SciCombinator

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Journal: Physiologia plantarum

28

Galactinol synthase (GolS, EC 2.4.1.123) catalyzes formation of galactinol and the subsequent synthesis of raffinose family oligosaccharides (RFOs). The relationship of GolS to drought and salt tolerance has been well documented, however, little information is available about the role of GolS gene in cold tolerance. A coding sequence of MfGolS1 cDNA was cloned from Medicago sativa subsp. falcata (i.e. M. falcata), a species that exhibits greater cold tolerance than alfalfa (Medicago sativa). MfGolS1 transcript was not detected in untreated vegetative tissues using RNA blot hybridization; however, it was greatly induced in leaves, but not in stem and petiole, after cold treatment. Higher levels of MfGolS1 transcript was induced and maintained in M. falcata than in M. sativa during cold acclimation. Accordingly, more sugars including sucrose, galactinol, raffinose and stachyose were accumulated in M. falcata than in M. sativa. The data indicated that MfGolS1 transcript and its resultant sugar accumulation were associated with the differential cold tolerance between M. falcata and M. sativa. MfGolS1 transcript was weakly induced by dehydration and salt stresses, but not responsive to abscisic acid (ABA). MfGolS1 could be induced by myo-inositol, which is proposed to participate in cold-induced MfGolS1 expression. Overexpression of MfGolS1 in tobacco resulted in elevated tolerance to freezing and chilling in transgenic plants as a result of enhanced levels of galactinol, raffinose, and stachyose. Tolerance to drought and salt stresses was also increased in the transgenic tobacco plants. It is suggested that MfGolS1 plays an important role in plant tolerance to abiotic stresses.

Concepts: DNA, Gene, Gene expression, Molecular biology, Seed, Alfalfa, Medicago, Result

27

Somatic embryogenesis (SE) represents a useful experimental system for studying the regulatory mechanisms of embryo development. In this study, the effect of carbohydrates and osmoticum on storage reserve accumulation and germination of Norway spruce (Picea abies (L.) Karst) somatic embryos were investigated. Using time lapse photography, we monitored development from proliferation of proembryogenic masses (PEMs) to maturation of somatic embryos in two P. abies cell lines cultured on two maturation treatments. A combination of sugar assays, metabolic and proteomic analyses were used to quantify storage reserves in the mature somatic embryos. The maturation treatment containing a non-permeating osmoticum, polyethylene glycol (PEG, 7.5%) and maltose (3%) as the carbohydrate gave significantly high maturation and low germination frequencies of somatic embryos compared to the treatment with only 3% sucrose. Somatic embryos treated with 3% sucrose contained high levels of sucrose, raffinose and late embryogenesis abundant (LEA) proteins. These compounds are known to be involved in the acquisition of desiccation tolerance during seed development and maturation. In addition the sucrose treatment significantly increased the content of starch in the somatic embryos while the maltose and PEG treatment resulted in somatic embryos with a high content of storage proteins. The high levels of sucrose, raffinose and LEA proteins in the embryos treated with 3% sucrose suggest that sucrose may improve the germination of somatic embryos by promoting the acquisition of desiccation tolerance.

Concepts: Embryo, Glucose, Carbohydrate, Spruce, Christmas tree, Norway Spruce, Picea, Plant embryogenesis

27

As the key enzyme in the biosynthesis of blue flower color pigments, flavonoid 3',5'-hydroxylase (F3'5'H) can catalyze the conversion of its major substrates, 2-S naringenin (NAR) and dihydrokaempferol (DHK), into 3',4',5'-hydroxylated pentahydroxyflavanone (PHF) and dihydromyricetin (DHM), respectively. Unlike other F3'5'Hs belonging to the CYP75A subfamily, Asteraceae-specific F3'5'Hs belong to the CYP75B subfamily. Furthermore, cineraria F3'5'H expressed in yeast exhibited not only F3'H (flavonoid 3'-hydroxylase) activity but also F3'5'H activity in vitro. In this study, Southern blotting showed that there was only one copy of a homolog of the F3'5'H gene PCFH in the Pericallis × hybrida genome. This gene could be detected by Northern blot in the primary developmental stages of ligulate florets of the purple and blue-flowered cultivars, and its transcripts also accumulated in the leaves. Heterologous expression of PCFH could produce new delphinidin derivatives in the corollas of transgenic tobacco plants, increased the content of cyanidin derivatives, and lead to the blue- and red-shifting of flower color in T(0) generation plants. These results indicate that cineraria F3'5'H exhibited both F3'5'H activity and F3'H activity in vivo. The types and contents of anthocyanins and flower color phenotypes of the T(1) generation were similar to those of T(0) generation plants. PCFH exhibited stable inheritance and normal functions between generations. This study supplies new evidence to understand Asteraceae-specific F3'5'Hs and provides important references for the further study of molecular breeding of blue-flowered chrysanthemums using the PCFH gene.

Concepts: Gene, Gene expression, Molecular biology, Enzyme, Western blot, In vivo, In vitro, Southern blot

27

MYB transcriptional factors, characterized by the presence of conserved DNA binding domains (MYB domain), are involved in diverse processes including plant growth, development, metabolic and stress responses. In this study, a new R2R3-type MYB gene, NbPHAN, was identified in Nicotiana benthamiana. The NbPHAN encodes a protein of 362 amino acids and shares high sequence identities with the AS1-RS2-PHANs (ARPs) from other plant species. The NbPHAN protein targets to and forms homodimers in the nucleus. The MYB domain and C-terminal region of NbPHAN determine its subcellular localization and homodimerization, respectively. Using virus-induced gene silencing, we showed that the NbPHAN-silenced leaves exhibited severe downward curling and abnormal growth of blades along the main veins through suppressing the expression of the NTH20 gene. In addition, we found NbPHAN plays an important role in drought tolerance. The NbPHAN-silenced plants exhibited severe wilting and increased rate of water loss than that found in the non-silenced plants when growing under the water deficit condition. Although ABA accumulation was not altered in the NbPHAN-silenced plants as compared with that in the non-silenced plants, several other stress inducible genes were clearly repressed under the water deficit condition. Our results provide strong evidence that other than controlling leaf development, the ARP genes can also regulate plant tolerance to drought stress.

Concepts: DNA, Protein, Gene, Gene expression, Bacteria, Amino acid, Organism, RNA

26

The biochemical mechanisms underlying the involvement of cytosolic APXs in photosynthesis are still unknown. In this study, rice plants doubly silenced in these genes (APX1/2) were exposed to moderate light (ML) and high light (HL) to assess the role of cAPXs in photosynthetic efficiency. APX1/2 mutants that were exposed to ML overexpressed seven and five proteins involved in photochemical activity and photorespiration, respectively. These plants also increased the pheophytin and chlorophyll levels, but the amount of five proteins that are important for Calvin cycle did not change. These responses in mutants were associated with Rubisco carboxylation rate, PSII activity and potential photosynthesis, which were similar to non-transformed plants. The upregulation of photochemical proteins may be part of a compensatory mechanism for APX1/2 deficiency but apparently the finer-control for photosynthesis efficiency is dependent on Calvin cycle proteins. Conversely, under HL the mutants employed a different strategy, triggering downregulation of proteins related to photochemical activity, Calvin cycle and decreasing the levels of photosynthetic pigments. These changes were associated to strong impairment in PSII activity and Rubisco carboxylation. The upregulation of some photorespiratory proteins was maintained under that stressful condition and this response may have contributed to photo-protection in rice plants deficient in cAPXs. The data reveal that the two cytosolic APXs are not essential for photosynthesis in rice or, alternatively, the deficient plants are able to trigger compensatory mechanisms to photosynthetic acclimation under ML and HL conditions. These mechanisms involve differential regulation in protein expression related to photochemistry, Calvin cycle and photorespiration.

Concepts: Photosynthesis, Metabolism, Chloroplast, Photosystem, Calvin cycle, Photorespiration, RuBisCO, C4 carbon fixation

23

In the 20th century, annual mean temperatures in the European Alps rose by almost 1 K and are predicted to rise further, increasing the impact of temperature on alpine plants. The role of light in the heat hardening of plants is still not fully understood. Here, the alpine dwarf shrub Vaccinium gaultherioides was exposed in situ to controlled short-term heat spells (150 min with leaf temperatures 43-49°C) and long-term heat waves (7 d, 30°C) under different irradiation intensities. Lethal leaf temperatures (LT50 ) were calculated. Low solar irradiation (max. 250 PPFD) during short-term heat treatments mitigated heat stress, shown by reduced leaf tissue damage and higher Fv /Fm than in darkness. The increase in xanthophyll cycle activity and ascorbate concentration was more pronounced under low light, and free radical scavenging activity increased independent of light conditions. During long-term heat wave exposure, heat tolerance increased from 3.7 to 6.5°C with decreasing mean solar irradiation intensity (585 to 115 PPFD). Long-term exposure to heat under low light enhanced heat hardening and increased photosynthetic pigment, dehydroascorbate and violaxanthin concentration. In conclusion, V. gaultherioides is able to withstand temperatures of around 50°C, and its heat hardening can be enhanced by low light during both short and long-term heat treatment. Data showing the specific role of light during short and long-term heat exposure and the potential risk of lethal damage in alpine shrubs as a result of rising temperature are discussed.

Concepts: Energy, Light, Redox, Temperature, Hyperthermia, Thermodynamics, Vitamin C, Shrub

23

Plants live in a world where they are challenged by abiotic and biotic stresses. In response to unfavorable conditions or an acute challenge like a pathogen attack, plants employ various signaling pathways that regulate expression of defense genes and other mechanisms to provide resistance or stress adaptation. Identification of the regulatory steps in defense signaling has seen much progress in recent years. Many of the identified signaling pathways show interactions with each other, exemplified by the modulation of the jasmonic acid response by salicylic acid. Accordingly, defense regulation is more appropriately thought of as a web of interactions, rather than linear pathways. Here we describe various regulatory components and how they interact to provide an appropriate defense response. One of the common assays to monitor the output of defense signaling, as well as interaction between signaling pathways, is the measurement of altered gene expression. We illustrate that, while this is a suitable assays to monitor defense regulation, it can also inadvertently provide overstated conclusions about interaction among signaling pathways.

Concepts: DNA, Gene, Genetics, Gene expression, Bacteria, Organism, Cellular differentiation, Regulation

23

In oxygenic photosynthesis there are two “light states” - adaptations of the photosynthetic apparatus to spectral composition that otherwise favours either Photosystem I or Photosystem II. In chloroplasts of green plants, the transition to light state 2 depends on phosphorylation of apoproteins of a membrane-intrinsic antenna, the chlorophyll-a/b-binding, light-harvesting complex II (LHC II), and on the resulting redistribution absorbed excitation energy from Photosystem II to Photosystem I. The transition to light state 1 reverses these events and requires a phospho-LHC II phosphatase. Current structures of LHC II reveal little about possible steric effects of phosphorylation. The surface-exposed N-terminal domain of an LHC II polypeptide contains its phosphorylation site and is disordered in its unphosphorylated form. A molecular recognition hypothesis proposes that state transitions are a consequence of movement of LHC II between binding sites on photosystems I and II. In state 1, LHC II forms part of the antenna of photosystem II. In state 2, a unique but as yet unidentified 3-D structure of phospho-LHC II may attach it instead to Photosystem I. One possibility is that the LHC II N-terminus becomes ordered upon phosphorylation, adopting a local alpha-helical secondary structure that initiates changes in LHC II tertiary and quaternary structure that sever contact with photosystem II while securing contact with photosystem I. In order to understand redistribution of absorbed excitation energy in photosynthesis we need to know the structure of LHC II in its phosphorylated form, and in its complex with photosystem I.

Concepts: Photosynthesis, Protein, Protein structure, Chloroplast, Plant, Posttranslational modification, Photosystem, Chlorophyll

22

Adventitious root formation is a process in which roots are induced, from determined or differentiated cells that have not been specified to develop a root, at positions where they do not normally occur during development. In forest tree species, a decline in the capacity to form adventitious roots from similar cell types in stem cuttings is associated with tree age and maturity. This decline limits the success of vegetative propagation of selected adult trees. The joint action of local signals and a dynamic cascade of regulatory changes in gene expression, resulting in stereotypical cell division patterns, regulate cell fate changes that enable a somatic differentiated cell to reactivate meristem programs towards the induction of an adventitious root meristem. This article is protected by copyright. All rights reserved.

2

Hypoxia (oxygen deprivation) causes metabolic disturbances at physiological, biochemical and genetic levels and results in decreased plant growth and development. PHOSPHOLIPASE D (PLD)-mediated signalling was reported for abiotic and biotic stress signalling events in plants. To investigate the participatory role of PLDs also in hypoxia signalling, we used wild type of Arabidopsis thaliana and 10 pld isoform mutants containing C2-domain. Hypoxia-induced changes in three major signalling players, namely cytosolic free calcium (Ca2+cyt ), reactive oxygen species (ROS) and phosphatidic acid (PA), were determined in mesophyll protoplasts. The Ca2+cyt and ROS levels were monitored by fluorescence microscopy and confocal imaging, while PA levels were quantified by an enzymatic method. Our findings reveal that the elevations of cytosolic calcium and PA are reduced in all the ten mutants dysfunctional in PLD isoforms. The hypoxia-related changes in both calcium and ROS show different kinetic patterns depending on the type of PLD studied. Pharmacological experiments confirm that both external and internal sources contribute to calcium and ROS accumulation under hypoxia. PLDα1-3, PLDβ1 and PLDγ1-3 are likely involved in calcium signalling under hypoxia as well as in PA production, while all investigated PLDs, except for PLDγ3, take part in ROS elevation. This article is protected by copyright. All rights reserved.