Early Light-inducible Proteins Research Articles

Guijing2501 (Citrus unshiu) Has Stronger Cold Tolerance Due to Higher Photoprotective Capacity as Revealed by Comparative Transcriptomic and Physiological Analysis and Overexpression of Early Light-Induced Protein

Cold is one of the major limiting factors for citrus production, particularly extreme cold waves. Therefore, it is of great importance to develop cold-tolerant varieties and clarify their cold tolerance mechanisms in citrus breeding. In this study, comparative transcriptomic and physiological analyses were performed to dissect the cold tolerance mechanism of Guijing2501 (GJ2501), a new satsuma mandarin (Citrus unshiu) variety with about 1 °C lower LT50 (the median lethal temperature) relative to Guijing (GJ). The physiological analysis results revealed that GJ2501 is more cold-tolerant with less photoinhibition, PSII photodamage, and MDA accumulation, but higher POD activity than GJ under cold stress. Comparative transcriptomic analysis identified 4200 DEGs between GJ and GJ2501, as well as 4884 and 5580 up-regulated DEGs, and 5288 and 5862 down-regulated DEGs in response to cold stress in GJ and GJ2501, respectively. “Photosynthesis, light harvesting” and “photosystem” were the specific and most significantly enriched GO terms in GJ2501 in response to cold stress. Two CuELIP1 genes (encoding early light-induced proteins) related to the elimination of PSII photodamage and photoinhibition were remarkably up-regulated (by about 1000-fold) by cold stress in GJ2501 as indicated by RT-qPCR verification. Overexpression of CuELIP1 from GJ2501 in transgenic Arabidopsis protected PSII against photoinhibition under cold stress. Taken together, the cold tolerance of GJ2501 may be ascribed to its higher photoprotective capacity under cold stress.

Grapevine plantlets respond to different monochromatic lights by tuning photosynthesis and carbon allocation.

The quality of planting materials is the foundation for productivity, longevity, and berry quality of perennial grapevines with a long lifespan. Manipulating the nursery light spectrum may speed up the production of healthy and high-quality planting vines but the underlying mechanisms remain elusive. Herein, the effects of different monochromatic lights (green, blue, and red) on grapevine growth, leaf photosynthesis, whole-plant carbon allocation, and transcriptome reprograming were investigated with white light as control. Results showed that blue and red lights were favorable for plantlet growth in comparison with white light. Blue light repressed excessive growth, significantly increased the maximum net photosynthetic rate (Pn) of leaves by 39.58% and leaf specific weight by 38.29%. Red light increased the dry weight of the stem by 53.60%, the starch content of the leaf by 53.63%, and the sucrose content of the stem by 230%. Green light reduced all photosynthetic indexes of the grape plantlet. Photosynthetic photon flux density (PPFD)/Ci-Pn curves indicated that blue light affected photosynthetic rate depending on the light intensity and CO2 concentration. RNA-seq analysis of different organs (leaf, stem, and root) revealed a systematic transcriptome remodeling and VvCOP1 (CONSTITUTIVELY PHOTOMORPHOGENIC 1), VvHY5 (ELONGATED HYPOCOTYL5), VvHYH (HY5 HOMOLOG), VvELIP (early light-induced protein) and VvPIF3 (PHYTOCHROME INTERACTING FACTOR 3) may play important roles in this shoot-to-root signaling. Furthermore, the correlation network between differential expression genes and physiological traits indicated that VvpsbS (photosystem II subunit S), Vvpsb28 (photosystem II subunit 28), VvHYH, VvSUS4 (sucrose synthase 4), and VvALDA (fructose-bisphosphate aldolase) were pertinent candidate genes in responses to different light qualities. Our results provide a foundation for optimizing the light recipe of grape plantlets and strengthen the understanding of light signaling and carbon metabolism under different monochromatic lights.

Sep2, a light-harvesting complex-like protein, is involved in light stress response by binding to free chlorophylls

The light-harvesting complex (LHC) protein superfamily includes the LHC family and LHC-like family, each of which contains one or two conserved chlorophyll (Chl)-binding motifs. The LHC-like family includes early light-inducible proteins (Elips), light harvesting-like3 (Lil3s), one-helix proteins (Ohps), and stress-enhanced proteins (Seps). Currently, only the Chl-binding ability and biological function of Elips, Lil3s, and Ohps have been reported. However, the role of Seps remains unknown. In the present study, we investigated and compared the functions of two Arabidopsis (Arabidopsis thaliana) Sep proteins, Sep1 and Sep2, in Chl breakdown and in response to light stress. Our results showed that Sep1 and Sep2 contained a conserved Chl-binding motif and were localized within chloroplasts. The expression of Sep1 and Sep2 in leaves was mainly induced by light stress. Overexpression of Sep2 caused a decrease in the levels of Chl by promoting Chl breakdown, whereas Sep1 overexpression did not. Moreover, reduced accumulation of reactive oxygen species was observed in the Sep2-overexpressing Arabidopsis leaves compared with that in wild-type leaves under long-term light stress. The Sep2-induced Chl breakdown was compromised when the conserved residue (glutamic acidatposition105) of the Chl-binding motif was mutated. However, neither wild-type Sep1 nor mutant Sep1 caused Chl breakdown in Arabidopsis leaves. These findings suggested that Sep2, as a member of the LHC-like family, might regulate the levels of free Chl by binding to Chl, thereby participating in light-stress response.

Different Responses to Water Deficit of Two Common Winter Wheat Varieties: Physiological and Biochemical Characteristics.

Since water scarcity is one of the main risks for the future of agriculture, studying the ability of different wheat genotypes to tolerate a water deficit is fundamental. This study examined the responses of two hybrid wheat varieties (Gizda and Fermer) with different drought resistance to moderate (3 days) and severe (7 days) drought stress, as well as their post-stress recovery to understand their underlying defense strategies and adaptive mechanisms in more detail. To this end, the dehydration-induced alterations in the electrolyte leakage, photosynthetic pigment content, membrane fluidity, energy interaction between pigment-protein complexes, primary photosynthetic reactions, photosynthetic and stress-induced proteins, and antioxidant responses were analyzed in order to unravel the different physiological and biochemical strategies of both wheat varieties. The results demonstrated that Gizda plants are more tolerant to severe dehydration compared to Fermer, as evidenced by the lower decrease in leaf water and pigment content, lower inhibition of photosystem II (PSII) photochemistry and dissipation of thermal energy, as well as lower dehydrins' content. Some of defense mechanisms by which Gizda variety can tolerate drought stress involve the maintenance of decreased chlorophyll content in leaves, increased fluidity of the thylakoid membranes causing structural alterations in the photosynthetic apparatus, as well as dehydration-induced accumulation of early light-induced proteins (ELIPs), an increased capacity for PSI cyclic electron transport and enhanced antioxidant enzyme activity (SOD and APX), thus alleviating oxidative damage. Furthermore, the leaf content of total phenols, flavonoids, and lipid-soluble antioxidant metabolites was higher in Gizda than in Fermer.

Core cellular and tissue-specific mechanisms enable desiccation tolerance in Craterostigma.

Resurrection plants can survive prolonged life without water (anhydrobiosis) in regions with seasonal drying. This desiccation tolerance requires the coordination of numerous cellular processes across space and time, and individual plant tissues face unique constraints related to their function. Here, we analyzed the complex, octoploid genome of the model resurrection plant Craterostigma (C. plantagineum), and surveyed spatial and temporal expression dynamics to identify genetic elements underlying desiccation tolerance. Homeologous genes within the Craterostigma genome have divergent expression profiles, suggesting the subgenomes contribute differently to desiccation tolerance traits. The Craterostigma genome contains almost 200 tandemly duplicated early light-induced proteins, a hallmark trait of desiccation tolerance, with massive upregulation under water deficit. We identified a core network of desiccation-responsive genes across all tissues, but observed almost entirely unique expression dynamics in each tissue during recovery. Roots and leaves have differential responses related to light and photoprotection, autophagy and nutrient transport, reflecting their divergent functions. Our findings highlight a universal set of likely ancestral desiccation tolerance mechanisms to protect cellular macromolecules under anhydrobiosis, with secondary adaptations related to tissue function.

Transcriptional profiling analysis providing insights into desiccation tolerance mechanisms of the desert moss Syntrichia caninervis.

Syntrichia caninervis is a desiccation tolerant moss and is the dominant bryophyte found in biological soil crusts in the Gurbantunggut desert. In this study, we assessed the transcriptome profiles of S. caninervis gametophytes during the dehydration-rehydration (D-R) process (across 9 time points) using Illumina sequencing. In total, 22489 transcripts were identified, including 5337 novel transcripts, that mapped to the reference genome. A total of 12548 transcripts exhibited significant alterations in the D-R samples compared with the control samples. The differentially expressed transcripts (DETs) possessed several enriched Gene Ontology terms, such as "water stress response", "oxidation-reduction process", "membrane metabolism", "photosynthesis", and "transcription factor activity". Moreover, during early dehydration stress, the DETs were significantly enriched in stress-related pathways from the Kyoto Encyclopedia of Genes and Genomes, such as "phenylpropanoid biosynthesis", "alpha-linolenic acid metabolism", and "fructose and mannose metabolism". Photosynthesis-related transcripts (e.g., ScPsa H, ScRubisco, and ScLhcb1) were inhibited during the dehydration treatment and significantly accumulated during the late rehydration period. Most transcripts from the late embryogenesis abundant proteins (LEA) and early light-inducible protein (ELIP) families strongly accumulated at the late dehydration stage. These pathways were positively correlated with the content changes of absolute water content and Fv/Fm values, alongside peroxidase and superoxide dismutase activities. Seven transcription factor families, including AP2-ERF, bHLH, G2-like, MYB, NAC, WRKY, and bZIP, were enriched in DETs during D-R treatment. This study is the first transcriptome analysis using the S. caninervis genome for gene annotation and multigroup D-R treatment points. Our results demonstrated the detailed dynamic changes in the transcriptome of S. caninervis during the D-R process. These results also improve understanding of desiccation tolerant plants' adaptations to desiccation stress at the transcription level and provide promising gene resources for transgenic crop breeding.

Protein Changes in Shade and Sun Haberlea rhodopensis Leaves during Dehydration at Optimal and Low Temperatures

Haberlea rhodopensis is a unique resurrection plant of high phenotypic plasticity, colonizing both shady habitats and sun-exposed rock clefts. H. rhodopensis also survives freezing winter temperatures in temperate climates. Although survival in conditions of desiccation and survival in conditions of frost share high morphological and physiological similarities, proteomic changes lying behind these mechanisms are hardly studied. Thus, we aimed to reveal ecotype-level and temperature-dependent variations in the protective mechanisms by applying both targeted and untargeted proteomic approaches. Drought-induced desiccation enhanced superoxide dismutase (SOD) activity, but FeSOD and Cu/ZnSOD-III were significantly better triggered in sun plants. Desiccation resulted in the accumulation of enzymes involved in carbohydrate/phenylpropanoid metabolism (enolase, triosephosphate isomerase, UDP-D-apiose/UDP-D-xylose synthase 2, 81E8-like cytochrome P450 monooxygenase) and protective proteins such as vicinal oxygen chelate metalloenzyme superfamily and early light-induced proteins, dehydrins, and small heat shock proteins, the latter two typically being found in the latest phases of dehydration and being more pronounced in sun plants. Although low temperature and drought stress-induced desiccation trigger similar responses, the natural variation of these responses in shade and sun plants calls for attention to the pre-conditioning/priming effects that have high importance both in the desiccation responses and successful stress recovery.

Protective Strategies of Haberlea rhodopensis for Acquisition of Freezing Tolerance: Interaction between Dehydration and Low Temperature

Resurrection plants are able to deal with complete dehydration of their leaves and then recover normal metabolic activity after rehydration. Only a few resurrection species are exposed to freezing temperatures in their natural environments, making them interesting models to study the key metabolic adjustments of freezing tolerances. Here, we investigate the effect of cold and freezing temperatures on physiological and biochemical changes in the leaves of Haberlea rhodopensis under natural and controlled environmental conditions. Our data shows that leaf water content affects its thermodynamical properties during vitrification under low temperatures. The changes in membrane lipid composition, accumulation of sugars, and synthesis of stress-induced proteins were significantly activated during the adaptation of H. rhodopensis to both cold and freezing temperatures. In particular, the freezing tolerance of H. rhodopensis relies on a sucrose/hexoses ratio in favor of hexoses during cold acclimation, while there is a shift in favor of sucrose upon exposure to freezing temperatures, especially evident when leaf desiccation is relevant. This pattern was paralleled by an elevated ratio of unsaturated/saturated fatty acids and significant quantitative and compositional changes in stress-induced proteins, namely dehydrins and early light-induced proteins (ELIPs). Taken together, our data indicate that common responses of H. rhodopensis plants to low temperature and desiccation involve the accumulation of sugars and upregulation of dehydrins/ELIP protein expression. Further studies on the molecular mechanisms underlying freezing tolerance (genes and genetic regulatory mechanisms) may help breeders to improve the resistance of crop plants.

Light regulates chlorophyll biosynthesis via ELIP1 during the storage of Chinese cabbage

Chlorophyll loss is a major problem during green vegetable storage. However, the molecular mechanism is still unclear. In this study, a 21 days of storage experiments showed chlorophyll content was higher in light-stored Chinese cabbage (Brassica chinensis L.) leaves than those in dark-stored samples. Transcriptome analyses were performed on these samples to determine the effects of light. Among 311 differentially expressed genes (DEGs), early light-induced protein 1 (ELIP1) was identified as the main control gene for chlorophyll synthesis. Tissues and subcellular localization indicated that ELIP1 was localized in the nucleus. Motifs structure analyses, chromatin immunoprecipitation (ChIP) assays, luciferase reporter assays, and overexpression experiments demonstrated that ELIP1 regulated the expressions of genomes uncoupled 4 (GUN4), Glutamyl-tRNA reductase family protein (HEMA1), and Mg-protoporphyrin IX methyltransferase (CHLM) by binding to G-box-like motifs and affected chlorophyll biosynthesis during the storage of Chinese cabbage. It is a possible common tetrapyrrole biosynthetic pathway for chlorophylls, hemes, and bilin pigments in photosynthetic organisms. Our research also revealed that white light can be used as a regulatory factor to improve the storage ability and extent shelf life of Chinese cabbage.

Photoprotection conferring plant tolerance to freezing stress through rescuing photosystem in evergreen Rhododendron.

Light stress is one of the important stresses for winter survival in evergreens, especially for plants with broad leaves, like evergreen rhododendrons. Photoprotection has been shown to upregulate dramatically in rhododendrons during winter, but whether it directly contributes to enhancing the freezing tolerance is still unknown. In this study, we found that the expression and circadian rhythm of an early light-induced protein (ELIP)-RhELIP3-which exerts photoprotection in Rhododendron 'Elsie Lee', could be impacted by both photoperiod and low temperature, with low temperature being the predominant inducer. Arabidopsis overexpressing RhELIP3 displayed significantly stronger freezing tolerance and better photosystem II function after a 3-day recovery from freezing treatment. Moreover, RhHY5 binds with theRhELIP3 promoter to activate its expression. Arabidopsis overexpressing RhHY5 exhibited stronger freezing tolerance and better photosystem II function. AtELIP1 and AtELIP2 were significantly induced in RhHY5-overexpressed Arabidopsis at low temperatures. We also discovered that RhBBX24 binds directly to RhELIP3 promoter and suppresses its expression. RhBBX24 can also interact with RhHY5 and inhibit the interaction of RhHY5-RhELIP3. RhELIP3, RhHY5,and RhBBX24 exhibited similar circadian rhythms under low temperature with short period. Overall, our investigation highlights that photoprotection is involved in improving the freezing tolerance of evergreen rhododendrons.

Photoprotection contributes to freezing tolerance as revealed by RNA-seq profiling of rhododendron leaves during cold acclimation and deacclimation over time.

Cold acclimation (CA) and deacclimation (DA), which are often accompanied by changes in freezing tolerance (FT), carbohydrates and hormones, are crucial for winter survival, especially under global warming. Plants with weak CA and premature DA caused by warm winters and/or unseasonal warm spells can be easily injured by adverse reactions to cold. Thus, understanding the molecular mechanisms of FT is imperative. In this study, we used high-throughput RNA-seq to profile the CA and DA of leaves of overwintering Rhododendron "Miyo-no-Sakae" over time; these leaves do not undergo dormancy but do undergo photoprotection during CA, and they do not grow during DA. Using Mfuzz and weighted gene coexpression network analysis, we identified specific transcriptional characteristics in each phase of CA and DA and proposed networks involving coexpressed genes and physiological traits. In particular, we discovered that the circadian rhythm is critical for obtaining the strongest FT, and high expression of circadian rhythm-related genes might be linked to sugar accumulation during winter. Furthermore, evergreen leaves exhibited robust photoprotection during winter, as revealed by high values of nonphotochemical quenching, high expression of transcripts annotated as "early light-induced proteins", loss of granum stacks and destacking of thylakoids, all of which were alleviated during DA. The strong requirement of photoprotection could be the reason for decreased abscisic acid (ABA) and jasmonic acid (JA) contents during CA, and decreases in ABA and JA contents may contribute to decreases in lignin content. Our data suggest that the molecular mechanisms of FT in overwintering leaves are unique, which may be due to the high requirements for photoprotection during winter.

Identification of resurrection genes from the transcriptome of dehydrated and rehydrated Selaginella tamariscina

ABSTRACT Selaginella tamariscina is a lycophyta species that survives under extremely dry conditions via the mechanism of resurrection. This phenomenon involves the regulation of numerous genes that play vital roles in desiccation tolerance and subsequent rehydration. To identify resurrection-related genes, we analyzed the transcriptome between dehydration conditions and rehydration conditions of S. tamariscina. The de novo assembly generated 124,417 transcripts with an average size of 1,000 bp and 87,754 unigenes. Among these genes, 1,267 genes and 634 genes were up and down regulated by rehydration compared to dehydration. To understand gene function, we annotated Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG). The unigenes encoding early light-inducible protein (ELIP) were down-regulated, whereas pentatricopeptide repeat-containing protein (PPR), late embryogenesis abundant proteins (LEA), sucrose nonfermenting protein (SNF), trehalose phosphate phosphatase (TPP), trehalose phosphate synthase (TPS), and ABC transporter G family (ABCG) were significantly up-regulated in response to rehydration conditions by differentially expressed genes (DEGs) analysis. Several studies provide evidence that these genes play a role in stress environment. The ELIP and PPR genes are involved in chloroplast protection during dehydration and rehydration. LEA, SNF, and trehalose genes are known to be oxidant scavengers that protect the cell structure from the deleterious effect of drought. TPP and TPS genes were found in the starch and sucrose metabolism pathways, which are essential sugar-signaling metabolites regulating plant metabolism and other biological processes. ABC-G gene interacts with abscisic acid (ABA) phytohormone in the stomata opening during stress conditions. Our findings provide valuable information and candidate resurrection genes for future functional analysis aimed at improving the drought tolerance of crop plants.

Transcriptomic Effects of Acute Ultraviolet Radiation Exposure on Two Syntrichia Mosses.

Ultraviolet radiation (UVR) is a major environmental stressor for terrestrial plants. Here we investigated genetic responses to acute broadband UVR exposure in the highly desiccation-tolerant mosses Syntrichia caninervis and Syntrichia ruralis, using a comparative transcriptomics approach. We explored whether UVR protection is physiologically plastic and induced by UVR exposure, addressing the following questions: (1) What is the timeline of changes in the transcriptome with acute UVR exposure in these two species? (2) What genes are involved in the UVR response? and (3) How do the two species differ in their transcriptomic response to UVR? There were remarkable differences between the two species after 10 and 30 min of UVR exposure, including no overlap in significantly differentially abundant transcripts (DATs) after 10 min of UVR exposure and more than twice as many DATs for S. caninervis as there were for S. ruralis. Photosynthesis-related transcripts were involved in the response of S. ruralis to UVR, while membrane-related transcripts were indicated in the response of S. caninervis. In both species, transcripts involved in oxidative stress and those important for desiccation tolerance (such as late embryogenesis abundant genes and early light-inducible protein genes) were involved in response to UVR, suggesting possible roles in UVR tolerance and cross-talk with desiccation tolerance in these species. The results of this study suggest potential UVR-induced responses that may have roles outside of UVR tolerance, and that the response to URV is different in these two species, perhaps a reflection of adaptation to different environmental conditions.

Early Light-Inducible Protein (ELIP) Can Enhance Resistance to Cold-Induced Photooxidative Stress in Chlamydomonas reinhardtii.

Cold weather is one of the biggest challenges in establishing a large-scale microalgae culture facility in temperate regions. In order to develop a strain that is resistant to low temperatures and still maintains high photosynthetic efficiency, transgenic studies have been conducted targeting many genes. Early light-inducible proteins (ELIPs) located in thylakoid membranes are known to protect photosynthetic machinery from various environmental stresses in higher plants. An ELIP homolog was identified from Chlamydomonas reinhardtii and named ELIP3. The role of the gene was analyzed in terms of photosynthetic CO2 assimilation under cold stress. Western blot results showed a significant accumulation of ELIP3 when the cells were exposed to cold stress (4°C). High light stress alone did not induce the accumulation of the protein. Enhanced expression of ELIP3 helped survival of the cell under photo-oxidative stress. The influx of CO2 to the photobioreactor induced strong accumulation of ELIP3, and enhanced survival of the cell under high light and cold stress. When the oxidative stress was reduced by adding a ROS quencher, TEMPOL, to the media the expression of ELIP3 was reduced. A knockdown mutant showed much lower photosynthetic efficiency than wild type in low temperature, and died rapidly when it was exposed to high light and cold stress. The overexpression mutant survived significantly longer in the same conditions. Interestingly, knockdown mutants showed negative phototaxis, while the overexpression mutant showed positive phototaxis. These results suggest that ELIP3 may be involved in the regulation of the redox state of the cell and takes important role in protecting the photosystem under photooxidative stress in low temperatures.

The genome of Prasinoderma coloniale unveils the existence of a third phylum within green plants

Genome analysis of the pico-eukaryotic marine green alga Prasinoderma coloniale CCMP 1413 unveils the existence of a novel phylum within green plants (Viridiplantae), the Prasinodermophyta, which diverged before the split of Chlorophyta and Streptophyta. Structural features of the genome and gene family comparisons revealed an intermediate position of the P. coloniale genome (25.3 Mb) between the extremely compact, small genomes of picoplanktonic Mamiellophyceae (Chlorophyta) and the larger, more complex genomes of early-diverging streptophyte algae. Reconstruction of the minimal core genome of Viridiplantae allowed identification of an ancestral toolkit of transcription factors and flagellar proteins. Adaptations of P. coloniale to its deep-water, oligotrophic environment involved expansion of light-harvesting proteins, reduction of early light-induced proteins, evolution of a distinct type of C4 photosynthesis and carbon-concentrating mechanism, synthesis of the metal-complexing metabolite picolinic acid, and vitamin B1, B7 and B12 auxotrophy. The P. coloniale genome provides first insights into the dawn of green plant evolution.

Transcriptome Analysis of Genes Involved in Cold Hardiness of Peach Tree (Prunus persica) Shoots during Cold Acclimation and Deacclimation.

We analyzed the transcriptomes in the shoots of five-year-old ‘Soomee’ peach trees (Prunus persica) during cold acclimation (CA), from early CA (end of October) to late CA (middle of January), and deacclimation (DA), from late CA to late DA (middle of March), to identify the genes involved in cold hardiness. Cold hardiness of the shoots increased from early to late CA, but decreased from late CA to late DA, as indicated by decreased and increased the median lethal temperature (LT50), respectively. Transcriptome analysis identified 17,208 assembled transcripts during all three stages. In total, 1891 and 3008 transcripts were differentially expressed with a |fold change| > 2 (p < 0.05) between early and late CA, and between late CA and late DA, respectively. Among them, 1522 and 2830, respectively, were functionally annotated with gene ontology (GO) terms having a greater proportion of differentially expressed genes (DEGs) associated with molecular function than biological process or cellular component categories. The biochemical pathways best represented both periods from early to late CA and from late CA to late DA were ‘metabolic pathway’ and ‘biosynthesis of secondary metabolites’. We validated these transcriptomic results by performing reverse transcription quantitative polymerase chain reaction on the selected DEGs showing significant fold changes. The relative expressions of the selected DEGs were closely related to the LT50 values of the peach tree shoots: ‘Soomee’ shoots exhibited higher relative expressions of the selected DEGs than shoots of the less cold-hardy ‘Odoroki’ peach trees. Irrespective of the cultivar, the relative expressions of the DEGs that were up- and then down-regulated during CA, from early to late CA, and DA, from late CA to late DA, were more closely correlated with cold hardiness than those of the DEGs that were down- and then up-regulated. Therefore, our results suggest that the significantly up- and then down-regulated DEGs are associated with cold hardiness in peach tree shoots. These DEGs, including early light-induced protein 1, chloroplastic, 14-kDa proline-rich protein DC2.15, glutamate dehydrogenase 2, and triacylglycerol lipase 2, could be candidate genes to determine cold hardiness.

Functional Aspects of Early Light-Induced Protein (ELIP) Genes from the Desiccation-Tolerant Moss Syntrichia caninervis.

The early light-induced proteins (ELIPs) are postulated to act as transient pigment-binding proteins that protect the chloroplast from photodamage caused by excessive light energy. Desert mosses such as Syntrichia caninervis, that are desiccation-tolerant and homoiochlorophyllous, are often exposed to high-light conditions when both hydrated and dry ELIP transcripts are accumulated in response to dehydration. To gain further insights into ELIP gene function in the moss S. caninervis, two ELIP cDNAs cloned from S. caninervis, ScELIP1 and ScELIP2 and both sequences were used as the basis of a transcript abundance assessment in plants exposed to high-light, UV-A, UV-B, red-light, and blue-light. ScELIPs were expressed separately in an Arabidopsis ELIP mutant Atelip. Transcript abundance for ScELIPs in gametophytes respond to each of the light treatments, in similar but not in identical ways. Ectopic expression of either ScELIPs protected PSII against photoinhibition and stabilized leaf chlorophyll content and thus partially complementing the loss of AtELIP2. Ectopic expression of ScELIPs also complements the germination phenotype of the mutant and improves protection of the photosynthetic apparatus of transgenic Arabidopsis from high-light stress. Our study extends knowledge of bryophyte photoprotection and provides further insight into the molecular mechanisms related to the function of ELIPs.

ORANGE Represses Chloroplast Biogenesis in Etiolated Arabidopsis Cotyledons via Interaction with TCP14.

The conversion of etioplasts into chloroplasts in germinating cotyledons is a crucial transition for higher plants, enabling photoautotrophic growth upon illumination. Tight coordination of chlorophyll biosynthesis and photosynthetic complex assembly is critical for this process. ORANGE (OR), a DnaJ-like zinc finger domain-containing protein, was reported to trigger the biogenesis of carotenoid-accumulating plastids by promoting carotenoid biosynthesis and sequestration. Both nuclear and plastidic localizations of OR have been observed. Here, we show that Arabidopsis (Arabidopsis thaliana) OR physically interacts with the transcription factor TCP14 in the nucleus and represses its transactivation activity. Through this interaction, the nucleus-localized OR negatively regulates expression of EARLY LIGHT-INDUCIBLE PROTEINS (ELIPs), reduces chlorophyll biosynthesis, and delays development of thylakoid membranes in the plastids of germinating cotyledons. Nuclear abundance of OR decreased upon illumination. Together with an accumulation of TCP14 in the nucleus, this derepresses chloroplast biogenesis during de-etiolation. TCP14 is epistatic to OR and expression of ELIPs is directly regulated by the binding of TCP14 to Up1 elements in the ELIP promoter regions. Our results demonstrate that the interaction between OR and TCP14 in the nucleus leads to repression of chloroplast biogenesis in etiolated seedlings and provide new insights into the regulation of early chloroplast development.plantcell;31/12/2996/FX1F1fx1.

Production of marker-free tomato plants expressing the supersweet protein thaumatin II gene under the control of predominantly fruit-specific promoters

Despite the lack of evidence of the danger of genetically modified organisms the presence of marker and antibiotic-resistant genes in transgenic plants causes concern to consumers. Genetically modified plants with viral and bacterial genes are adopted by consumers, but with concerns; in addition, constitutive promoters have a number of disadvantages in industrial-scale cultivation of plants. In our study, we used the pMF vector system (Wageningen Plant Research, Wageningen, Netherlands), which combines inducible site-specific recombinase and a bifunctional selectable gene to obtain marker-free tomato plants. The gene of interest was the supersweet thaumatin II protein from the tropical plant Thaumatococcus daniellii under the control of tomato predominantly fruit-specific early-light inducible protein (ELIP) or E8 promoters and tomato Rubisco terminator. The use of this gene in our laboratory allowed enhancing sweetness, as well as improving the taste characteristics of fruit such as apple, strawberries, carrots, tomatoes, and pears. By using different strategies of early and delayed selection we developed a protocol for obtaining fully marker-free tomato plants, which was checked by polymerase chain reaction and Southern blotting. The thaumatin II gene expression was confirmed by reverse transcription-PCR and western blotting analyses. The fruit of transgenic and marker-free tomato plants displayed a sweet taste. A quantitative comparative assessment of the level of expression of the thaumatin protein under the control of two promoters was carried out using enzyme-linked immunosorbent assay. Multiple and/or incomplete T-DNA inserts that often occur during transformation of Solanaceae greatly reduced the efficiency of the system used. The strong tomato ELIP promoter provides a high level of expression of the supersweet thaumatin II protein gene in the fruit of marker-free tomato plants.

Functional characterization of a strong promoter of the early light-inducible protein gene from tomato.

The tomato ELIP gene promoter is mainly active in the ripening fruit. Considering its high activity, the promoter could be used for molecular breeding of plants in the future. The ability to obtain new varieties of transgenic plants with economically valuable traits relies on a high level of target gene expression, which is largely controlled by a gene promoter. Hence, research aimed at finding and characterizing new tissue-specific promoters that direct gene expression in specific plant tissues or at certain developmental stages has become the most important field of plant biotechnology. Here, we cloned and characterized the promoter of the early light-inducible protein (ELIP) gene from tomato (Solanum lycopersicum cv. Yalf). ELIPs are produced in the presence of light and putatively function in the chloroplast-to-chromoplast conversion, playing a photorepairing role in the photosynthetic system. Analysis of the promoter sequence revealed multiple cis-acting elements related to light responsiveness, and other motifs involved in plant hormone response and circadian control. To determine the functionality of the promoter, seven 5'-deletion variants were fused with the β-glucuronidase (GUS) reporter gene and introduced into tomato. Histochemical analysis of transgenic tomato plants revealed different levels of GUS activity in most analyzed tissues, depending on the promoter fragment used. The intensity of staining was considerably higher in ripening fruits than in unripe and non-fruit tissues. Quantitative analysis indicated that the level of GUS activity with the longest (full-length) version of the ELIP promoter in ripened fruits was comparable to that in plants expressing the constitutive CaMV35S promoter. Further, the location of both negative and positive regulatory motifs was identified. The described ELIP promoter is a potential tool for various applications in plant biotechnology.