The A subunit of vacuolar H+-ATPase gene (CmVHA-A) plays opposite roles in plant growth and drought tolerance of chrysanthemum under different growing conditions.

As the most prominent proton pumps in plants, vacuolar H+-ATPases (VHAs) comprise multiple subunits that are important for physiological processes and stress tolerance in plants. However, few studies on the roles of subunit genes of VHAs in chrysanthemum have been reported to date. In this study, the gene of A subunit of V-ATPase in chrysanthemum (CmVHA-A) was cloned and identified. CmVHA-A was conserved with VHA-A proteins from other plants. Expression analysis showed that CmVHA-A was highly expressed in most tissues of chrysanthemum except for the flower bud, and was readily induced by polyethylene glycol (PEG) treatment. Functional analysis demonstrated that CmVHA-A exerted a negative influence on the growth and development of shoot and root of chrysanthemum under normal conditions. RNA-sequencing (RNA-seq) analysis revealed the possible explanations for phenotypic differences between transgenic and wild-type (WT) plants. Under drought conditions, CmVHA-A positively affected the drought tolerance of chrysanthemum by enhancing antioxidase activity and alleviating photosynthetic disruption. Overall, CmVHA-A plays opposite roles in plant growth and drought tolerance of chrysanthemums under different growing conditions.

Effects of exogenous melatonin on photosynthesis and physiological characteristics of chry-santhemum seedlings under high temperature stress.

We examined the effects of exogenous melatonin (MT) on the resistance of Chrysanthemum morifolium 'Jinba' to high temperature stress. Chrysanthemum leaves were sprayed with 200 µmol·L-1MT, and then subjected to high temperature stress at 40 ℃ (day)/ 35 ℃ (night). The ultrastructure of chloroplast and thylakoid of chrysanthemum leaves were observed, and the photosynthetic and physiological indices were measured. The results showed that the chloroplast and thyla-koid structures of chrysanthemum were damaged under high temperature stress. The chlorophyll contents and maximum fluorescence (Fm) were significantly reduced, while the OJIP curve changed with the fluorescence of K and J points increased. The net photosynthetic rate (Pn), transpiration rate (Tr) and stomatal conductance (gs) were significantly decreased, while the internal CO2 concentration (Ci) was significantly increased. The relative conductivity (REC), malondialdehyde (MDA), reactive oxygen species (ROS), osmotic adjustment substances content and antioxidant enzyme activity all increased significantly. Spraying exogenous MT onto leaves could maintain the integrity of chloroplast and thylakoid structure under high temperature in chrysanthemum and significantly decrease the increment in the K and J points of OJIP curve. Exogenous application of MT alleviated the inhibition of high temperature stress on photosynthesis and fluorescence of chrysanthemum, as indicated by significantly higher Fm, Pn, gs, Tr and photosynthetic pigment contents and lower Ci. Exogenous MT also significantly reduced the REC, MDA and ROS contents of chrysanthemum under high temperature stress, and enhanced the osmotic adjustment substances content and antioxidant enzyme activity in chrysanthemum leaves. It suggested that exogenous MT could protect the integrity of chloroplast structure of chrysanthemum leaves, enhance photosynthesis, inhibit the excessive production of ROS in the plants under high temperature stress, improve the activity of antioxidant enzyme system, reduce the level of membrane peroxidation and keep the integrity of lipid membrane, and thus improve the ability of chrysanthemum plants to resist high temperature stress.

New insights into the role of MADS-box transcription factor gene CmANR1 on root and shoot development in chrysanthemum (Chrysanthemum morifolium).

BACKGROUND: MADS-box transcription factors (TFs) are the key regulators of multiple developmental processes in plants; among them, a chrysanthemum MADS-box TF CmANR1 has been isolated and described as functioning in root development in response to high nitrate concentration signals. However, how CmANR1 affects root and shoot development remains unclear. RESULTS: We report that CmANR1 plays a positive role in root system development in chrysanthemum throughout the developmental stages of in vitro tissue cultures. Metabolomics combined with transcriptomics assays show that CmANR1 promotes robust root system development by facilitating nitrate assimilation, and influencing the metabolic pathways of amino acid, glycolysis, and the tricarboxylic acid cycle (TCA) cycle. Also, we found that the expression levels of TFs associated with the nitrate signaling pathways, such as AGL8, AGL21, and LBD29, are significantly up-regulated in CmANR1-transgenic plants relative to the wild-type (WT) control; by contrast, the expression levels of RHD3-LIKE, LBD37, and GATA23 were significantly down-regulated. These results suggest that these nitrate signaling associated TFs are involved in CmANR1-modulated control of root development. In addition, CmANR1 also acts as a positive regulator to control shoot growth and development. CONCLUSIONS: These findings provide potential mechanisms of MADS-box TF CmANR1 modulation of root and shoot development, which occurs by regulating a series of nitrate signaling associated TFs, and influencing the metabolic pathways of amino acid and glycolysis, as well as TCA cycle and nitrate assimilation.

Proteomic profiling of root system development proteins in chrysanthemum overexpressing the CmTCP20 gene.

Plant root systems ensure the efficient absorption of water and nutrients and provide anchoring into the soil. Although root systems are a highly plastic set of traits that vary both between and among species, the basic root system morphology is controlled by inherent genetic factors. TCP20 has been identified as a key regulator of root development in plants, and yet its underlying mechanism has not been fully elucidated, especially in chrysanthemum. We found that overexpression of the CmTCP20 gene promoted both adventitious and lateral root development in chrysanthemum. To get further insight into the molecular mechanisms controlling root system development, we conducted a study employing tandem mass tag proteomic to characterize the differential root system development proteomes from CmTCP20-overexpressing and wild-type chrysanthemum root samples. Of the proteins identified, 234 proteins were found to be differentially abundant (>1.5-fold cut off, p < 0.05) in CmTCP20-overexpressing versus wild-type chrysanthemum root samples. Functional enrichment analysis indicated that the CmTCP20 gene may participate in "phytohormone signal transduction". Our findings provide a valuable perspective on the mechanisms of both adventitious and lateral root development via CmTCP20 modulation at the proteome level in chrysanthemum.

CmTCP20 Plays a Key Role in Nitrate and Auxin Signaling-Regulated Lateral Root Development in Chrysanthemum.

Lateral root (LR) formation and development play a vital role in plant development by permitting the establishment of branched root systems. It is well known that nutrient availability controls LR development. Moreover, LR development is fine-tuned by a myriad of hormonal signals. Many transcription factors (TFs) participate in LR development. Here, we discuss the TFs involved in the nitrate and auxin signaling pathways and how these function in the regulation of LR formation and development in chrysanthemum. AtTCP20 is a plant-specific TF, which can modulate LR development in response to nitrate. The roles of CmTCP20 in LR development were identified by overexpression in chrysanthemum and heterologous expression in Arabidopsis. Overexpression of CmTCP20 significantly increased the number and average length of LRs compared with the wild type in chrysanthemum and Arabidopsis. We also found that CmTCP20 positively influenced auxin accumulation in the LRs at least partly by improving auxin biosynthesis, transport and response, thereby promoting LR development. Moreover, we found that CmTCP20 interacts with an auxin response factor, CmARF8, which also can be induced by nitrate and combined to proximal sites in the upstream promoter region of CmCYCB1;1 to positively regulate the cell cycle. The CmTCP20-CmARF8 heterodimer links nitrate and auxin signaling and converts cell-cycle signals to regulate LR initiation and growth.

The MADS transcription factor CmANR1 positively modulates root system development by directly regulating CmPIN2 in chrysanthemum.

Plant root systems are essential for many physiological processes, including water and nutrient absorption. MADS-box transcription factor (TF) genes have been characterized as the important regulators of root development in plants; however, the underlying mechanism is largely unknown, including chrysanthemum. Here, it was found that the overexpression of CmANR1, a chrysanthemum MADS-box TF gene, promoted both adventitious root (AR) and lateral root (LR) development in chrysanthemum. Whole transcriptome sequencing analysis revealed a series of differentially expressed unigenes (DEGs) in the roots of CmANR1-transgenic chrysanthemum plants compared to wild-type plants. Functional annotation of these DEGs by alignment with Gene Ontology (GO) terms and biochemical pathway Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that CmANR1 TF exhibited "DNA binding" and "catalytic" activity, as well as participated in "phytohormone signal transduction". Both chromatin immunoprecipitation-polymerase chain reaction (ChIP-PCR) and gel electrophoresis mobility shift assays (EMSA) indicated the direct binding of CmPIN2 to the recognition site CArG-box motif by CmANR1. Finally, a firefly luciferase imaging assay demonstrated the transcriptional activation of CmPIN2 by CmANR1 in vivo. Overall, our results provide novel insights into the mechanisms of MADS-box TF CmANR1 modulation of both AR and LR development, which occurs by directly regulating auxin transport gene CmPIN2 in chrysanthemum.

Chrysanthemum MADS-box transcription factor CmANR1 modulates lateral root development via homo-/heterodimerization to influence auxin accumulation in Arabidopsis.

Root system architecture is an important agronomic trait by which plants both acquire water and nutrients from the soil and adapt to survive in a complex environment. The adaptation of plant root systems to environmental constraints largely depends on the growth and development of lateral roots (LRs). MADS-box transcription factors (TFs) are important known regulators of plant growth, development, and response to environmental stimuli. However, the potential mechanisms by which they regulate LRs development remain poorly understood. Here, we identified a MADS-box chrysanthemum gene CmANR1, homologous to the Arabidopsis gene AtANR1, which plays a key role in the regulation of LR development. qRT-PCR assays indicated that CmANR1 was primarily expressed in chrysanthemum roots and was rapidly induced by exposure to high nitrate concentrations. Ectopic expression of CmANR1 in Arabidopsis significantly increased the number and length of emerged LRs compared to the wild-type (col) control, but had no obvious affect on primary root (PR) development. We also found that CmANR1 positively influenced auxin accumulation in LRs at least partly by improving auxin biosynthesis and transport, thereby promoting LR development. Furthermore, we found that ANR1 formed homo- and heterodimers through interactions with itself and AGL21 at its C-terminal domain. Overall, our findings provide considerable new information about the mechanisms by which the chrysanthemum MADS-box TF CmANR1 mediates LR development by directly altering auxin accumulation.

[Evaluation of cold resistance of four wild Carex speices]. / 四种野生苔草属植物的耐寒性评价.

Four wild Carex speices (C. rigescens, C. lancifolia, C. leucochlora and C. humilis) collected from Mount Taishan were used as experimental materials. The adaption across the winter and physiological characteristics resistant to cold stress were investigated, semi-lethal temperature (LT50) was calculated and fuzzy subordination method was used to generally evaluate the Carex resistant to cold stress. The results showed that 4 Carex species all survived in the winter safely and restored well to grow in the following spring. The green period of the species was between 260 d and 310 d, and percentage of the withered leaves was between 12% and 95%, the range of LT50 was from -18.65 ℃ to -11.74 ℃. With intensifying cold stress, the contents of MDA, proline (Pro) and soluble protein increased at first and then decreased, while the soluble sugar content increased with the treatment time. C. rigescens with poor cold tolerance showed the early accumulation of MDA, Pro and soluble sugar. The value of soluble protein peaked at the late stage of low tempe-rature stress, and the Carex with stronger cold-resistance showed the smaller value. The SOD activity in the leaves of C. lancifolia was higher than that of the other three species in the beginning of treatments. The cold resistance of four Carex species was in order of C. lancifolia>C. leucochlora>C. humilis>C. rigescens.

[Effects of nitrogen fertilization on the nitrogen uptake, accumulation, and seed quality of oil peony]. / æ–½ç”¨æ°®è‚¥å¯¹æ²¹ç”¨ç‰¡ä¸¹å¶ç‰‡æ°®ç´ å¸æ”¶ç§¯ç´¯ä¸Žç±½ç²’å“è´¨çš„å½±å“.

Field experiments, including four levels of N application 0, 18, 24, 30 g N·m-2, were carried out to clarify the effects of nitrogen fertilization on N accumulation and translocation in lea-ves as well as the seed quality of oil peony (Paeonia ostii 'Fengdan Group'). The results showed that the nitrogen application significantly increased the height, canopy, flower diameter and flower dry mass. The heights under the treatments 24 and 30 g N·m-2 increased by 14.7% and 15.2% compared with CK, respectively. Moreover, the nitrogen application improved seed yield. The highest seed yields were acquired under the treatments 24 and 30 g N·m-2, which were 15.2% and 15.4% higher than that of CK, respectively. The N accumulation and translocation in leaves and the N accumulation in seeds all increased with the nitrogen application level. The greatest leaf contribution proportion was acquired under the treatment 24 g N·m-2. The nitrogen application significantly increased the contents of protein N, total amino acid, and some saturated and unsaturated fatty acids in seeds. In this experiment, the N input of 24 g N·m-2 was optimal to obtain the higher N translocation amount, N translocation efficiency and N contribution proportion from leaves to seeds, seed yield as well as the contents of protein N, amino acid and unsaturated fatty acids.

[Effects of controlled-release fertilizer on chrysanthemum leaf chlorophyll fluorescence characteristics and ornamental quality].

Taking cut flower chrysanthemum 'Baima' as test material, a pot experiment was conducted to study the effects of controlled-release fertilizer on the leaf chlorophyll fluorescence parameters, chlorophyll and nutrient contents, and ornamental quality of chrysanthemum. Under no fertilization, the maximal photochemical efficiency of PS II in dark (F(v)/F(m)), potential photochemical efficiency of PS II (F(v)/F(0)), and quantum yield of PS II electron transport (phi(PS II)) decreased significantly, compared with those under fertilization. With the application of conventional compound fertilizers CCFA (N:P:K=20:8:10) and CCFB (N:P:K= 14:14:14), the F(v)/F(m), F(v)/F(0) and phi(PS II) had a slight increase in early period (30-60 d) but a remarkable decrease in mid and later periods (75 - 120 d), compared with those under the application of controlled-release fertilizers CRFA (N:P:K = 20:8:10) and CRFB (N:P:K= 14:14:14). Under the application of CRFA, the F(v)/F(m), phi(PS II), and photochemical quenching (q(P)) had somewhat increase, as compared with the application of CRFB. The non-photochemical quenching (NPQ) under the application of CRFA and CRFB decreased significantly, compared with that under the application of CCFA and CCFB and the control. The chlorophyll content had a similar change trend with F(v)/F(m), F(v)/F(0), and phi(PS II). The leaf N, P, and K contents, flower stalk length and stalk diameter, flower diameter, and flower fresh and dry mass at harvest stage all increased under the application of CRFA and CRFB, compared with those under the application of CCFA and CCFB and the control, and the flower fresh and dry mass was significantly higher under the application of CRFA than of CRFB. This study showed that controlled-release fertilizer could improve the ornamental quality of chrysanthemum via improving the leaf chlorophyll content, photochemical transduction rate, and nutrient uptake, and CRFA had better effects than CRFB.

[Effects of ca(2+) -carrier A23187 and Ca(2+) -chelator EGTA on the flower formation of chrysanthemum under photoperiodic induction and the Ca2+ distribution and carbohydrate contents in leaves during the flower formation].

This paper studied the effects of Ca(2+) -carrier A23187 and Ca(2+) -chelator EGTA on the bud differentiation of cut flower chrysanthemum (Dendranthema grandiflorium 'Shenma') under photoperiodic induction, as well as the Ca2+ distribution and the sucrose, soluble sugar, and starch contents in 'Shenma' leaves during the differentiation. In the control, the leaf Ca2+ content was lower at the vegetative stage of apical bud (I), increased rapidly and reached a peak at the stage of initial differentiation (II), and decreased then. At stage I, the Ca2+ was mainly allocated in vacuole, cell wall, and cell lacuna; while at stage II, it was more in cytoplasm. Compared with the control, the leaf Ca2+ content of A23187-treated plants increased significantly, and the days of initiation and ending of bud differentiation were advanced by 2 days and 3 days, respectively. On the other hand, the leaf Ca2+ content of EGTA-treated plants decreased significantly, and the days of initiation and ending of bud differentiation were postponed by 4 days and 8 days, respectively. For both A23187- and EGTA-treated plants, their leaf Ca2+ at stage II was more allocated in cytoplasm. The leaf sucrose and soluble sugar contents of A23187-treated plants reached a peak on the 2nd day after treatment, and the time to reach the peak was shortened by 2 days, compared with the control, which was consistent with the peak time of Ca2+. The leaf sucrose and soluble sugar contents of EGTA-treated plants had no significant changes on the 2nd day of treatment, but increased rapidly and reached the peak on the 8th day of treatment (stage II), and then decreased. However, the leaf sucrose and soluble sugar contents during the whole period of bud differentiation were higher than those before photoperiodic induction. The leaf starch content of A23187-treated plants and the control decreased 2 days after treatment, while that of EGTA-treated plants began to decrease 8 days after treatment, and maintained at a lower level by the end of bud differentiation. The results indicated that Ca2+ and carbohydrates participated in the flower formation of chrysanthemum under photoperiodic induction.

[Effects of low temperature- and weak light stress and its recovery on the photosynthesis and chlorophyll fluorescence parameters of cut flower chrysanthemum].

The cut flower chrysanthemum 'Jinba' was respectively treated with lower temperature and weaker light (16 degrees C/ 12 degrees C, PFD 100 micromol x m(-2) x s(-1)) and critical low temperature and weak light (12 degrees C/8 degrees C, PFD 60 micromol x m(-2) x s(-1)) for 11 days, and then transferred to normal condition (22 degrees C/18 degrees C, PFD 450 micromol x m(-2) x s(-1)) for 11 days, aimed to study the low temperature- and weak light stress and its recovery on the photosynthesis and chlorophyll fluorescence of chrysanthemum leaves. Under the stress of lower temperature and weaker light, the net photosynthetic rate (P(n)) and stomatal limitation (L(s)) of chrysanthemum leaves decreased while the intercellular CO2 concentration (C(i)) increased, the maximal photochemical efficiency of PS II (F(v)/F(m)) in dark and the initial fluorescence (F(o)) had no obvious change, but the maximal photochemical efficiency of PS II (F(v)'/F(m)') in light increased after an initial decrease. Contrarily, under the stress of critical low temperature and weak light, the F(o) increased, and the F(v)/F(m) and F(v)'/F(m)' decreased significantly. The quantum yield of PS II electron transport (phi(PS II)), photochemical quenching (q(p)), and apparent photosynthetic electron transfer rate (ETR) of chrysanthemum leaves decreased with increasing stress and time, and recovered quickly after the release of lower temperature- and weaker light stress but more slowly after the release of critical low temperature- and weak light stress. At the same time, the photochemistry react rate (Prate) decreased, but the hot dissipation of antenna (Drate) and the energy dissipation of PS II (Ex) increased under the stress conditions. Drate was the main pathway of superfluous light allocation.

[Effects of exogenous Ca2+ on leaf photosynthetic apparatus and active oxygen scavenging enzyme system of chrysanthemum under high temperature stress].

Taking cut flower chrysanthemum 'Jinba' as test material, the effects of exogenous Ca2+ on its photosynthetic system and antioxidant enzyme activities under high temperature stress were investigated, with the possible action mechanisms of Ca2+ discussed. The results showed that under high temperature stress, Ca2+ addition greatly inhibited the net photosynthesis rate (P(n)) and quantum yield of PS II electron transport (phi(PS II)). After 24 h treatment, the P(n) and psi(PS II) were increased by 31.11% and 21.88% , respectively, and the initial fluorescence (F(o)) decreased by 13.19%, compare with the control. Ca2+ addition also greatly enhanced the activities of SOD, POD and CAT, and thus, the active oxygen was scavenged timely. After 24 h treatment, the MDA accumulation and REC were 29.20% and 35.81% lower than the control, respectively. In conclusion, Ca2+ addition could efficiently protect chrysanthemum leaves from the damage in photosynthetic apparatus under short-term high temperature stress.

[Effects of high temperature stress on photosynthesis and chlorophyll fluorescence of cut flower chrysanthemum (Dendranthema grandiflora 'Jinba')].

Cut flower chrysanthemum (Dendranthema grandflora 'Jinba') plants were treated with 40 degrees C/35 degrees C or 33 degrees C/28 degrees C (day/night) for 11 days and then transferred to 23 degrees C/18 degrees C for 5 days to study the changes in their photosynthesis and fluorescence parameters under high temperature stress and normal temperature recovery. The results showed that on the 5th day of 33 degrees C/28 degrees C treatment, net photosynthesis (P(n)) decreased gradually and stomatal conductance (G(s)) decreased evidently; while after recovery for 5 days, both P(n) and G(s) resumed to 80% of the control. At 40 degrees C/35 degrees C, P(n) and G(s) decreased dramatically. The increase of intercellular CO2 concentration (C(i)) at the early stage under given high temperatures showed that the photosynthesis inhibition by high temperature stress was resulted from non-stomatal limitations. However, 9 days later, stomatal limitation became the mainly cause of photosynthesis inhibition. The intrinsic photochemical efficiency (F(v)/F(m)), quantum yield of PS II (phi(PS II), and the efficiency of excitation energy capture by open PS II reaction center (F(v)'/F(m)') at 33 degrees C/28 degrees C and 40 degrees C/35 degrees C all decreased, with antenna heat dissipation increased, indicating that reaction center was protected by decreased light capture and efficiency of electron transfer through PS II. The photochemical quenching (q(p)) at 33 degrees C/28 degrees C descended first and turned to rise then, suggesting that the electron transfer was firstly restrained by the stress. Contrastively, q(p) rose continuously at 40 degrees C/35 degrees C, indicating that oxygen-evolving complex (OEC) was the location in chrysanthemum photosynthesis apparatus most sensitive to extreme high temperature.