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1.  Control of yellow and purple nutsedge in elevated CO2 environments with glyphosate and halosulfuron 
Atmospheric concentrations of carbon dioxide (CO2) have significantly increased over the past century and are expected to continue rising in the future. While elevated levels of CO2 will likely result in higher crop yields, weed growth is also highly likely to increase, which could increase the incidence of herbicide resistant biotypes. An experiment was conducted in 2012 to determine the effects of an elevated CO2 environment on glyphosate and halosulfuron efficacy for postemergence control of purple and yellow nutsedge (Cyperus rotundus L. and C. esculentus L.). Both species of nutsedge where grown in 3.0-L containers under either ambient or elevated (ambient + 200 μmol mol−1) CO2 in open-top field chambers and treated with either 0.5×, 1.0×, or 1.5× of the manufacturer's labeled rate of halosulfuron, glyphosate, or a tank mix of the two herbicides. The growth of both nutsedge species responded positively to elevated CO2, purple nutsedge had increased shoot and root dry weights and yellow nutsedge had increased shoot, root, and tuber dry weights and counts. Few treatment differences were observed among the herbicides at any of the rates tested. At 3 weeks following herbicide application, both purple and yellow nutsedge were adequately controlled by both herbicides and combinations at all rates tested, regardless of CO2 concentration. Based on this study, it is likely that predicted future CO2 levels will have little impact on the efficacy of single applications of halosulfuron or glyphosate for control of purple and yellow nutsedge at the growth stages described here, although scenarios demanding more persistent control efforts remain a question.
PMCID: PMC4299438  PMID: 25653664
climate change; invasive species; herbicide; nutsedge; resistance management
2.  The elucidation of stress memory inheritance in Brassica rapa plants 
Plants are able to maintain the memory of stress exposure throughout their ontogenesis and faithfully propagate it into the next generation. Recent evidence argues for the epigenetic nature of this phenomenon. Small RNAs (smRNAs) are one of the vital epigenetic factors because they can both affect gene expression at the place of their generation and maintain non-cell-autonomous gene regulation. Here, we have made an attempt to decipher the contribution of smRNAs to the heat-shock-induced transgenerational inheritance in Brassica rapa plants using sequencing technology. To do this, we have generated comprehensive profiles of a transcriptome and a small RNAome (smRNAome) from somatic and reproductive tissues of stressed plants and their untreated progeny. We have demonstrated that the highest tissue-specific alterations in the transcriptome and smRNAome profile are detected in tissues that were not directly exposed to stress, namely, in the endosperm and pollen. Importantly, we have revealed that the progeny of stressed plants exhibit the highest fluctuations at the smRNAome level but not at the transcriptome level. Additionally, we have uncovered the existence of heat-inducible and transgenerationally transmitted tRNA-derived small RNA fragments in plants. Finally, we suggest that miR168 and braAGO1 are involved in the stress-induced transgenerational inheritance in plants.
PMCID: PMC4300914  PMID: 25653665
epigenetics; small RNAs; transcriptome; B. rapa; transgenerational inheritance
3.  A multi-tissue genome-scale metabolic modeling framework for the analysis of whole plant systems 
Genome scale metabolic modeling has traditionally been used to explore metabolism of individual cells or tissues. In higher organisms, the metabolism of individual tissues and organs is coordinated for the overall growth and well-being of the organism. Understanding the dependencies and rationale for multicellular metabolism is far from trivial. Here, we have advanced the use of AraGEM (a genome-scale reconstruction of Arabidopsis metabolism) in a multi-tissue context to understand how plants grow utilizing their leaf, stem and root systems across the day-night (diurnal) cycle. Six tissue compartments were created, each with their own distinct set of metabolic capabilities, and hence a reliance on other compartments for support. We used the multi-tissue framework to explore differences in the “division-of-labor” between the sources and sink tissues in response to: (a) the energy demand for the translocation of C and N species in between tissues; and (b) the use of two distinct nitrogen sources (NO−3 or NH+4). The “division-of-labor” between compartments was investigated using a minimum energy (photon) objective function. Random sampling of the solution space was used to explore the flux distributions under different scenarios as well as to identify highly coupled reaction sets in different tissues and organelles. Efficient identification of these sets was achieved by casting this problem as a maximum clique enumeration problem. The framework also enabled assessing the impact of energetic constraints in resource (redox and ATP) allocation between leaf, stem, and root tissues required for efficient carbon and nitrogen assimilation, including the diurnal cycle constraint forcing the plant to set aside resources during the day and defer metabolic processes that are more efficiently performed at night. This study is a first step toward autonomous modeling of whole plant metabolism.
PMCID: PMC4302846  PMID: 25657653
multi-tissue; genome-scale; modeling; plant metabolism; AraGEM
4.  Overexpression of copper/zinc superoxide dismutase from mangrove Kandelia candel in tobacco enhances salinity tolerance by the reduction of reactive oxygen species in chloroplast 
Na+ uptake and transport in Kandelia candel and antioxidative defense were investigated under rising NaCl stress from 100 to 300 mM. Salinized K. candel roots had a net Na+ efflux with a declined flux rate during an extended NaCl exposure. Na+ buildup in leaves enhanced H2O2 levels, superoxide dismutase (SOD) activity, and increased transcription of CSD gene encoding a Cu/Zn SOD. Sequence and subcellular localization analyses have revealed that KcCSD is a typical Cu/Zn SOD in chloroplast. The transgenic tobacco experimental system was used as a functional genetics model to test the effect of KcCSD on salinity tolerance. KcCSD-transgenic lines were more Na+ tolerant than wild-type (WT) tobacco in terms of lipid peroxidation, root growth, and survival rate. In the latter, 100 mM NaCl led to a remarkable reduction in chlorophyll content and a/b ratio, decreased maximal chlorophyll a fluorescence, and photochemical efficiency of photosystem II. NaCl stress in WT resulted from H2O2 burst in chloroplast. Na+ injury to chloroplast was less pronounced in KcCSD-transgenic plants due to upregulated antioxidant defense. KcCSD-transgenic tobacco enhanced SOD activity by an increment in SOD isoenzymes under 100 mM NaCl stress from 24 h to 7 day. Catalase activity rose in KcCSD overexpressing tobacco plants. KcCSD-transgenic plants better scavenged NaCl-elicited reactive oxygen species (ROS) compared to WT ones. In conclusion, K. candel effectively excluded Na+ in roots during a short exposure; and increased CSD expression to reduce ROS in chloroplast in a long-term and high saline environment.
PMCID: PMC4302849  PMID: 25657655
Kandelia candel; Na+ flux; superoxide anion; hydrogen peroxide; salt; catalase; superoxide dismutase
5.  Disintegration of microtubules in Arabidopsis thaliana and bladder cancer cells by isothiocyanates 
Isothiocyanates (ITCs) from biodegradation of glucosinolates comprise a group of electrophiles associated with growth-inhibitory effects in plant- and mammalian cells. The underlying modes of action of this feature are not fully understood. Clarifying this has involved mammalian cancer cells due to ITCs' chemopreventive potential. The binding of ITCs to tubulins has been reported as a mechanism by which ITCs induce cell cycle arrest and apoptosis. In the present study we demonstrate that ITCs disrupt microtubules in Arabidopsis thaliana contributing to the observed inhibited growth phenotype. We also confirmed this in rat bladder cancer cells (AY-27) suggesting that cells from plant and animals share mechanisms by which ITCs affect growth. Exposure of A. thaliana to vapor-phase of allyl ITC (AITC) inhibited growth and induced a concurrent bleaching of leaves in a dose-dependent manner. Transcriptional analysis was used to show an upregulation of heat shock-genes upon AITC-treatment. Transgenic A. thaliana expressing GFP-marked α-tubulin was employed to show a time- and dose-dependent disintegration of microtubules by AITC. Treatment of AY-27 with ITCs resulted in a time- and dose-dependent decrease of cell proliferation and G2/M-arrest. AY-27 transiently transfected to express GFP-tagged α-tubulin were treated with ITCs resulting in a loss of microtubular filaments and the subsequent formation of apoptotic bodies. In conclusion, our data demonstrate an ITC-induced mechanism leading to growth inhibition in A. thaliana and rat bladder cancer cells, and expose clues to the mechanisms underlying the physiological role of glucosinolates in vivo.
PMCID: PMC4303138  PMID: 25657654
isothiocyanates; plant defense; Arabidopsis thaliana; bladder cancer; glucosinolate-myrosinase pathway
6.  Perception of Arabidopsis AtPep peptides, but not bacterial elicitors, accelerates starvation-induced senescence 
Members of the AtPep group of Arabidopsis endogenous peptides have frequently been reported to induce pattern-triggered immunity (PTI) and to increase resistance to diverse pathogens by amplifying the innate immune response. Here, we made the surprising observation that dark-induced leaf senescence was accelerated by the presence of Peps. Adult leaves as well as leaf discs of Col-0 wild type plants showed a Pep-triggered early onset of chlorophyll breakdown and leaf yellowing whereas pepr1 pepr2 double mutant plants were insensitive. In addition, this response was dependent on ethylene signaling and inhibited by the addition of cytokinins. Notably, addition of the bacterial elicitors flg22 or elf18, both potent inducers of PTI, did not provoke an early onset of leaf senescence. Continuous darkness leads to energy deprivation and starvation and therewith promotes leaf senescence. We found that continuous darkness also strongly induced PROPEP3 transcription. Moreover, Pep-perception led to a rapid induction of PAO, APG7, and APG8a, genes indispensable for chlorophyll degradation as well as autophagy, respectively, and all three hallmarks of starvation and senescence. Notably, addition of sucrose as a source of energy inhibited the Pep-triggered early onset of senescence. In conclusion, we report that Pep-perception accelerates dark/starvation-induced senescence via an early induction of chlorophyll degradation and autophagy. This represents a novel and unique characteristic of PEPR signaling, unrelated to PTI.
PMCID: PMC4304247  PMID: 25667591
Plant elicitor peptide (Pep); starvation; senescence; MAMP; Arabidopsis; PEPR; PTI
7.  The concept of the sexual reproduction cycle and its evolutionary significance 
The concept of a “sexual reproduction cycle (SRC)” was first proposed by Bai and Xu (2013) to describe the integration of meiosis, sex differentiation, and fertilization. This review discusses the evolutionary and scientific implications of considering these three events as part of a single process. Viewed in this way, the SRC is revealed to be a mechanism for efficiently increasing genetic variation, facilitating adaptation to environmental challenges. It also becomes clear that, in terms of cell proliferation, it is appropriate to contrast mitosis with the entire SRC, rather than with meiosis alone. Evolutionarily, it appears that the SRC was first established in unicellular eukaryotes and that all multicellular organisms evolved within that framework. This concept provides a new perspective into how sexual reproduction evolved, how generations should be defined, and how developmental processes of various multicellular organisms should properly be compared.
PMCID: PMC4304253  PMID: 25667590
sexual reproduction cycle; meiosis; heterogametogenesis; fertilization; generation
8.  Regulation of gametogenesis and zoosporogenesis in Ulva linza (Chlorophyta): comparison with Ulva mutabilis and potential for laboratory culture 
Green Ulvophyte macroalgae represent attractive model systems for understanding growth, development, and evolution. They are untapped resources for food, fuel, and high-value compounds, but can also form nuisance blooms. To fully analyze green seaweed morphogenesis, controlled laboratory-based culture of these organisms is required. To date, only a single Ulvophyte species, Ulva mutabilis Føyn, has been manipulated to complete its whole life cycle in laboratory culture and to grow continuously under axenic conditions. Such cultures are essential to address multiple key questions in Ulva development and in algal–bacterial interactions. Here we show that another Ulva species, U. linza, with a broad geographical distribution, has the potential to be grown in axenic culture similarly to U. mutabilis. U. linza can be reliably induced to sporulate (form gametes and zoospores) in the laboratory, by cutting the relevant thallus tissue into small pieces and removing extracellular inhibitors (sporulation and swarming inhibitors). The germ cells work as an ideal feed stock for standardized algae cultures. The requirement of U. linza for bacterial signals to induce its normal morphology (particularly of the rhizoids) appears to have a species-specific component. The axenic cultures of these two species pave the way for future comparative studies of algal–microbial interactions.
PMCID: PMC4306291
green algae; gametogenesis; zoosporogenesis; morphogenesis; life cycle; algal–bacterial interactions; axenic culture; sporulation inhibitor
9.  GIGANTEA – an emerging story 
GIGANTEA (GI) is a plant specific nuclear protein and functions in diverse physiological processes such as flowering time regulation, light signaling, hypocotyl elongation, control of circadian rhythm, sucrose signaling, starch accumulation, chlorophyll accumulation, transpiration, herbicide tolerance, cold tolerance, drought tolerance, and miRNA processing. It has been five decades since its discovery but the biochemical function of GI and its different domains are still unclear. Although it is known that both GI transcript and GI protein are clock controlled, the regulation of its abundance and functions at the molecular level are still some of the unexplored areas of intensive research. Since GI has many important pleotropic functions as described above scattered through literature, it is worthwhile and about time to encapsulate the available information in a concise review. Therefore, in this review, we are making an attempt to summarize (i) the various interconnected roles that GI possibly plays in the fine-tuning of plant development, and (ii) the known mutations of GI that have been instrumental in understanding its role in distinct physiological processes.
PMCID: PMC4306306
GIGANTEA; flowering time regulation; circadian clock control; GI mutants; light signaling
10.  Optimization of ATP synthase function in mitochondria and chloroplasts via the adenylate kinase equilibrium 
The bulk of ATP synthesis in plants is performed by ATP synthase, the main bioenergetics engine of cells, operating both in mitochondria and in chloroplasts. The reaction mechanism of ATP synthase has been studied in detail for over half a century; however, its optimal performance depends also on the steady delivery of ATP synthase substrates and the removal of its products. For mitochondrial ATP synthase, we analyze here the provision of stable conditions for (i) the supply of ADP and Mg2+, supported by adenylate kinase (AK) equilibrium in the intermembrane space, (ii) the supply of phosphate via membrane transporter in symport with H+, and (iii) the conditions of outflow of ATP by adenylate transporter carrying out the exchange of free adenylates. We also show that, in chloroplasts, AK equilibrates adenylates and governs Mg2+ contents in the stroma, optimizing ATP synthase and Calvin cycle operation, and affecting the import of inorganic phosphate in exchange with triose phosphates. It is argued that chemiosmosis is not the sole component of ATP synthase performance, which also depends on AK-mediated equilibrium of adenylates and Mg2+, adenylate transport, and phosphate release and supply.
PMCID: PMC4309032
adenylate kinase; ATP synthase; chemiosmosis; chloroplasts; magnesium; mitochondria
11.  Exploring the importance of sulfate transporters and ATP sulphurylases for selenium hyperaccumulation—a comparison of Stanleya pinnata and Brassica juncea (Brassicaceae) 
Selenium (Se) hyperaccumulation, the capacity of some species to concentrate Se to levels upwards of 0.1% of dry weight, is an intriguing phenomenon that is only partially understood. Questions that remain to be answered are: do hyperaccumulators have one or more Se-specific transporters? How are these regulated by Se and sulfur (S)? In this study, hyperaccumulator Stanleya pinnata was compared with related non-hyperaccumulator Brassica juncea with respect to S-dependent selenate uptake and translocation, as well as for the expression levels of three sulfate/selenate transporters (Sultr) and three ATP sulphurylases (APS). Selenium accumulation went down ~10-fold with increasing sulfate supply in B. juncea, while S. pinnata only had a 2–3-fold difference in Se uptake between the highest (5 mM) and lowest sulfate (0 mM) treatments. The Se/S ratio was generally higher in the hyperaccumulator than the non-hyperaccumulator, and while tissue Se/S ratio in B. juncea largely reflected the ratio in the growth medium, S. pinnata enriched itself up to 5-fold with Se relative to S. The transcript levels of Sultr1;2 and 2;1 and APS1, 2, and 4 were generally much higher in S. pinnata than B. juncea, and the species showed differential transcript responses to S and Se supply. These results indicate that S. pinnata has at least one transporter with significant selenate specificity over sulfate. Also, the hyperaccumulator has elevated expression levels of several sulfate/selenate transporters and APS enzymes, which likely contribute to the Se hyperaccumulation and hypertolerance phenotype.
PMCID: PMC4304243
Brassica juncea; Stanleya pinnata; selenium; sulfur; uptake; ATP-sulphurylase; gene expression
12.  Facile high-throughput forward chemical genetic screening by in situ monitoring of glucuronidase-based reporter gene expression in Arabidopsis thaliana 
The use of biologically active small molecules to perturb biological functions holds enormous potential for investigating complex signaling networks. However, in contrast to animal systems, the search for and application of chemical tools for basic discovery in the plant sciences, generally referred to as “chemical genetics,” has only recently gained momentum. In addition to cultured cells, the well-characterized, small-sized model plant Arabidopsis thaliana is suitable for cultivation in microplates, which allows employing diverse cell- or phenotype-based chemical screens. In such screens, a chemical's bioactivity is typically assessed either through scoring its impact on morphological traits or quantifying molecular attributes such as enzyme or reporter activities. Here, we describe a facile forward chemical screening methodology for intact Arabidopsis seedlings harboring the β-glucuronidase (GUS) reporter by directly quantifying GUS activity in situ with 4-methylumbelliferyl-β-D-glucuronide (4-MUG) as substrate. The quantitative nature of this screening assay has an obvious advantage over the also convenient histochemical GUS staining method, as it allows application of statistical procedures and unbiased hit selection based on threshold values as well as distinction between compounds with strong or weak bioactivity. At the same time, the in situ bioassay is very convenient requiring less effort and time for sample handling in comparison to the conventional quantitative in vitro GUS assay using 4-MUG, as validated with several Arabidopsis lines harboring different GUS reporter constructs. To demonstrate that the developed assays is particularly suitable for large-scale screening projects, we performed a pilot screen for chemical activators or inhibitors of salicylic acid-mediated defense signaling using the Arabidopsis PR1p::GUS line. Importantly, the screening methodology provided here can be adopted for any inducible GUS reporter line.
PMCID: PMC4310277
chemical screening; chemical genetics; high-throughput screening; bioactive small molecules; β-glucuronidase activity; reporter gene expression; salicylic acid
13.  When plants produce not enough or at all: metabolic engineering of flavonoids in microbial hosts 
As a result of the discovery that flavonoids are directly or indirectly connected to health, flavonoid metabolism and its fascinating molecules that are natural products in plants, have attracted the attention of both the industry and researchers involved in plant science, nutrition, bio/chemistry, chemical bioengineering, pharmacy, medicine, etc. Subsequently, in the past few years, flavonoids became a top story in the pharmaceutical industry, which is continually seeking novel ways to produce safe and efficient drugs. Microbial cell cultures can act as workhorse bio-factories by offering their metabolic machinery for the purpose of optimizing the conditions and increasing the productivity of a selective flavonoid. Furthermore, metabolic engineering methodology is used to reinforce what nature does best by correcting the inadequacies and dead-ends of a metabolic pathway. Combinatorial biosynthesis techniques led to the discovery of novel ways of producing natural and even unnatural plant flavonoids, while, in addition, metabolic engineering provided the industry with the opportunity to invest in synthetic biology in order to overcome the currently existing restricted diversification and productivity issues in synthetic chemistry protocols. In this review, is presented an update on the rationalized approaches to the production of natural or unnatural flavonoids through biotechnology, analyzing the significance of combinatorial biosynthesis of agricultural/pharmaceutical compounds produced in heterologous organisms. Also mentioned are strategies and achievements that have so far thrived in the area of synthetic biology, with an emphasis on metabolic engineering targeting the cellular optimization of microorganisms and plants that produce flavonoids, while stressing the advances in flux dynamic control and optimization. Finally, the involvement of the rapidly increasing numbers of assembled genomes that contribute to the gene- or pathway-mining in order to identify the gene(s) responsible for producing species-specific secondary metabolites is also considered herein.
PMCID: PMC4310283
flavonoid biosynthesis; unnatural flavonoids; metabolic engineering; dynamic regulation; metabolic control; secondary metabolites; combinatorial biosynthesis
14.  To grow old: regulatory role of ethylene and jasmonic acid in senescence 
Senescence, the final stage in the development of an organ or whole plant, is a genetically programmed process controlled by developmental and environmental signals. Age-related signals underlie the onset of senescence in specific organs (leaf, flower, and fruit) as well as the whole plant (monocarpic senescence). Rudimentary to most senescence processes is the plant hormone ethylene, a small gaseous molecule critical to diverse processes throughout the life of the plant. The role of ethylene in senescence was discovered almost 100 years ago, but the molecular mechanisms by which ethylene regulates senescence have been deciphered more recently primarily through genetic and molecular studies in Arabidopsis. Jasmonic acid (JA), another plant hormone, is emerging as a key player in the control of senescence. The regulatory network of ethylene and JA involves the integration of transcription factors, microRNAs, and other hormones. In this review, we summarize the current understanding of ethylene’s role in senescence, and discuss the interplay of ethylene with JA in the regulation of senescence.
PMCID: PMC4310285
ethylene; jasmonic acid; leaf senescence; cross-talk; miRNA; transcription factors
15.  Development and Characterization of Somatic Hybrids of Ulva reticulata Forsskål (×) Monostroma oxyspermum (Kutz.)Doty 
Ulvophycean species with diverse trait characteristics provide an opportunity to create novel allelic recombinant variants. The present study reports the development of seaweed variants with improved agronomic traits through protoplast fusion between Monostroma oxyspermum (Kutz.) Doty and Ulva reticulata Forsskål. A total of 12 putative hybrids were screened based on the variations in morphology and total DNA content over the fusion partners. DNA-fingerprinting by inter simple sequence repeat (ISSR) and amplified fragment length polymorphism (AFLP) analysis confirmed genomic introgression in the hybrids. The DNA fingerprint revealed sharing of parental alleles in regenerated hybrids and a few alleles that were unique to hybrids. The epigenetic variations in hybrids estimated in terms of DNA methylation polymorphism also revealed sharing of methylation loci with both the fusion partners. The functional trait analysis for growth showed a hybrid with heterotic trait (DGR% = 36.7 ± 1.55%) over the fusion partners U. reticulata (33.2 ± 2.6%) and M. oxyspermum (17.8 ± 1.77%), while others were superior to the mid-parental value (25.2 ± 2.2%) (p < 0.05). The fatty acid (FA) analysis of hybrids showed notable variations over fusion partners. Most hybrids showed increased polyunsaturated FAs (PUFAs) compared to saturated FAs (SFAs) and mainly includes the nutritionally important linoleic acid, α-linolenic acid, oleic acid, stearidonic acid, and docosahexaenoic acid. The other differences observed include superior cellulose content and antioxidative potential in hybrids over fusion partners. The hybrid varieties with superior traits developed in this study unequivocally demonstrate the significance of protoplast fusion technique in developing improved varients of macroalgae.
PMCID: PMC4310296
heterosis; hybrid; Monostroma; protoplast; Ulva
16.  Toward a more physiologically and evolutionarily relevant definition of metal hyperaccumulation in plants 
PMCID: PMC4311607
hyperaccumulation; tolerance; metals; metalloids; physiology; evolution; traits
17.  To die or not to die? Lessons from lesion mimic mutants 
Programmed cell death (PCD) is a ubiquitous genetically regulated process consisting in an activation of finely controlled signaling pathways that lead to cellular suicide. Although some aspects of PCD control appear evolutionary conserved between plants, animals and fungi, the extent of conservation remains controversial. Over the last decades, identification and characterization of several lesion mimic mutants (LMM) has been a powerful tool in the quest to unravel PCD pathways in plants. Thanks to progress in molecular genetics, mutations causing the phenotype of a large number of LMM and their related suppressors were mapped, and the identification of the mutated genes shed light on major pathways in the onset of plant PCD such as (i) the involvements of chloroplasts and light energy, (ii) the roles of sphingolipids and fatty acids, (iii) a signal perception at the plasma membrane that requires efficient membrane trafficking, (iv) secondary messengers such as ion fluxes and ROS and (v) the control of gene expression as the last integrator of the signaling pathways.
PMCID: PMC4311611
plant; programmed cell death; lesion mimic mutants; genetics approaches; immunity responses
18.  The absence of protein Y4yS affects negatively the abundance of T3SS Mesorhizobium loti secretin, RhcC2, in bacterial membranes 
Mesorhizobium loti MAFF303099 has a functional type III secretion system (T3SS) that is involved in the determination of nodulation competitiveness on Lotus. The M. loti T3SS cluster contains gene y4yS (mlr8765) that codes for a protein of unknown function (Y4yS). A mutation in the y4yS gene favors the M. loti symbiotic competitive ability on Lotus tenuis cv. Esmeralda and affects negatively the secretion of proteins through T3SS. Here we localize Y4yS in the bacterial membrane using a translational reporter peptide fusion. In silico analysis indicated that this protein presents a tetratricopeptide repeat (TPR) domain, a signal peptide and a canonical lipobox LGCC in the N-terminal sequence. These features that are shared with proteins required for the formation of the secretin complex in type IV secretion systems and in the Tad system, together with its localization, suggest that the y4yS-encoded protein is required for the formation of the M. loti T3SS secretin (RhcC2) complex. Remarkably, analysis of RhcC2 in the wild-type and M. loti y4yS mutant strains indicated that the absence of Y4yS affects negatively the accumulation of normal levels of RhcC2 in the membrane.
PMCID: PMC4311626
symbiosis; rhizobium; rhizobia; secretion system; secretin; pilotin
19.  Time to dig deep into the plant proteome: a hunt for low-abundance proteins 
PMCID: PMC4311630
low-abundance proteins; high-abundance proteins; two-dimensional gel electrophoresis; RuBisCO; post-translational modifications
20.  Photopolarization of Fucus zygotes is determined by time sensitive vectorial addition of environmental cues during axis amplification 
Fucoid zygotes have been extensively used to study cell polarization and asymmetrical cell division. Fertilized eggs are responsive to different environmental cues (e.g., light, gravity) for a long period before the polarity is fixed and the cells germinate accordingly. First, it is commonly believed that the direction and sense of the polarization vector are established simultaneously as indicated by the formation of an F-actin patch. Secondly, upon reorientation of the zygote, a new polar gradient is formed and it is assumed that the position of the future rhizoid pole is only influenced by the latter. Here we tested these two hypotheses investigating photopolarization in Fucus zygotes by reorienting zygotes 90° relative to a unilateral light source at different time points during the first cell cycle. We conclude that fixation of direction and sense of the polarization vector is indeed established simultaneously. However, the experiments yielded a distribution of polarization axes that cannot be explained if only the last environmental cue is supposed to determine the polarization axis. We conclude that our observations, together with published findings, can only be explained by assuming imprinting of the different polarization vectors and their integration as a vectorial sum at the moment of axis fixation. This way cells will average different serially perceived cues resulting in a polarization vector representative of the dynamic intertidal environment, instead of betting exclusively on the perceived vector at the moment of axis fixation.
PMCID: PMC4315017
Fucus; polarization; asymmetrical cell division; positional information; Brown algae; intrinsic factors; embryogenesis; patterning
21.  OsSDS is essential for DSB formation in rice meiosis 
SDS is a meiosis specific cyclin-like protein and required for DMC1 mediated double-strand break (DSB) repairing in Arabidopsis. Here, we found its rice homolog, OsSDS, is essential for meiotic DSB formation. The Ossds mutant is normal in vegetative growth but both male and female gametes are inviable. The Ossds meiocytes exhibit severe defects in homologous pairing and synapsis. No γH2AX immunosignals in Ossds meiocytes together with the suppression of chromosome fragmentation in Ossds-1 Osrad51c, both provide strong evidences that OsSDS is essential for meiotic DSB formation. Immunostaining investigations revealed that meiotic chromosome axes are normally formed but both SC installation and localization of recombination elements are failed in Ossds. We suspected that this cyclin protein has been differentiated pretty much between monocots and dicots on its function in meiosis.
PMCID: PMC4315026
rice; OsSDS; meiosis; DSB formation
22.  Lineage-specific evolution of Methylthioalkylmalate synthases (MAMs) involved in glucosinolates biosynthesis 
Methylthioalkylmalate synthases (MAMs) encoded by MAM genes are central to the diversification of the glucosinolates, which are important secondary metabolites in Brassicaceae species. However, the evolutionary pathway of MAM genes is poorly understood. We analyzed the phylogenetic and synteny relationships of MAM genes from 13 sequenced Brassicaceae species. Based on these analyses, we propose that the syntenic loci of MAM genes, which underwent frequent tandem duplications, divided into two independent lineage-specific evolution routes and were driven by positive selection after the divergence from Aethionema arabicum. In the lineage I species Capsella rubella, Camelina sativa, Arabidopsis lyrata, and A. thaliana, the MAM loci evolved three tandem genes encoding enzymes responsible for the biosynthesis of aliphatic glucosinolates with different carbon chain-lengths. In lineage II species, the MAM loci encode enzymes responsible for the biosynthesis of short-chain aliphatic glucosinolates. Our proposed model of the evolutionary pathway of MAM genes will be useful for understanding the specific function of these genes in Brassicaceae species.
PMCID: PMC4315028
glucosinolates; MAM genes; syntenic; evolution; Brassicaceae
23.  Regulation of floral stem cell termination in Arabidopsis 
In Arabidopsis, floral stem cells are maintained only at the initial stages of flower development, and they are terminated at a specific time to ensure proper development of the reproductive organs. Floral stem cell termination is a dynamic and multi-step process involving many transcription factors, chromatin remodeling factors and signaling pathways. In this review, we discuss the mechanisms involved in floral stem cell maintenance and termination, highlighting the interplay between transcriptional regulation and epigenetic machinery in the control of specific floral developmental genes. In addition, we discuss additional factors involved in floral stem cell regulation, with the goal of untangling the complexity of the floral stem cell regulatory network.
PMCID: PMC4313600
Arabidopsis; floral meristem; stem cell; determinacy; flower development
24.  Brassica napus L. cultivars show a broad variability in their morphology, physiology and metabolite levels in response to sulfur limitations and to pathogen attack 
Under adequate sulfur supply, plants accumulate sulfate in the vacuoles and use sulfur-containing metabolites as storage compounds. Under sulfur-limiting conditions, these pools of stored sulfur-compounds are depleted in order to balance the nitrogen to sulfur ratio for protein synthesis. Stress conditions like sulfur limitation and/or pathogen attack induce changes in the sulfate pool and the levels of sulfur-containing metabolites, which often depend on the ecotypes or cultivars. We are interested in investigating the influence of the genetic background of canola (Brassica napus) cultivars in sulfur-limiting conditions on the resistance against Verticillium longisporum. Therefore, four commercially available B. napus cultivars were analyzed. These high-performing cultivars differ in some characteristics described in their cultivar pass, such as several agronomic traits, differences in the size of the root system, and resistance to certain pathogens, such as Phoma and Verticillium. The objectives of the study were to examine and explore the patterns of morphological, physiological and metabolic diversity in these B. napus cultivars at different sulfur concentrations and in the context of plant defense. Results indicate that the root systems are influenced differently by sulfur deficiency in the cultivars. Total root dry mass and length of root hairs differ not only among the cultivars but also vary in their reaction to sulfur limitation and pathogen attack. As a sensitive indicator of stress, several parameters of photosynthetic activity determined by PAM imaging showed a broad variability among the treatments. These results were supported by thermographic analysis. Levels of sulfur-containing metabolites also showed large variations. The data were interrelated to predict the specific behavior during sulfur limitation and/or pathogen attack. Advice for farming are discussed.
PMCID: PMC4313603
canola; diurnal rhythm; elemental sulfur; metabolites; Verticillium longisporum
25.  Key players of singlet oxygen-induced cell death in plants 
The production of reactive oxygen species (ROS) is an unavoidable consequence of oxygenic photosynthesis. Singlet oxygen (1O2) is a highly reactive species to which has been attributed a major destructive role during the execution of ROS-induced cell death in photosynthetic tissues exposed to excess light. The study of the specific biological activity of 1O2 in plants has been hindered by its high reactivity and short lifetime, the concurrent production of other ROS under photooxidative stress, and limited in vivo detection methods. However, during the last 15 years, the isolation and characterization of two 1O2-overproducing mutants in Arabidopsis thaliana, flu and ch1, has allowed the identification of genetically controlled 1O2 cell death pathways and a 1O2 acclimation pathway that are triggered at sub-cytotoxic concentrations of 1O2. The study of flu has revealed the control of cell death by the plastid proteins EXECUTER (EX)1 and EX2. In ch1, oxidized derivatives of β-carotene, such as β-cyclocitral and dihydroactinidiolide, have been identified as important upstream messengers in the 1O2 signaling pathway that leads to stress acclimation. In both the flu and ch1 mutants, phytohormones act as important promoters or inhibitors of cell death. In particular, jasmonate has emerged as a key player in the decision between acclimation and cell death in response to 1O2. Although the flu and ch1 mutants show many similarities, especially regarding their gene expression profiles, key differences, such as EXECUTER-independent cell death in ch1, have also been observed and will need further investigation to be fully understood.
PMCID: PMC4316694
singlet oxygen; oxidative stress; cell death; acclimation; EXECUTER; β-cyclocitral; phytohormones; oxylipins

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