, the carriers are often not metal-specific. Therefore, essential and non-essential metals may compete for carriers. As a result, unbalanced soil metal concentrations can be reflected in the plants and in the chloroplasts. Metal deficiency or excess causes reduced growth and decreased productivity of crops. It can lead to human malnutrition. Several complex physiological processes can be responsible for the reduced biomass observed in plants with unbalanced metal concentration. In this review, we have focused on the structural and functional alterations of chloroplasts under metal deficiency or excess. Interestingly, besides specific differences, our data indicate several similarities in the response of chloroplasts to metal deficiency or excess. Indeed, oxidative stress and several ultrastructural alterations, e.g., increase in the number and size of plastoglobuli, disorganized grana and disturbed thylakoids, and swelling of the intrathylakoidal space, are observed in both cases. This indicates that changes in chloroplast ion homeostasis rather than the specific effect of a metal are responsible for decreased plant productivity. Therefore, sustainable agriculture has to take into consideration solutions that enable undisturbed metal and ion homeostasis in chloroplasts of crop plants grown even in soils with unbalanced metal concentrations.

Photosynthetic response of clusterbean chloroplasts to UV-B radiation: Energy imbalance and loss in redox homeostasis between Q A and Q B of photosystem II

The effects of ultraviolet-B (UV-B: 280–320 nm) radiation on the photosynthetic pigments, primary photochemical reactions of thylakoids and the rate of carbon assimilation ( P n) in the cotyledons of clusterbean ( Cyamopsis tetragonoloba) seedlings have been examined. The radiation induces an imbalance between the energy absorbed through the photophysical process of photosystem (PS) II and the energy consumed for carbon assimilation. Decline in the primary photochemistry of PS II induced by UV-B in the background of relatively stable P n, has been implicated in the creation of the energy imbalance . The radiation induced damage of PS II hinders the flow of electron from Q A to Q B resulting in a loss in the redox homeostasis between the Q A to Q B leading to an accumulation of Q A −. The accumulation of Q A − generates an excitation pressure that diminishes the PS II-mediated O 2 evolution, maximal photochemical potential ( F v/ F m) and PS II quantum yield ( Φ PS II). While UV-B radiation inactivates the carotenoid-mediated protective mechanisms, the accumulation of flavonoids seems to have a small role in protecting the photosynthetic apparatus from UV-B onslaught. The failure of protective mechanisms makes PS II further vulnerable to the radiation and facilitates the accumulation of malondialdehyde (MDA) indicating the involvement of reactive oxygen species (ROS) metabolism in UV-B-induced damage of photosynthetic apparatus of clusterbean cotyledons.

Factors altering the membrane fluidity of spinach thylakoid as determined by fluorescence polarization

Alterations in fluidity of thylakoid membranes isolated from spinach chloroplasts in response to sodium bisulfite (NaHSO3), hydrogen peroxide (H2O2), sodium dodecyl sulfate (SDS), bovine serum albumin (BSA), and free linoleic acid (LA) were investigated by means of a fluorescence polarization study with 1,6-diphenyl-1,3,5-hexatriene as the fluorescence probe. A decrease in fluidity and an increase in microviscosity of membrane were caused by NaHSO3 and H2O2 treatment. In contrast, SDS and BSA were found to increase thylakoid membranes fluidity and decrease microviscosity, in which the corresponding correlation coefficients were −0.9995 to −0.9516 (SDS) and −0.9359 (BSA), respectively. No changes in thylakoid membranes fluidity induced by free LA were found until its concentration above 5 mM where the polarization value (P value) declined (increased fluidity). The results suggest that the changes in thylakoids membrane fluidity might depend on the characteristics, mechanism and extent of the interactions between membrane components and compounds added.

Two interacting coiled-coil proteins, WEB1 and PMI2, maintain the chloroplast photorelocation movement velocity in Arabidopsis

Chloroplasts move toward weak light (accumulation response) and away from strong light (avoidance response). The fast and accurate movement of chloroplasts in response to ambient light conditions is essential for efficient photosynthesis and photodamage prevention in chloroplasts. Here, we report that two Arabidopsis mutants, weak chloroplast movement under blue light 1 (web1) and web2, are defective in both the avoidance and the accumulation responses. Map-based cloning revealed that both genes encode coiled-coil proteins and that WEB2 is identical to the plastid movement impaired 2 (PMI2) gene. The velocities of chloroplast movement in web1 and pmi2 were approximately threefold lower than that in the wild type. Defects in the avoidance response of web1 and pmi2 were suppressed by mutation of the J-domain protein required for chloroplast accumulation response 1 (JAC1) gene, which is essential for the accumulation response; these results indicate that WEB1 and PMI2 play a role in suppressing JAC1 under strong light conditions. A yeast two-hybrid analysis and a nuclear recruitment assay identified a physical interaction between WEB1 and PMI2, and transient expression analysis of CFP-WEB1 and YFP-PMI2 revealed that they colocalized in the cytosol. Bimolecular fluorescence complementation analysis confirmed the interaction of these proteins in the cytosol. Blue light-induced changes in short chloroplast actin filaments (cp-actin filaments) were impaired in both web1 and pmi2. Our findings suggest that a cytosolic WEB1-PMI2 complex maintains the velocity of chloroplast photorelocation movement via cp-actin filament regulation.

Differential Positioning of C4 Mesophyll and Bundle Sheath Chloroplasts: Aggregative Movement of C4 Mesophyll Chloroplasts in Response to Environmental Stresses

In C(4) plants, mesophyll (M) chloroplasts are randomly distributed along the cell walls, while bundle sheath (BS) chloroplasts are typically located in either a centripetal or centrifugal position. We investigated whether these intracellular positions are affected by environmental stresses. When mature leaves of finger millet (Eleusine coracana) were exposed to extremely high intensity light, most M chloroplasts aggregatively re-distributed to the BS side, whereas the intracellular arrangement of BS chloroplasts was unaffected. Compared with the homologous light-avoidance movement of M chloroplasts in C(3) plants, it requires extremely high light (3,000-4,000 micromol m(-2) s(-1)) and responds more slowly (distinctive movement observed in 1 h). The high light-induced movement of M chloroplasts was also observed in maize (Zea mays), another C(4) species, but with a distinct pattern of redistribution along the sides of anticlinal walls, analogous to C(3) plants. The aggregative movement of M chloroplasts occurred at normal light intensities (250-500 micromol m(-2) s(-1)) in response to environmental stresses, such as drought, salinity and hyperosmosis. Moreover, the re-arrangement of M chloroplasts was observed in field-grown C(4) plants when exposed to mid-day sunlight, but also under midsummer drought conditions. The migration of M chloroplasts was controlled by actin filaments and also induced in a light-dependent fashion upon incubation with ABA, which may be the physiological signal transducer. Together these results suggest that M and BS cells of C(4) plants have different mechanisms controlling intracellular chloroplast positioning, and that the aggregative movement of C(4) M chloroplasts is thought to be a protective response under environmental stress conditions.

In vivo reorganization of the actin cytoskeleton in leaves of Nicotiana tabacum L. transformed with plastin-GFP. Correlation with light-activated chloroplast responses.

BackgroundThe actin cytoskeleton is involved in the responses of plants to environmental signals. Actin bundles play the role of tracks in chloroplast movements activated by light. Chloroplasts redistribute in response to blue light in the mesophyll cells of Nicotiana tabacum. The aim of this work was to study the relationship between chloroplast responses and the organization of actin cytoskeleton in living tobacco cells. Chloroplast movements were measured photometrically as changes in light transmission through the leaves. The actin cytoskeleton, labeled with plastin-GFP, was visualised by confocal microscopy.ResultsThe actin cytoskeleton was affected by strong blue and red light. No blue light specific actin reorganization was detected. EGTA and trifluoperazine strongly inhibited chloroplast responses and disrupted the integrity of the cytoskeleton. This disruption was reversible by Ca2+ or Mg2+. Additionally, the effect of trifluoperazine was reversible by light. Wortmannin, an inhibitor of phosphoinositide kinases, potently inhibited chloroplast responses but did not influence the actin cytoskeleton at the same concentration. Also this inhibition was reversed by Ca2+ and Mg2+. Magnesium ions were equally or more effective than Ca2+ in restoring chloroplast motility after treatment with EGTA, trifluoperazine or wortmannin.ConclusionThe architecture of the actin cytoskeleton in the mesophyll of tobacco is significantly modulated by strong light. This modulation does not affect the direction of chloroplast redistribution in the cell. Calcium ions have multiple functions in the mechanism of the movements. Our results suggest also that Mg2+ is a regulatory molecule cooperating with Ca2+ in the signaling pathway of blue light-induced tobacco chloroplast movements.

Influences of magnesium deficiency and cerium on antioxidant system of spinach chloroplasts

Magnesium-deficiency conditions applied to spinach cultures caused an oxidative stress status in spinach chloroplast monitored by an increase in reactive oxygen species (ROS) accumulation. The enhancement of lipids peroxide of spinach chloroplast grown in magnesium-deficiency media suggested an oxidative attack that was activated by a reduction of antioxidative defense mechanism measured by analysing the activities of superoxide dismutase, catalase, ascorbate peroxidase, guaiacol peroxidase, and glutathione reductase, as well as antioxidants such as carotenoids and glutathione content. As the antioxidative response of chloroplast was reduced in spinach grown in magnesium-deficiency media, it caused a significant reduction of spinach plant weight, old leaves turning chlorosis. However, cerium treatment grown in magnesium-deficiency conditions decreased the malondialdehyde and ROS, and increased activities of the antioxidative defense system, and improved spinach growth. Together, the experimental study implied that cerium could partly substitute for magnesium and increase the oxidative stress-resistance of spinach chloroplast grown in magnesium-deficiency conditions, but the mechanisms need further study.

Bacterial Alarmone, Guanosine 5′‐Diphosphate 3′‐Diphosphate (ppGpp), Predominantly Binds the β′ Subunit of Plastid‐Encoded Plastid RNA Polymerase in Chloroplasts

It's alarming: Bacterial alarmone guanosine 5'-diphosphate 3'-diphosphate (ppGpp), which is a key regulatory molecule that controls the stringent response, also exists in chloroplasts of plant cells. Cross-linking experiments with 6-thioguanosine 5'-diphosphate 3'-diphosphate (6-thioppGpp) and chloroplast RNA polymerase indicate that ppGpp binds the beta' subunit of plastid-encoded plastid RNA polymerase that corresponds to the Escherichia coli beta' subunit. Chloroplasts, which are thought to have originated from cyanobacteria, have their own genetic system that is similar to that of the bacteria from which they were derived. Recently, bacterial alarmone guanosine 5'-diphosphate 3'-diphosphate (ppGpp, 1), a key regulatory molecule that controls the stringent response, was identified in the chloroplasts of plant cells. Similar to its function in bacteria, ppGpp inhibits chloroplast RNA polymerase; this suggests that ppGpp mediates gene expression through the stringent response in chloroplasts. However, a detailed mechanism of ppGpp action in chloroplasts remains elusive. We synthesized 6-thioguanosine 5'-diphosphate 3'-diphosphate (6-thioppGpp) as a photoaffinity probe of ppGpp; this probe thus enabled the investigation of ppGpp binding to chloroplast RNA polymerase. We found that 6-thioppGpp, as well as ppGpp, inhibits chloroplast RNA synthesis in vitro in a dose-dependent manner. Cross-linking experiments with 6-thioppGpp and chloroplast RNA polymerase indicated that ppGpp binds the beta' subunit (corresponding to the Escherichia coli beta' subunit) of plastid-encoded plastid RNA polymerase composed of alpha, beta, beta', beta'', and sigma subunits. Furthermore, ppGpp did not inhibit transcription in plastid nucleoids prepared from tobacco BY-2 cells; this suggests that ppGpp does not inhibit nuclear-encoded plastid RNA polymerase.

Light-Dependent Intracellular Positioning of Mitochondria in Arabidopsis thaliana Mesophyll Cells

Mitochondria, the power house of the cell, are one of the most dynamic cell organelles. Although there are several reports on actin- or microtubule-dependent movement of mitochondria in plant cells, intracellular positioning and motility of mitochondria under different light conditions remain open questions. Mitochondria were visualized in living Arabidopsis thaliana leaf cells using green fluorescent protein fused to a mitochondrion-targeting signal. In darkness, mitochondria were distributed randomly in palisade cells. In contrast, mitochondria accumulated along the periclinal walls, similar to the accumulation response of chloroplasts, when treated with weak blue light (470 nm, 4 micromol m(-2) s(-1)). Under strong blue light (100 micromol m(-2) s(-1)), mitochondria occupied the anticlinal positions similar to the avoidance response of chloroplasts and nuclei. While strong red light (660 nm, 100 micromol m(-2) s(-1)) induced the accumulation of mitochondria along the inner periclinal walls, green light exhibited little effect on the distribution of mitochondria. In addition, the mode of movement of individual mitochondria along the outer periclinal walls under different light conditions was precisely analyzed by time-lapse fluorescence microscopy. A gradual increase in the number of static mitochondria located in the vicinity of chloroplasts with a time period of blue light illumination clearly demonstrated the accumulation response of mitochondria. Light-induced co-localization of mitochondria with chloroplasts strongly suggested their mutual metabolic interactions. This is the first characterization of the light-dependent redistribution of mitochondria in plant cells.

Acquisition of plastid movement responsiveness to light during mesophyll cell differentiation

A culture of Nicotiana tabacum leaf protoplasts, regenerating in vitro, was used to study light-induced plastid translocations. Experiments were carried out for 5 months starting with protoplasts, through single cells, microcolonies and callus to the differentiated mesophyll of regenerated plants. Although the actin cytoskeleton was fully developed at every stage of culture, blue light-mediated directional movements of chloroplasts were observed only after the full differentiation of callus into leaf tissues. These chloroplast rearrangements were similar to those observed in control plants grown from seeds. Under strong blue light, chloroplasts gathered at the cell walls parallel to the light direction (profile position); under weak blue light, they gathered at the walls perpendicular to the light direction (face position). No light-dependent plastid arrangements were found in undifferentiated cell cultures even after cell wall recovery. A characteristic pattern of plastids in the dividing cells was independent of light signals. Only trace chloroplast responses to strong blue light were detected in the first leaves regenerating from callus. We hypothesize that factors which control the developmental status of a cell, i.e. division and differentiation, take over the control of plastid redistribution from light signals.

Low-temperature induced changes in the ultrastructure of maize mesophyll chloroplasts strongly depend on the chilling pattern/intensity and considerably differ among inbred and hybrid genotypes

The ultrastructure and dimensions of chloroplasts in leaf mesophyll cells were quantitatively examined in three parental inbred lines of maize (Zea mays L.) and their four hybrids subjected to two types of four-week low-temperature (LT) treatment: the abrupt onset of chilling temperatures (“severe chilling”, SC) and the gradual, more moderate one (“moderate chilling”, MC). The relationship between the response of individual genotypes to one or the other type of chilling was analyzed as well as the possibility to predict the behaviour of chloroplasts in hybrids from that of their parents. Although selected parameters of chloroplast ultrastructure (e.g. volume densities of granal and intergranal thylakoids, plastoglobuli, and peripheral reticulum) and dimensions changed due to the exposure of maize plants to LT, no general pattern of such changes was found for this species due to the observed intraspecific variability. The response of some genotype to SC could not be predicted from its behaviour under MC (and vice versa) and no clear rules could be applied for the inheritance of chloroplast response to chilling in the general sense. Thus, great caution should be always taken when interpreting the results of studies aimed at the dissection of chloroplast ultrastructure as affected by LT, particularly in case such studies are made with one genotype or under one type of chilling only.

The Bacterial Stringent Response, Conserved in Chloroplasts, Controls Plant Fertilization

The chloroplast, an essential organelle for plants, performs a wide variety of metabolic processes for host cells, which include photosynthesis as well as amino acid and fatty acid biosynthesis. The organelle conserves many bacterial systems in its functions, implicating its origin from symbiosis of a photosynthetic bacterium. In bacterial cells, the stringent response acts as a global regulatory system for gene expression mediated by a small nucleotide, guanosine 5'-diphosphate 3'-diphosphate (ppGpp), that is necessary for cell adaptation to diverse environmental stimuli such as amino acid starvation. Recent studies indicated that proteins similar to the bacterial ppGpp synthase/hydrolyase are conserved in plants, although their precise roles are not known. Here we show that the stringent response in chloroplasts is crucial for normal plant fertilization. Specifically, one of the Arabidopsis ppGpp synthase homologs, CRSH (Ca(2+)-activated RelA/SpoT homolog), exhibits calcium-dependent ppGpp synthesis activity in vitro, and is localized in chloroplasts in vivo. A knockdown mutation of CRSH in Arabidopsis results in a significant reduction in silique size and seed production, indicating that plant reproduction is under the control of chloroplast function through a ppGpp-mediated stringent response.