Increased Chlorophyll Accumulation Research Articles

Detoxification and catabolism of mesotrione and fomesafen facilitated by a Phase II reaction acetyltransferase in rice

IntroductionThe excessive dosage of pesticides required for agronomic reality results in growing contamination of pesticide residues in environment, thus bringing high risks to crop production and human health. ObjectivesThis study aims to unveil a novel mechanism for catabolism of two pesticides MTR and FSA facilitated by an uncharacterized Phase II reaction enzyme termed acetyltransferase-1 (ACE1) in rice and to make assessment of its potential for bioremediation to minimize the risks to crop production and food safety. MethodsWe developed genetically improved cultivars overexpressing OsACE1 (OE) and knockout mutant lines by CRISPR-Cas9 technology to identify the MTR and FSA detoxic and metabolic functions and characterized their metabolites and conjugates by HPLC-LTQ-MS/MS. ResultsOsACE1 overexpression conferred rice resistance to toxicity of MTR/FSA compared to wild-type, manifested by improved plant elongation and biomass, attenuated cellular injury, and increased chlorophyll accumulation. The OE plants accumulated significantly less parent MTR/FSA and more degradative metabolites, and removed MTR/FSA from their growth medium by 1.38 and 1.61 folds over the wild-type. In contrast, knocking out OsACE1 led to compromised growth fitness and intensified toxic symptoms under MTR/FSA stress and accumulation of more toxic MTR and FSA in rice. The reduced metabolites of MTR and FSA detected in the Cas9 plants suggest the impaired capability of OsACE1 function. ConclusionsThese results signified that OsACE1 expression is required for detoxifying the two poisoning chemicals in rice and plays a critical role in accelerating breakdown of the pesticides mainly through Phase II reaction mechanism pathways.

Comprehensive Analysis of the SBP Family in Blueberry and Their Regulatory Mechanism Controlling Chlorophyll Accumulation.

SQUAMOSA Promoter Binding Protein (SBP) family genes act as central players to regulate plant growth and development with functional redundancy and specificity. Addressing the diversity of the SBP family in crops is of great significance to precisely utilize them to improve agronomic traits. Blueberry is an important economic berry crop. However, the SBP family has not been described in blueberry. In the present study, twenty VcSBP genes were identified through data mining against blueberry transcriptome databases. These VcSBPs could be clustered into eight groups, and the gene structures and motif compositions are divergent among the groups and similar within each group. The VcSBPs were differentially expressed in various tissues. Intriguingly, 10 VcSBPs were highly expressed at green fruit stages and dramatically decreased at the onset of fruit ripening, implying that they are important regulators during early fruit development. Computational analysis showed that 10 VcSBPs were targeted by miR156, and four of them were further verified by degradome sequencing. Moreover, their functional diversity was studied in Arabidopsis. Noticeably, three VcSBPs significantly increased chlorophyll accumulation, and qRT-PCR analysis indicated that VcSBP13a in Arabidopsis enhanced the expression of chlorophyll biosynthetic genes such as AtDVR, AtPORA, AtPORB, AtPORC, and AtCAO. Finally, the targets of VcSBPs were computationally identified in blueberry, and the Y1H assay showed that VcSBP13a could physically bind to the promoter region of the chlorophyll-associated gene VcLHCB1. Our findings provided an overall framework for individually understanding the characteristics and functions of the SBP family in blueberry.

SlMYB72 Regulates the Metabolism of Chlorophylls, Carotenoids, and Flavonoids in Tomato Fruit.

Tomato (Solanum lycopersicum) fruit ripening is accompanied by the degradation of chlorophylls and the accumulation of carotenoids and flavonoids. Tomato SlMYB72 belongs to the R2R3 MYB subfamily, is located in the nucleus, and possesses transcriptional activator activity. Down-regulation of the SlMYB72 gene produced uneven-colored fruits; that is, dark green spots appeared on immature and mature green fruits, whereas yellow spots appeared on red fruits. Down-regulation of SlMYB72 increased chlorophyll accumulation, chloroplast biogenesis and development, and photosynthesis rate in fruits. This down-regulation decreased lycopene content, promoted β-carotene production and chromoplast development, and increased flavonoid accumulation in fruits. RNA sequencing analysis revealed that down-regulation of SlMYB72 altered the expression levels of genes involved in the biosynthesis of chlorophylls, carotenoids, and flavonoids. SlMYB72 protein interacted with the auxin response factor SlARF4. SlMYB72 directly targeted protochlorophyllide reductase, Mg-chelatase H subunit, and knotted1-like homeobox2 genes and regulated chlorophyll biosynthesis and chloroplast development. SlMYB72 directly bound to phytoene synthase, ζ-carotene isomerase, and lycopene β-cyclase genes and regulated carotenoid biosynthesis. SlMYB72 directly targeted 4-coumarate-coenzyme A ligase and chalcone synthase genes and regulated the biosynthesis of flavonoids and phenolic acid. The uneven color phenotype in RNA interference-SlMYB72 fruits was due to uneven silencing of SlMYB72 and uneven expression of chlorophyll, carotenoid, and flavonoid biosynthesis genes. In summary, this study identified important roles for SlMYB72 in the regulation of chlorophyll, carotenoid, and flavonoid metabolism and provided a potential target to improve fruit nutrition in horticultural crops.

Complementation studies of the Arabidopsis fc1 mutant substantiate essential functions of ferrochelatase 1 during embryogenesis and salt stress.

Ferrochelatase (FC) is the final enzyme for haem formation in the tetrapyrrole biosynthesis pathway and encoded by two genes in higher plants. FC2 exists predominantly in green tissue, whereas FC1 is constitutively expressed. We intended to substantiate the specific roles of FC1. The embryo-lethal fc1-2 mutant was used to express the two genomic FC-encoding sequences under the FC1 and FC2 promoter and explore the complementation of the FC1 deficiency. Apart from the successful complementation with FC1, expression of FC2 under control of the FC1 promoter (pFC1::FC2) compensates for missing FC1 but not by FC2 promoter expression. The complementing lines pFC1FC2(fc1/fc1) succeeded under standard growth condition but failed under salt stress. The pFC1FC2(fc1/fc1) line exhibited symptoms of leaf senescence, including accelerated loss of haem and chlorophyll and elevated gene expression for chlorophyll catabolism. In contrast, ectopic FC1 expression (p35S::FC1) resulted in increased chlorophyll accumulation. The limited ability of FC2 to complement fc1 is explained by a faster turnover of FC2 mRNA during stress. It is suggested that FC1-produced haem is essential for embryogenesis and stress response. The pFC1::FC2 expression readily complements the fc1-2 embryo lethality, whereas higher FC1 transcript content contributes essentially to stress tolerance.

Heterologous expression of a novel Poa pratensis gibberellin 2-oxidase gene, PpGA2ox, caused dwarfism, late flowering, and increased chlorophyll accumulation in Arabidopsis

Gibberellin 2-oxidases (GA2oxs) irreversibly convert bioactive gibberellins (GAs) and their immediate precursors into inactive GAs via 2-β hydroxylation and so regulate gibberellin content in plants. However, to the best of our knowledge, little has been known about the GA2oxs and its function in cool season turfgrass Poa pratensis. In this study, rapid amplification of cDNA end (RACE) was employed to isolate PpGA2ox from P. pratensis. The open reading frame of PpGA2ox was 1 047 bp in length, corresponding to 348 amino acids. PpGA2ox was localized in both nucleus and cytoplasm. The expression of PpGA2ox could be up-regulated by 10 μM gibberellic acid, 5 μM methyl jasmonate, or 10 μM indole-3-acetic acid. In addition, its native promoter could drive GUS expression in both leaf apex and shoot apical region. Moreover, overexpression of PpGA2ox in Arabidopsis led to GA-deficiency leading to dwarf phenotype, delayed flowering time, and increased chlorophyll content. Our study suggests that PpGA2ox could be a candidate gene for breeding new cultivars of P. pratensis.

Light quantity and quality supplies sharply affect growth, morphological, physiological and quality traits of basil

Plants sharply adapt their growth and physiology to light availability. This study aimed at evaluating the effect of light quantity and quality manipulation on growth, morphological traits, pigment and secondary metabolites content in basil as well as comprehending the mechanisms which regulate such responses.Two experiments were carried out under greenhouse in 2014 (spring transplanting, Spr_Tr) and 2015 (summer transplanting, Sum_Tr). On a complete randomized block design, plants of basil were exposed to three modifications of the transmitted solar radiation with colored plastic films: yellow (YF), green (GF) and blue films (BF), plus a control (Control).Leaf pairs, axillary shoots, total fresh and dry biomass, specific leaf area, soil-plant analysis development, reflectance indices (Normalized Different Vegetation Index670, NDVI670, and Optimized Soil-Adjusted Vegetation Index, OSAVI), total chlorophyll, chlorophyll a, chlorophyll b, carotenoids, single and total polyphenol content and radical scavenging activity were recorded and examined.Shading induced stem elongation, a greater leaf area expansion and a lower leaf thickness; moreover, shaded plants increased chlorophyll accumulation (on average +29.4% and +21.6% during Spr_Tr and Sum_Tr, respectively). YF treatment allowed always the highest biomass accumulation (averaged over crop cycle: 2.1 and 3.4 g plant−1 during Spr_Tr and Sum_Tr, respectively).OSAVI and NDVI670 seem the more suitable indicators for chlorophyll accumulation. Light manipulation influenced specific phenolic compounds concentration. The application of colored films lowered rosmarinic and caftaric acids (by 29.8% and 33.2%, respectively, averaged over treatments and crop cycle). Antiradical activity was linearly correlated only with caffeic acid.Light manipulation represents a promising tool for the manipulation of basil morphological, physiological and quality traits.

Silencing SlELP2L, a tomato Elongator complex protein 2-like gene, inhibits leaf growth, accelerates leaf, sepal senescence, and produces dark-green fruit.

The multi-subunit complex Elongator interacts with elongating RNA polymerase II (RNAPII) and is thought to facilitate transcription through histone acetylation. Elongator is highly conserved in eukaryotes, yet has multiple kingdom-specific functions in diverse organisms. Recent genetic studies performed in Arabidopsis have demonstrated that Elongator functions in plant growth and development, and in response to biotic and abiotic stress. However, little is known about its roles in other plant species. Here, we study the function of an Elongator complex protein 2-like gene in tomato, here designated as SlELP2L, through RNAi-mediated gene silencing. Silencing SlELP2L in tomato inhibits leaf growth, accelerates leaf and sepal senescence, and produces dark-green fruit with reduced GA and IAA contents in leaves, and increased chlorophyll accumulation in pericarps. Gene expression analysis indicated that SlELP2L-silenced plants had reduced transcript levels of ethylene- and ripening-related genes during fruit ripening with slightly decreased carotenoid content in fruits, while the expression of DNA methyltransferase genes was up-regulated, indicating that SlELP2L may modulate DNA methylation in tomato. Besides, silencing SlELP2L increases ABA sensitivity in inhibiting seedling growth. These results suggest that SlELP2L plays important roles in regulating plant growth and development, as well as in response to ABA in tomato.

Assembly of Synthetic Locked Phycocyanobilin Derivatives with Phytochrome in Vitro and in Vivo in Ceratodon purpureus and Arabidopsis

Phytochromes are photoreceptors with a bilin chromophore in which light triggers the conversion between the red light-absorbing form, Pr, and the far-red-light-absorbing form, Pfr. Here we performed in vitro and in vivo studies using locked phycocyanobilin derivatives, termed 15 Z anti phycocyanobilin (15ZaPCB) and 15 E anti PCB (15EaPCB). Recombinant bacterial and plant phytochromes incorporated either chromophore in a noncovalent or covalent manner. All adducts were photoinactive. The absorption spectra of the 15ZaPCB and 15EaPCB adducts were comparable with those of the Pr and Pfr form, respectively. Feeding of 15EaPCB, but not 15ZaPCB, to protonemal filaments of the moss Ceratodon purpureus resulted in increased chlorophyll accumulation, modulation of gravitropism, and induction of side branches in darkness. The effect of locked chromophores on phytochrome responses, such as induction of seed germination, inhibition of hypocotyl elongation, induction of cotyledon opening, randomization of gravitropism, and gene regulation, were investigated in wild-type Arabidopsis thaliana and the phytochrome-chromophore-deficient long hypocotyl mutant hy1. All phytochrome responses were induced in darkness by 15EaPCB, not only in the mutant but also in the wild type. These studies show that the 15Ea stereochemistry of the chromophore results in the formation of active Pfr-like phytochrome in the cell. Locked chromophores might be used to investigate phytochrome responses in many other organisms without the need to isolate mutants. The induction of phytochrome responses in the hy1 mutant by 15EaPCB were however less efficient than by red light irradiation given to biliverdin-rescued seeds or seedlings.

Cytokinin and abscisic acid control plastid gene transcription during barley seedling de-etiolation

The regulatory role of cytokinins and abscisic acid (ABA) during de-etiolation of monocotyledonous plants is not sufficiently studied. The effects of cytokinin—6-benzyladenine (BA) and ABA on leaf greening and chloroplast gene transcription were studied on leaves detached from etiolated barley (Hordeum vulgare L.) seedlings. The influence of both phytohormones was age-dependent. Cytokinin practically had no effect on the youngest (3-day-old) leaves, increased chlorophyll accumulation in 6-day-old leaves, and this effect decreased in 9-day-old leaves. ABA reduced chlorophyll accumulation in the youngest leaves, and this effect was blocked by cytokinin. ABA alone had practically no effect on greening of leaves detached from 6- and 9-day-old seedlings, but fully inhibited activation of this process by cytokinin. Thus, antagonism between ABA and cytokinin was observed for seedlings of all ages. The rate of transcription of chloroplast genes was studied by run-on transcription in isolated chloroplasts in the course of etiolated seedling growth in darkness, and age-dependent decrease was demonstrated. It was established that cytokinin activated transcription of 11 chloroplast genes among 30 studied, and ABA suppressed transcription of 12 plastid genes. Such differential effect of cytokinin and ABA on chloroplast gene transcription in monocot seedlings during etiolated leaf greening was not known before.

Alleviation of Copper-Induced Oxidative Damage in Chlamydomonas reinhardtii by Carbon Monoxide

Carbon monoxide (CO) is an endogenous gaseous molecule in plants and animals. Recent studies have shown that it is one of the most essential cellular components regulating many aspects of plant growth and development. However, whether CO regulates the green algae adaptive response to heavy metal toxicity is unknown. The present study investigated the role of CO in regulating Cu-induced oxidative stress in eukaryotic algae Chlamydomonas reinhardtii. Cells pretreated with 5μM CO for 30min and followed by exposure to 5μM Cu(II) for 4days showed attenuated toxicity. The CO-improved growth of algae was correlated with reduced lipid peroxidation and increased chlorophyll accumulation. The beneficial effect of CO was confirmed by histochemical staining with reactive oxygen species. Further, treatment with 5μM CO increased the activity of catalase with Cu. However, a reduced superoxide dismutase activity was observed in the CO+Cu-treated algae compared to the control (activity of Cu treatment alone). Under the same condition, the activity of ascorbate peroxidase was not significantly changed. These results suggest that CO can play an important role in regulating the response of algae to Cu stress.

MicroRNA171c-Targeted SCL6-II, SCL6-III, and SCL6-IV Genes Regulate Shoot Branching in Arabidopsis

MicroRNAs (miRNAs) are ∼21-nucleotide noncoding RNAs that play critical roles in regulating plant growth and development through directing the degradation of target mRNAs. Axillary meristem activity, and hence shoot branching, is influenced by a complicated network that involves phytohormones such as auxin, cytokinin, and strigolactone. GAI, RGA, and SCR (GRAS) family members take part in a variety of developmental processes, including axillary bud growth. Here, we show that the Arabidopsis thaliana microRNA171c (miR171c) acts to negatively regulate shoot branching through targeting GRAS gene family members SCARECROW-LIKE6-II (SCL6-II), SCL6-III, and SCL6-IV for cleavage. Transgenic plants overexpressing MIR171c (35Spro-MIR171c) and scl6-II scl6-III scl6-IV triple mutant plants exhibit a similar reduced shoot branching phenotype. Expression of any one of the miR171c-resistant versions of SCL6-II, SCL6-III, and SCL6-IV in 35Spro-MIR171c plants rescues the reduced shoot branching phenotype. Scl6-II scl6-III scl6-IV mutant plants exhibit pleiotropic phenotypes such as increased chlorophyll accumulation, decreased primary root elongation, and abnormal leaf and flower patterning. SCL6-II, SCL6-III, and SCL6-IV are located to the nucleus, and show transcriptional activation activity. Our results suggest that miR171c-targeted SCL6-II, SCL6-III, and SCL6-IV play an important role in the regulation of shoot branch production.

Triadimefon induced physiological and ultra structural changes for moisture stress protection in Bougainvillea (Bougainvillea spectabilis Willd) at nursery stage

Abstract: Hardwood stem cuttings of bougainvillea (Bougainvillea spectabilis Willd) consisting of two to four leaves were treated with triadimefon (triazole derivative) in the form of foliar spray at 50 ppm and 100 ppm, while the control plants were treated with water spray. The (treatment) plants were irrigated once in two days, once in four days and the control plants were irrigated daily. The experiment was conducted at the Al Maqam campus of the U.A.E. University, Al Ain. The experimental design was completely randomized with the two factors being irrigation and chemical treatment, consisting of nine treatment combinations, replicated three times. The relative water content (RWC) in both concentrations was similar, while the control plants showed high plant-water content. Photosynthetic pigments viz., Chlorophyll a, increased with increasing concentration of the chemical, while the Chlorophyll b and Total Chlorophyll showed significant differences with different treatment combinations of irrigation frequencies and triadimefon treatment. Combination of triadimefon with irrigation frequencies produced sun-type chloroplasts and increased chlorophyll accumulation. Compared with the control, the stomata from the abaxial epidermal peels in the triadimefon treated leaves were sunken, narrow and appeared partially closed. Both triadimefon treatments increased the number of trichomes compared to the leaf samples of control plants. Formation of epicuticular wax in treated plants was noticed, which was probably formed to act as a barrier against water loss. The results of the present study may add to the plant growth regulatory effects data published previously on the synthesis of photosynthetic pigments, and other growth responses conducive for protection against moisture stress.

Accumulation of photosynthesis gene transcripts in response to sodium chloride by salt-tolerant alfalfa cells

A cell line from diploid alfalfa (Medicago sativa L.), HG2-N1, which is stably tolerant to 1% (0.171 M) NaCl in the medium and noticeably more green than the parent line (HG2), was investigated to determine if increased chlorophyll accumulation was related to chloroplast development toward greater photosynthetic activity and increases in chloroplast-gene expression in this cell line. The ctDNA (chloroplast-DNA) copy number in HG2-N1 was found to be increased by 50% from that of the salt-sensitive parent HG2 line. RNA accumulation, as detected by hybridization to a nuclear actin gene, appeared comparable in HG2 and HG2-N1. In contrast, mRNA levels of the plastid-encoded psbA gene were increased in the salt-tolerant HG2-N1 grown on normal medium. The mRNA levels of HG2-N1 were further increased in response to added NaCl in the medium. The mRNA levels from other chloroplast genes necessary for photosynthesis (psbD, psaB, atpB, rbcL), as well as from several nuclear genes (pCab4, pCab1, rbcS) encoding polypeptides participating in photosynthesis, also increased in the salt-tolerant HG2-N1 when it was grown in the presence of NaCl whereas actin-mRNA and chloroplast-rRNA levels remained comparable under growth conditions ± NaCl. Several independently isolated salt-tolerant cell lines showed a similar response to salt, as monitored by accumulation of rbcL mRNA, indicating that transcript accumulation for photosynthesis genes is correlated with the salt response in these salt-tolerant alfalfa cell lines.

Promotion of plant growth by polymers of lactic acid

Polymers of L-lactic acid are shown to promote plant growth. Dry weight of duckweed (Lemna minor L.) and corn (Zea mays L) was more than doubled when plants were grown in media containing the dimer of L-lactic acid, L-lactoyllactic acid. Higher polymers were equally effective at increasing plant biomass. Monomeric lactic acid and polymers of D-lactic acid showed no biological activity. Increased plant biomass was accompanied by increased chlorophyll accumulation and root growth. Promotion of chlorophyll accumulation and biomass may be due to increased ability to assimilate nutrients as plants treated with L-lactoyllactic acid showed no decrease in biomass when grown in medium that was growth limiting for control plants.