The NAC factor LpNAL delays leaf senescence by repressing two chlorophyll catabolic genes in perennial ryegrass.

Expression of chlorophyll (Chl) catabolic genes during leaf senescence is tightly controlled at the transcriptional level. Here, we identified a NAC family transcription factor, LpNAL, involved in regulating Chl catabolic genes via the yeast one-hybrid system based on truncated promoter analysis of STAYGREEN (LpSGR) in perennial ryegrass (Lolium perenne L.). LpNAL was found to be a transcriptional repressor, directly repressing LpSGR as well as the Chl b reductase gene, NONYELLOWING COLORING1. Perennial ryegrass plants over-expressing LpNAL exhibited delayed leaf senescence or stay-green phenotypes, whereas knocking down LpNAL using RNA interference accelerated leaf senescence. Comparative transcriptome analysis of leaves at 30 d after emergence in wild-type, LpNAL-overexpression, and knock-down transgenic plants revealed that LpNAL-regulated stay-green phenotypes possess altered light reactions of photosynthesis, antioxidant metabolism, ABA and ethylene synthesis and signaling, and Chl catabolism. Collectively, the transcriptional repressor LpNAL targets both Chl a and Chl b catabolic genes and acts as a brake to fine-tune the rate of Chl degradation during leaf senescence in perennial ryegrass.

Rhizobacteria-enhanced drought tolerance and post-drought recovery of creeping bentgrass involving differential modulation of leaf and root metabolism.

Rhizobacteria that produce 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase (ACCd) that inhibits ethylene production may mitigate stress damages. The objectives of this study were to examine whether a novel strain of ACCd-producing bacteria, Paraburkholderia aspalathi "WSF23," promotes plant tolerance to drought stress and post-stress recovery and determine changes in metabolic profiles in leaves and roots associated with the positive ACCd-bacteria effects in cool-season perennial grass species. Creeping bentgrass (Agrostis Stolonifera L. cv. "Penncross") plants were inoculated with P. aspalathi "WSF23" and exposed to drought by withholding irrigation for 35 days, followed by re-watering for 15 days in growth chambers. Inoculated plants demonstrated increased turf quality, canopy density, and root growth during drought stress and more rapid re-growth upon re-watering. Metabolomic analysis demonstrated that inoculation with P. aspalathi "WSF 23" increased the content of metabolites in the metabolic pathways related to stress defense, including osmoregulation, cell wall stability, and antioxidant protection in both leaves and roots, as well as nitrogen metabolism in roots of creeping bentgrass exposed to drought stress. The promotion of post-stress recovery by P. aspalathi "WSF 23" was mainly associated with enhanced carbohydrate and pyrimidine metabolism and zeatin biosynthesis pathways in leaves and increased carbohydrates, biosynthesis of DNA and proteins, cellular metabolism, and TCA cycle activity in roots. These results provide insights into the metabolic pathways regulated by "WSF23," with the PGPR conferring improvements in drought stress tolerance and post-drought recovery in a perennial grass species.

Identification of expansin genes as promoting or repressing factors for leaf elongation in tall fescue.

Expansins are cell-wall loosening proteins involved in plant cell expansion and elongation. Objectives of this study were to identify expansins related to leaf elongation in a perennial grass species and determine the relationship between the expression of expansin genes and leaf elongation. A total of 20 expansin genes were identified in tall fescue (Festuca arundinacea), out of which nine genes belonged to the EXPA- and 11 to the EXPB subfamily. Two genotypes ("TF007" and "TF116") with different growth rates were used to determine the correlation between expansins and leaf growth. Among the 20 expansins, 16 were differentially expressed in the leaf growth zone in "TF007" and "TF116." The further analysis of gene expression in different leaf segments of "TF007" and "TF116" revealed that the expression level of FaEXPB16 was positively correlated with leaf elongation rate, and "TF007" had a higher leaf elongation rate than "TF116" due to the greater expression level of FaEXPB16. FaEXPA7 exhibited significantly higher expression level in leaves of the rapid-growing genotypes than the slow-growing genotypes, suggesting that FaEXPA7 acts as a positive regulator for leaf elongation. FaEXPA7 also exhibited its highest expression level in the cell division zone located in the leaf base. FaEXPB3, FaEXPB4-2, and FaEXPB11-2 showed a negative correlation with the leaf elongation rate in "TF007" and "TF116" and were highly expressed in leaves of the slow-growing genotypes. As promoting or repressing factors for leaf growth, these five expansins could be used as candidate genes in developing the rapid or slow-growing perennial grass species.

NOL-mediated functional stay-green traits in perennial ryegrass (Lolium perenne L.) involving multifaceted molecular factors and metabolic pathways regulating leaf senescence.

Loss of chlorophyll (Chl) is a hallmark of leaf senescence, which may be regulated by Chl catabolic genes, including NON-YELLOW COLORING 1 (NYC1)-like (NOL). The objective of this study was to determine molecular factors and metabolic pathways underlying NOL regulation of leaf senescence in perennial grass species. LpNOL was cloned from perennial ryegrass (Lolium perenne L.) and found to be highly expressed in senescent leaves. Transient overexpression of LpNOL accelerated leaf senescence and Chl b degradation in Nicotiana benthamiana. LpNOL RNA interference (NOLi) in perennial ryegrass not only significantly blocked Chl degradation in senescent leaves, but also delayed initiation and progression of leaf senescence. This study found that NOL, in addition to functioning as a Chl b reductase, could enact the functional stay-green phenotype in perennial grass species, as manifested by increased photosynthetic activities in NOLi plants. Comparative transcriptomic analysis revealed that NOL-mediated functional stay-green in perennial ryegrass was mainly achieved through the modulation of Chl catabolism, light harvesting for photosynthesis, photorespiration, cytochrome respiration, carbohydrate catabolism, oxidative detoxification, and abscisic acid biosynthesis and signaling pathways.

Knock down of NON-YELLOW COLOURING 1-like gene or chlorophyllin application enhanced chlorophyll accumulation with antioxidant roles in suppressing heat-induced leaf senescence in perennial ryegrass.

Loss of chlorophyll and oxidative damage co-occur during heat-induced leaf senescence. This study aimed to determine the functions of a chlorophyll catabolic gene, NON-YELLOW COLOURING 1 (NYC1)-like (NOL), in regulating heat-induced leaf senescence and to characterize antioxidant roles of a chlorophyll derivative, sodium copper chlorophyllin (SCC), in suppressing heat-induced leaf senescence. In two separate experiments, one by comparing NOL RNAi transgenic and wild-type plants, and the other by analysing the effects of SCC treatment, perennial ryegrass (Lolium perenne) was exposed to heat stress (38/35 °C, day/night) or optimal temperature (25/20 °C). Results showed that both knock down of LpNOL and application of SCC suppressed heat-induced leaf senescence, as manifested by increased chlorophyll content, reduced electrolyte leakage, down-regulation of chlorophyll-catabolic genes and senescence-related genes, as well as enhanced antioxidant capacity in the peroxidase pathway for H2O2 scavenging. Ex vivo SCC incubation protected membranes from H2O2 damage in mesophyll protoplasts of perennial ryegrass. The suppression of leaf senescence by knocking down NOL or chlorophyllin application was associated with enhanced chlorophyll accumulation playing antioxidant roles in protecting leaves from heat-induced oxidative damage.

Cytokinin or ethylene regulation of heat-induced leaf senescence involving transcriptional modulation of WRKY in perennial ryegrass.

Heat stress is a major abiotic stress for temperate plant species with characteristic symptoms of premature leaf senescence. The objectives of this study were to evaluate the physiological effects of cytokinins (CK) and an ethylene inhibitor, aminoethoxyvinylglycine (AVG) on heat-induced leaf senescence in the temperate perennial grass species, perennial ryegrass (Lolium perenne), and to investigate whether WRKY transcription factors (TFs) could be associated with CK- or ethylene-mediated regulation of heat-induced leaf senescence by exogenously applying CK or AVG to perennial ryegrass. Perennial ryegrass plants foliar-sprayed with 6-benzylaminopurine (6-BA), and AVG exhibited prolonged stay-green phenotypes and a lesser degree of leaf senescence under heat stress (35/30°C), as shown by a decline in electrolyte leakage, malondialdehyde content, hydrogen peroxide, and superoxide content, and increased chlorophyll (Chl) content along with reduced activities of Chl-degrading enzymes (pheophytinase and chlorophyllase) and increased activity of Chl-synthesizing enzyme (porphobilinogen deaminase) due to 6-BA or AVG application. The suppression of heat-induced leaf senescence by 6-BA or AVG treatment corresponded with the upregulation of LpWRKY69 and LpWRKY70. The LpWRKY69 and LpWRKY70 promoters were predicted to share conserved cis-elements potentially recognized by TFs in the CK or ethylene pathways. These results indicate that LpWRKY69 and LpWRKY70 may negatively regulate heat-induced leaf senescence through CK or ethylene pathways, conferring heat tolerance in perennial ryegrass.

Choline-Mediated Lipid Reprogramming as a Dominant Salt Tolerance Mechanism in Grass Species Lacking Glycine Betaine.

Choline, as a precursor of glycine betaine (GB) and phospholipids, is known to play roles in plant tolerance to salt stress, but the downstream metabolic pathways regulated by choline conferring salt tolerance are still unclear for non-GB-accumulating species. The objectives were to examine how choline affects salt tolerance in a non-GB-accumulating grass species and to determine major metabolic pathways of choline regulating salt tolerance involving GB or lipid metabolism. Kentucky bluegrass (Poa pratensis) plants were subjected to salt stress (100 mM NaCl) with or without foliar application of choline chloride (1 mM) in a growth chamber. Choline or GB alone and the combined application increased leaf photochemical efficiency, relative water content and osmotic adjustment and reduced leaf electrolyte leakage. Choline application had no effects on the endogenous GB content and GB synthesis genes did not show responses to choline under nonstress and salt stress conditions. GB was not detected in Kentucky bluegrass leaves. Lipidomic analysis revealed an increase in the content of monogalactosyl diacylglycerol, phosphatidylcholine and phosphatidylethanolamine and a decrease in the phosphatidic acid content by choline application in plants exposed to salt stress. Choline-mediated lipid reprogramming could function as a dominant salt tolerance mechanism in non-GB-accumulating grass species.

LpNOL-knockdown suppression of heat-induced leaf senescence in perennial ryegrass involving regulation of amino acid and organic acid metabolism.

The nonyellow COLORING 1-like gene (NOL) is known for its roles in accelerating leaf senescence, but the underlying metabolic mechanisms for heat-induced leaf senescence remain unclear. The objectives of this study were to identify metabolites and associated metabolic pathways regulated by knockdown of NOL in perennial ryegrass (Lolium perenne) and to determine the metabolic mechanisms of NOL controlling heat-induced leaf senescence. Wild-type (WT; cv. "Pinnacle") and two lines (Noli-1 and Noli-2) of perennial ryegrass with LpNOL knockdown were exposed to heat stress at 35/33°C (day/night) or nonstress control temperatures at 25/22°C (day/night) for 30 days in growth chambers. Leaf electrolyte leakage, chlorophyll (Chl) content, photochemical efficiency (Fv /Fm ), and net photosynthetic rate (Pn) were measured as physiological indicators of leaf senescence, while gas chromatography-mass spectrometry was performed to identify metabolites regulated by LpNOL. Knockdown of LpNOL suppressed heat-induced leaf senescence and produced a stay-green phenotype in perennial ryegrass, as manifested by increased Chl content, photochemical efficiency, net photosynthetic rate, and cell membrane stability in Noli-1 and Noli-2. Five metabolites (valine, malic acid, threonic acid, shikimic acid, chlorogenic acid) were uniquely upregulated in LpNOL plants exposed to heat stress, and six metabolites (aspartic acid, glutamic acid, 5-oxoproline, phenylalanine, proline, tartaric acid) exhibited more pronounced increases in their content in LpNOL plants than the WT. LpNOL could regulate heat-induced leaf senescence in perennial ryegrass through metabolic reprogramming in the pathways of respiration, secondary metabolism, antioxidant metabolism, and protein synthesis.

Natural variation of physiological traits, molecular markers, and chlorophyll catabolic genes associated with heat tolerance in perennial ryegrass accessions.

BACKGROUND: Identification of genetic diversity in heat tolerance and associated traits is of great importance for improving heat tolerance in cool-season grass species. The objectives of this study were to determine genetic variations in heat tolerance associated with phenotypic and physiological traits and to identify molecular markers associated with heat tolerance in a diverse collection of perennial ryegrass (Lolium perenne L.). RESULTS: Plants of 98 accessions were subjected to heat stress (35/30 °C, day/night) or optimal growth temperature (25/20 °C) for 24 d in growth chambers. Overall heat tolerance of those accessions was ranked by principal component analysis (PCA) based on eight phenotypic and physiological traits. Among these traits, electrolyte leakage (EL), chlorophyll content (Chl), relative water content (RWC) had high correlation coefficients (- 0.858, 0.769, and 0.764, respectively) with the PCA ranking of heat tolerance. We also found expression levels of four Chl catabolic genes (CCGs), including LpNYC1, LpNOL, LpSGR, and LpPPH, were significant higher in heat sensitive ryegrass accessions then heat tolerant ones under heat stress. Furthermore, 66 pairs of simple sequence repeat (SSR) markers were used to perform association analysis based on the PCA result. The population structure of ryegrass can be grouped into three clusters, and accessions in cluster C were relatively more heat tolerant than those in cluster A and B. SSR markers significantly associated with above-mentioned traits were identified (R2 > 0.05, p < 0.01)., including two pairs of markers located on chromosome 4 in association with Chl content and another four pairs of markers in association with EL. CONCLUSION: The result not only identified useful physiological parameters, including EL, Chl content, and RWC, and their associated SSR markers for heat-tolerance breeding of perennial ryegrass, but also highlighted the involvement of Chl catabolism in ryegrass heat tolerance. Such knowledge is of significance for heat-tolerance breeding and heat tolerance mechanisms in perennial ryegrass as well as in other cool-season grass species.

Up-regulation of lipid metabolism and glycine betaine synthesis are associated with choline-induced salt tolerance in halophytic seashore paspalum.

Choline may affect salt tolerance by regulating lipid and glycine betaine (GB) metabolism. This study was conducted to determine whether alteration of lipid profiles and GB metabolism may contribute to choline regulation and genotypic variations in salt tolerance in a halophytic grass, seashore paspalum (Paspalum vaginatum). Plants of Adalayd and Sea Isle 2000 were subjected to salt stress (200-mM NaCl) with or without foliar application of choline chloride (1 mM). Genotypic variations in salt tolerance and promotive effects of choline application on salt tolerance were associated with both the up-regulation of lipid metabolism and GB synthesis. The genotypic variations in salt tolerance associated with lipid metabolism were reflected by the differential accumulation of phosphatidylcholine and phosphatidylethanolamine between Adalayd and Sea Isle 2000. Choline-induced salt tolerance was associated with of the increase in digalactosyl diacylglycerol (DGDG) content including DGDG (36:4 and 36:6) in both cultivars of seashore paspalum and enhanced synthesis of phosphatidylinositol (34:2, 36:5, and 36:2) and phosphatidic acid (34:2, 34:1, and 36:5), as well as increases in the ratio of digalactosyl diacylglycerol: monogalactosyl diacylglycerol (DGDG:MGDG) in salt-tolerant Sea Isle 2000. Choline regulation of salt tolerance may be due to the alteration in lipid metabolism in this halophytic grass species.

Differential regulatory pathways associated with drought-inhibition and post-drought recuperation of rhizome development in perennial grass.

BACKGROUND AND AIMS: Rhizomes are key organs for the establishment of perennial grass stands and adaptation to environmental stress. However, mechanisms regulating rhizome initiation and elongation under drought stress and during post-drought recovery remain unclear. The objective of this study is to investigate molecular factors and metabolic processes involved in drought effects and post-drought recovery in rhizome growth in perennial grass species by comparative transcriptomic and proteomic profiling. METHODS: Tall fescue (Festuca arundinacea) (B-type rhizome genotype, 'BR') plants were exposed to drought stress and re-watering in growth chambers. The number and length of rhizomes were measured following drought stress and re-watering. Hormone and sugar contents were analysed, and transcriptomic and proteomic analyses were performed to identify metabolic factors, genes and proteins associated with rhizome development. KEY RESULTS: Rhizome initiation and elongation were inhibited by drought stress, and were associated with increases in the contents of abscisic acid (ABA) and soluble sugars, but declines in the contents of indoleacetic acid (IAA), zeatin riboside (ZR) and gibberellin (GA4). Genes involved in multiple metabolic processes and stress defence systems related to rhizome initiation exhibited different responses to drought stress, including ABA signalling, energy metabolism and stress protection. Drought-inhibition of rhizome elongation could be mainly associated with the alteration of GA4 and antioxidants contents, energy metabolism and stress response proteins. Upon re-watering, new rhizomes were regenerated from rhizome nodes previously exposed to drought stress, which was accompanied by the decline in ABA content and increases in IAA, ZR and GA4, as well as genes and proteins for auxin, lipids, lignin and nitrogen metabolism. CONCLUSIONS: Drought-inhibition of rhizome initiation and elongation in tall fescue was mainly associated with adjustments in hormone metabolism, carbohydrate metabolism and stress-defence systems. Rhizome regeneration in response to re-watering involved reactivation of hormone and lipid metabolism, secondary cell-wall development, and nitrogen remobilization and cycling.

Proteomic Profiling for Metabolic Pathways Involved in Interactive Effects of Elevated Carbon Dioxide and Nitrogen on Leaf Growth in a Perennial Grass Species.

Elevated atmospheric CO2 and nitrogen are major environmental factors affecting shoot growth. The objectives of this study are to determine the interactive effects of elevated CO2 and nitrogen on leaf growth in tall fescue ( Festuca arundinacea) and to identify major proteins and associated metabolic pathways underlying CO2-regulation of leaf growth under insufficient and sufficient nitrate conditions using proteomic analysis. Plants of tall fescue treated with low nitrate level (0.25 mM, LN), moderate nitrate level (4 mM, MN) and high nitrate level (32 mM, HN) were exposed to ambient (400 µmol mol-1) and elevated (800 µmol mol-1) CO2 concentrations in environment-controlled growth chambers. Increased atmospheric CO2 concentration increased leaf length and shoot biomass, which corresponded to increased content of indo-acetic acid, gibberellic acid, cytokinins and reduced content of abscisic acid under sufficient nitrate conditions (MN and HN conditions). Low nitrate supply limited shoot growth and hormonal responses to elevated CO2. Proteomic analysis of plants exposed to elevated CO2 under LN and MN conditions demonstrated the increases in the abundance of many proteins due to elevated CO2 under MN condition involved with cell cycle and proliferation, transcription and translation, photosynthesis (ribosomal and chlorophyll a/b-binding proteins), amino acids synthesis, sucrose and starch metabolism, as well as ABA signaling pathways (ABA-induced proteins). Our results revealed major proteins and associated metabolic pathways associated with the interactive effects of elevated CO2 and nitrate regulating leaf growth in a perennial grass species.

Knockdown of STAYGREEN in Perennial Ryegrass (Lolium perenne L.) Leads to Transcriptomic Alterations Related to Suppressed Leaf Senescence and Improved Forage Quality.

Chl breakdown is a hallmark of leaf senescence. Protein degradation is tightly associated with accelerated Chl catabolism during leaf senescence. Therefore, blocking or reducing Chl breakdown and thereby improving Chl and leaf protein contents is desirable for agronomic improvement in perennial forage grasses. Perennial ryegrass (Lolium perenne L.) is one principle cool-season forage grass in temperate areas throughout the world. In this study, the perennial ryegrass STAY-GREEN gene (LpSGR) was cloned and characterized. LpSGR was highly expressed in developmentally or dark-induced senescent leaves. LpSGR was subcellularly localized in chloroplast and interacted with the other Chl catabolic enzymes. RNA interference (RNAi) of LpSGR in perennial ryegrass blocked the degradation of Chl, resulting in increased Chl content and photochemical efficiency in senescent leaves. The RNAi transgenic plants had significantly improved forage quality, with up to 46.1% increased protein content in the harvested biomass. Transcriptome comparison revealed that suppression of LpSGR led to multiple alterations in metabolic pathways in locations inside the chloroplast. Most transcription factors of senescence-associated hormonal signaling pathways (e.g. ABA, ethylene and jasmonic acid) had decreased expression levels in the RNAi plants. These results provided a foundation for the further study on the regulatory mechanism of LpSGR in perennial ryegrass for the purpose of forage improvement with delayed leaf senescence and higher forage quality.

Lipidomic reprogramming associated with drought stress priming-enhanced heat tolerance in tall fescue (Festuca arundinacea).

Stress priming by exposing plants to a mild or moderate drought could enhance plant tolerance to subsequent heat stress. Lipids play vital roles in stress adaptation, but how lipidomic profiles change, affecting the cross-stress tolerance, is largely unknown. The objectives of this study were to perform lipidomics, to analyse the content, composition, and saturation levels of lipids in leaves of tall fescue (Festuca arundinacea) following drought priming and subsequent heat stress, and to identify major lipids and molecular species associated with priming-enhanced heat tolerance. Plants were initially exposed to drought for 8 days by withholding irrigation and subsequently subjected to 25 days of heat stress (38/33°C day/night) in growth chambers. Drought-primed plants maintained significantly higher leaf relative water content, chlorophyll content, photochemical efficiency, and lower electrolyte leakage than nonprimed plants under heat stress. Drought priming enhanced the accumulation of phospholipids and glycolipids involved in membrane stabilization and stress signalling (phosphatidic acid, phosphatidylcholine, phosphatidylinositol, phosphatidylglycerol, and digalactosyl diacylglycerol) during subsequent exposure to heat stress. The reprogramming of lipid metabolism for membrane stabilization and signalling in response to drought priming and subsequent exposure to heat stress could contribute to drought priming-enhanced heat tolerance in cool-season grass species.

Abscisic acid mediation of drought priming-enhanced heat tolerance in tall fescue (Festuca arundinacea) and Arabidopsis.

Abscisic acid (ABA) may play roles in mediating cross stress tolerance in plants. The objectives of this study were to investigate the priming effects of drought and ABA on heat tolerance and to determine how ABA may be involved in enhanced heat tolerance by drought. Focusing on the transcriptional level, two independent experiments were conducted, using a perennial grass species, tall fescue (Festuca arundinacea) and Arabidopsis. In experiment 1, tall fescue plants were exposed to mild drought by withholding irrigation for 8 days (drought priming) and foliar sprayed with ABA or an ABA-synthesis inhibitor (fluridone). After that they were subsequently subjected to heat stress (38/33°C day/night) for 25 days in growth chambers. In experiment 2, Arabidopsis Columbia ecotype (wild-type) and ABA-deficient mutant (aba3-1, CS157) were pre-treated with drought priming and then exposed to heat stress (45/40°C) for 3 days. The physiological analysis demonstrated that both drought priming and foliar application of ABA-enhanced heat tolerance in tall fescue, while drought priming had no significant effects on heat tolerance in ABA-deficient Arabidopsis plants. Application of fluridone to tall fescue and ABA-deficient mutants of Arabidopsis exhibited diminished or attenuated positive effects of drought priming on heat tolerance. ABA mediation of acquired heat tolerance by drought priming was associated with the upregulation of CDPK3, MPK3, DREB2A, AREB3, MYB2, MYC4, HsfA2, HSP18, and HSP70. Our study revealed the roles of ABA in drought priming-enhanced heat tolerance, which may involve transcriptional regulation for stress signaling, ABA responses and heat protection.

Strigolactones Promote Leaf Elongation in Tall Fescue through Upregulation of Cell Cycle Genes and Downregulation of Auxin Transport Genes in Tall Fescue under Different Temperature Regimes.

Strigolactones (SLs) have recently been shown to play roles in modulating plant architecture and improving plant tolerance to multiple stresses, but the underlying mechanisms for SLs regulating leaf elongation and the influence by air temperature are still unknown. This study aimed to investigate the effects of SLs on leaf elongation in tall fescue (Festuca arundinacea, cv. 'Kentucky-31') under different temperature regimes, and to determine the interactions of SLs and auxin in the regulation of leaf growth. Tall fescue plants were treated with GR24 (synthetic analog of SLs), naphthaleneacetic acid (NAA, synthetic analog), or N-1-naphthylphthalamic acid (NPA, auxin transport inhibitor) (individually and combined) under normal temperature (22/18 °C) and high-temperature conditions (35/30 °C) in controlled-environment growth chambers. Exogenous application of GR24 stimulated leaf elongation and mitigated the heat inhibition of leaf growth in tall fescue. GR24-induced leaf elongation was associated with an increase in cell numbers, upregulated expression of cell-cycle-related genes, and downregulated expression of auxin transport-related genes in elongating leaves. The results suggest that SLs enhance leaf elongation by stimulating cell division and interference with auxin transport in tall fescue.

Transcriptomic analysis reveals unique molecular factors for lipid hydrolysis, secondary cell-walls and oxidative protection associated with thermotolerance in perennial grass.

BACKGROUND: Heat stress is the primary abiotic stress limiting growth of cool-season grass species. The objective of this study was to determine molecular factors and metabolic pathways associated with superior heat tolerance in thermal bentgrass (Agrostis scabra) by comparative analysis of transcriptomic profiles with its co-generic heat-sensitive species creeping bentgrass (A. stolonifera). RESULTS: Transcriptomic profiling by RNA-seq in both heat-sensitive A. stolonifera (cv. 'Penncross') and heat-tolerant A. scabra exposed to heat stress found 1393 (675 up- and 718 down-regulated) and 1508 (777 up- and 731 down-regulated) differentially-expressed genes, respectively. The superior heat tolerance in A. scabra was associated with more up-regulation of genes in oxidative protection, proline biosynthesis, lipid hydrolysis, hemicellulose and lignin biosynthesis, compared to heat-sensitive A. stolonifera. Several transcriptional factors (TFs), such as high mobility group B protein 7 (HMGB7), dehydration-responsive element-binding factor 1a (DREB1a), multiprotein-bridging factor 1c (MBF1c), CCCH-domain containing protein 47 (CCCH47), were also found to be up-regulated in A. scabra under heat stress. CONCLUSIONS: The unique TFs and genes identified in thermal A. scabra could be potential candidate genes for genetic modification of cultivated grass species for improving heat tolerance, and the associated pathways could contribute to the transcriptional regulation for superior heat tolerance in bentgrass species.

Characterization of Dehydrin protein, CdDHN4-L and CdDHN4-S, and their differential protective roles against abiotic stress in vitro.

BACKGROUND: Dehydrins play positive roles in regulating plant abiotic stress responses. The objective of this study was to characterize two dehydrin genes, CdDHN4-L and CdDHN4-S, generated by alternative splicing of CdDHN4 in bermudagrass. RESULTS: Overexpression of CdDHN4-L with φ-segment and CdDHN4-S lacking of φ-segment in Arabidopsis significantly increased tolerance against abiotic stresses. The growth phenotype of Arabidopsis exposed to NaCl at 100 mM was better in plants overexpressing CdDHN4-L than those overexpressing CdDHN4-S, as well as better in E.coli cells overexpressing CdDHN4-L than those overexpressing CdDHN4-S in 300 and 400 mM NaCl, and under extreme temperature conditions at - 20 °C and 50 °C. The CdDHN4-L had higher disordered characterization on structures than CdDHN4-S at temperatures from 10 to 90 °C. The recovery activities of lactic dehydrogenase (LDH) and alcohol dehydrogenase (ADH) in presence of CdDHN4-L and CdDHN4-S were higher than that of LDH and ADH alone under freeze-thaw damage and heat. Protein-binding and bimolecular fluorescence complementation showed that both proteins could bind to proteins with positive isoelectric point via electrostatic forces. CONCLUSIONS: These results indicate that CdDHN4-L has higher protective ability against abiotic stresses due to its higher flexible unfolded structure and thermostability in comparison with CdDHN4-S. These provided direct evidence of the function of the φ-segment in dehydrins for protecting plants against abiotic stress and to show the electrostatic interaction between dehydrins and client proteins.

The optimal CO2 concentrations for the growth of three perennial grass species.

BACKGROUND: Grasslands are one of the most representative vegetation types accounting for about 20% of the global land area and thus the response of grasslands to climate change plays a pivotal role in terrestrial carbon balance. However, many current climate change models, based on earlier results of the doubling-CO2 experiments, may overestimate the CO2 fertilization effect, and as a result underestimate the potentially effects of future climate change on global grasslands when the atmospheric CO2 concentration goes beyond the optimal level. Here, we examined the optimal atmospheric CO2 concentration effect on CO2 fertilization and further on the growth of three perennial grasses in growth chambers with the CO2 concentration at 400, 600, 800, 1000, and 1200 ppm, respectively. RESULTS: All three perennial grasses featured an apparent optimal CO2 concentration for growth. Initial increases in atmospheric CO2 concentration substantially enhanced the plant biomass of the three perennial grasses through the CO2 fertilization effect, but this CO2 fertilization effect was dramatically compromised with further rising atmospheric CO2 concentration beyond the optimum. The optimal CO2 concentration for the growth of tall fescue was lower than those of perennial ryegrass and Kentucky bluegrass, and thus the CO2 fertilization effect on tall fescue disappeared earlier than the other two species. By contrast, the weaker CO2 fertilization effect on the growth of perennial ryegrass and Kentucky bluegrass was sustained for a longer period due to their higher optimal CO2 concentrations than tall fescue. The limiting effects of excessively high CO2 concentrations may not only associate with changes in the biochemical and photochemical processes of photosynthesis, but also attribute to the declines in stomatal conductance and nitrogen availability. CONCLUSIONS: In this study, we found apparent differences in the optimal CO2 concentrations for the growth of three grasses. These results suggest that the growth of different types of grasses may respond differently to future elevated CO2 concentrations through the CO2 fertilization effect, and thus potentially alter the community composition and structure of grasslands. Meanwhile, our results may also be helpful for improving current process-based ecological models to more accurately predict the structure and function of grassland ecosystems under future rising atmospheric CO2 concentration and climate change scenarios.

Switchgrass PvDREB1C plays opposite roles in plant cold and salt tolerance in transgenic tobacco.

BACKGROUND: The C-repeat-binding factors/DRE-binding factors (CBF/DREBs) comprise a key transcription factor family involved in plant stress tolerance. Yet, there is limited information about switchgrass DREB genes and their functional roles. RESULTS: In this study, four cold-inducible PvDREB1s were identified from switchgrass (Panicum virgatum), among which PvDREB1C was the one responded to cold stress later than the other three PvDREB1s. Yet, ectopic overexpression of PvDREB1C led to significantly compromised, instead of improved cold tolerance in transgenic tobacco. On the other hand, PvDREB1C was transcriptionally down-regulated in response to salt stress, but overexpression of PvDREB1C improved plant salt tolerance in transgenic tobacco. The improved salt tolerance was associated with increased K+/Na+ ratio and Ca2+ content, higher cellular osmotic potential, and activation of stress-related functional genes in the leaves of transgenic plants under salt stress. CONCLUSIONS: The current results implied that PvDREB1C played opposite roles in plant cold and salt tolerance. Although DREB1s were known as positive stress regulators, particular attentions shall be paid to their potential negative regulatory role(s).