Potato oxidized hydroxypropyl starch/pectin-based indicator film with Clitoria ternatea anthocyanin and silver nanoparticles for monitoring chilled beef freshness.

Potato oxidized hydroxypropyl starch (POHS)/pectin (P) functional and smart beef freshness indicator films were prepared using butterfly pea (Clitoria ternatea) anthocyanin (BA) and silver nanoparticles (AgNPs). BA exhibited significant pH-responsive color changes. BA and AgNPs were evenly distributed within a polymer matrix to create a compatible film with POHS/P. The films containing BA and AgNPs had good UV resistance and maintained strong mechanical strength, barrier properties, and color stability. The color of the indicator film changed from purple to green when exposed to ammonia, with the 1 % POHS/P/BA/AgNPs film showing the most sensitive response. The films also demonstrated strong antibacterial and antioxidant properties. The freshness of beef was monitored using 1 % POHS/P/BA/AgNPs films and was identified as sub-fresh and spoiled on days 4 and 7, respectively. The relationship between the color change of the indicator label and the freshness of chilled beef was established: purple for fresh meat, blue for less fresh meat, and green for spoiled meat. Thus, the new POHS/P/BA/AgNPs film can serve as a smart packaging material to indicate food freshness and extend shelf life. These results suggest that POHS/P/BA/AgNPs films have significant potential as an active and smart food packaging material.

Unveiling the mechanism of source-sink rebalancing in cucumber-pumpkin heterografts: the buffering roles of rootstock cotyledon.

Grafting onto pumpkin rootstock is widely applied in cucumber production to improve growth and yield, as well as to overcome soil-borne diseases and enhance resistance to abiotic stresses. In this study, we constructed the cucumber-pumpkin heterografts with the one-cotyledon grafting method, and examined the effects of heterografting on biomass allocation and sugar partitioning, with cucumber and pumpkin self-grafts used as control. Compared with cucumber self-grafts, heterografting onto pumpkin rootstock promoted photosynthesis in cucumber scion, and led to higher sucrose contents in the 1st true leaf (source) and newly emerged leaf (sink). Thereby, the scion part of heterografts accumulated more biomass than cucumber self-grafts. In contrast, when compared to pumpkin self-grafts, grafting with cucumber scion reduced root vigor and biomass but promoted cotyledon growth in pumpkin rootstock. The roots (sink) of heterografts contained less sucrose and hexoses, and showed reduced sucrose synthase (SuSy) and hexokinase (HXK) activities. However, the rootstock cotyledon (source) contained more sucrose and starch, and showed higher activities of HXK, cell-wall invertase (CWIN), and enzymes for starch synthesis and degradation. Furthermore, removal or shade of rootstock cotyledon led to reduced growth of root and scion. Silencing of CmoMEX1a gene in rootstock cotyledon inhibited maltose export and reduced root growth of heterografts. These results indicated that rootstock cotyledon, especially its starch content, played a buffering role in the growth regulation of cucumber-pumpkin heterografts. Taken together, our results provided a major contribution to our understanding of source-sink sugar partitioning and scion-rootstock growth balancing in cucumber-pumpkin heterografts.

24-Epibrassinolide Facilitates Adventitious Root Formation by Coordinating Cell-Wall Polyamine Oxidase- and Plasma Membrane Respiratory Burst Oxidase Homologue-Derived Reactive Oxygen Species in Capsicum annuum L.

Adventitious root (AR) formation is a critical process in cutting propagation of horticultural plants. Brassinosteroids (BRs) have been shown to regulate AR formation in several plant species; however, little is known about their exact effects on pepper AR formation, and the downstream signaling of BRs also remains elusive. In this study, we showed that treatment of 24-Epibrassinolide (EBL, an active BR) at the concentrations of 20-100 nM promoted AR formation in pepper (Capsicum annuum). Furthermore, we investigated the roles of apoplastic reactive oxygen species (ROS), including hydrogen peroxide (H2O2) and superoxide radical (O2•-), in EBL-promoted AR formation, by using physiological, histochemical, bioinformatic, and biochemical approaches. EBL promoted AR formation by modulating cell-wall-located polyamine oxidase (PAO)-dependent H2O2 production and respiratory burst oxidase homologue (RBOH)-dependent O2•- production, respectively. Screening of CaPAO and CaRBOH gene families combined with gene expression analysis suggested that EBL-promoted AR formation correlated with the upregulation of CaPAO1, CaRBOH2, CaRBOH5, and CaRBOH6 in the AR zone. Transient expression analysis confirmed that CaPAO1 was able to produce H2O2, and CaRBOH2, CaRBOH5, and CaRBOH6 were capable of producing O2•-. The silencing of CaPAO1, CaRBOH2, CaRBOH5, and CaRBOH6 in pepper decreased the ROS accumulation and abolished the EBL-induced AR formation. Overall, these results uncover one of the regulatory pathways for BR-regulated AR formation, and extend our knowledge of the functions of BRs and of the BRs-ROS crosstalk in plant development.

Chitosan coating with grape peel extract: A promising coating to enhance the freeze-thaw stability of beef.

This study investigated the effect of chitosan coating with grape peel extracts (CH + GPE) on the physiochemical properties, protein and lipid oxidation, microstructure, and bacterial community diversity of beef during freeze-thaw (F-T) cycles. The results indicated that the CH + GPE groups had lower pH values, total aerobic count, total volatile base nitrogen, and thiobarbituric acid reactive substance values and better protection against color, water holding capacity, and sensory quality after five F-T cycles. The CH + GPE coating effectively inhibited beef microstructure destruction during the F-T cycles. High-throughput sequencing analysis revealed that the CH + GPE coating contributed to a decline in the bacterial diversity of beef and inhibited the growth of pathogenic bacteria. Interestingly, the CH + GPE coating affected the correlation between quality parameters and bacteria in beef. Consequently, the CH + GPE coating can be used as a novel packaging for preventing the loss of frozen meat quality due to temperature fluctuations.

ABA-responsive AREB1/ABI3-1/ABI5 cascade regulates IAA oxidase gene SlDAO2 to inhibit hypocotyl elongation in tomato.

Hypocotyl elongation is dramatically influenced by environmental factors and phytohormones. Indole-3-acetic acid (IAA) plays a prominent role in hypocotyl elongation, whereas abscisic acid (ABA) is regarded as an inhibitor through repressing IAA synthesis and signalling. However, the regulatory role of ABA in local IAA deactivation remains largely uncharacterized. In this study, we confirmed the antagonistic interplay of ABA and IAA during the hypocotyl elongation of tomato (Solanum lycopersicum) seedlings. We identified an IAA oxidase enzyme DIOXYGENASE FOR AUXIN OXIDATION2 (SlDAO2), and its expression was induced by both external and internal ABA signals in tomato hypocotyls. Moreover, the overexpression of SlDAO2 led to a reduced sensitivity to IAA, and the knockout of SlDAO2 alleviated the inhibitory effect of ABA on hypocotyl elongation. Furthermore, an ABA-responsive regulatory SlAREB1/SlABI3-1/SlABI5 cascade was identified to act upstream of SlDAO2 and to precisely control its expression. SlAREB1 directly bound to the ABRE present in the SlDAO2 promoter to activate SlDAO2 expression, and SlABI3-1 enhanced while SlABI5 inhibited the activation ability of SlAREB1 by directly interacting with SlAREB1. Our findings revealed that ABA might induce local IAA oxidation and deactivation via SlDAO2 to modulate IAA homoeostasis and thereby repress hypocotyl elongation in tomato.

Community-acquired Pseudomonas aeruginosa pneumonia manifested by bloody pleural effusion in a previously healthy infant: A case report.

BACKGROUND: Community-acquired Pseudomonas aeruginosa pneumonia in immunocompetent children is a rare occurrence. METHODS: A retrospective analysis of the clinical manifestations, imaging characteristics, laboratory examinations, and treatment of a child with community-acquired Pseudomonas aeruginosa pneumonia presented with bloody pleural effusion. RESULTS: The 1-year-old previously healthy patient, who developed community-acquired pneumonia caused by Pseudomonas aeruginosa and influenza virus. The patient manifested bloody pleural effusion although his condition improved after anti-infective therapy and closed thoracic drainage. After 10 days of hospitalization, his symptoms worsened, accompanied by hemoptysis, and the pathogen developed resistance to carbapenems. The antibiotic strategy was adjusted to combined antipseudomonal regimen. He developed low-grade fever and was extubated, although these manifestations and imaging were eventually alleviated. CONCLUSIONS: Community-acquired Pseudomonas aeruginosa pneumonia in children may be non-septic, with bloody pleural effusion as presentation, and the disease may progress after 10 days of treatment due to drug resistance in Pseudomonas aeruginosa. Early extubation should be considered after adequate drainage.

Development of a Nomogram for Predicting Refractory Mycoplasma pneumoniae Pneumonia in Children.

Background: In children, refractory Mycoplasma pneumoniae pneumonia (RMPP) may result in severe complications and high medical costs. There is research on a simple and easy-to-use nomogram for early prediction and timely treatment of RMPP. Methods: From December 2018 to June 2021, we retrospectively reviewed medical records of 299 children with Mycoplasma pneumoniae pneumonia (MPP) hospitalized in Tianjin Children's Hospital. According to their clinical manifestations, patients were divided into the RMPP group and the general Mycoplasma pneumoniae pneumonia (GMPP) group. The clinical manifestations, laboratory indicators, and radiological data of the two groups were obtained. Stepwise regression was employed for variable selection of RMPP. The predictive factors selected were used to construct a prediction model which presented with a nomogram. The performance of the prediction model was evaluated by C statistics, calibration curve, and receiver operating characteristic (ROC) curve. Results: The RMPP group significantly showed a higher proportion of females, longer fever duration, and longer hospital stay than the GMPP group (P < 0.05). Additionally, the RMPP group revealed severe clinical characteristics, including higher incidences of extrapulmonary complications, decreased breath sounds, unilateral pulmonary consolidation >2/3, and plastic bronchitis than the GMPP group (P < 0.05). The RMPP group had higher neutrophil ratio (N%), C-reactive protein (CRP), interleukin-6 (IL-6), lactic dehydrogenase (LDH), and D-dimer than the GMPP group (P < 0.05). Stepwise regression demonstrated that CRP [OR = 1.075 (95% CI: 1.020-1.133), P < 0.001], LDH [OR = 1.015 (95% CI: 1.010-1.020), P < 0.001], and D-dimer [OR = 70.94 (95% CI: 23.861-210.904), P < 0.001] were predictive factors for RMPP, and developed a prediction model of RMPP, which can be visualized and accurately quantified using a nomogram. The nomogram showed good discrimination and calibration. The area under the ROC curve of the nomogram was 0.881, 95% CI (0.843, 0.918) in training cohorts and 0.777, 95% CI (0.661, 0.893) in validation cohorts, respectively. Conclusion: C-reactive protein, LDH, and D-dimer were predictive factors for RMPP. The simple and easy-to-use nomogram assisted us in quantifying the risk for predicting RMPP, and more accurately and conveniently guiding clinicians to recognize RMPP, and contribute to a rational therapeutic choice.

Stage-specific events in tomato graft formation and the regulatory effects of auxin and cytokinin.

Grafting is widely used worldwide because of its obvious advantages, especially in solanaceous vegetable crops. However, the molecular mechanisms underlying graft formation are unknown. In this study, internode tissues from above and below the graft junction were harvested, and we performed weighted gene co-expression network analysis (WGCNA) to describe the temporal and spatial transcriptional dynamics that occur during graft formation in tomato. The wounding stress response involved in JA, ETH, and oxylipins mainly occurred at 1 h after grafting (HAG). From 3 to 12 HAG, the biological processes of snRNA and snoRNA modification and the gibberellin-mediated signaling pathway functioned both above and below the graft junction. However, auxin transport and signaling, DNA replication, and xylem and phloem pattern formation were restricted to the scion, whereas the cytokinin-activated signaling pathway and the cellular response to sucrose starvation was restricted to the rootstock. At 24-72 HAG, cell division occurred above the graft junction, and photosynthesis-related pathways were activated below the graft junction. The levels of auxin and cytokinin reached their maxima above and below the graft junction at 12 HAG, respectively. Exogenous application of certain concentrations of IAA and 6-BA will promote xylem and phloem transport capacity. The current work has analyzed the stage-specific events and hub genes during the developmental progression of tomato grafting. We found that auxin and cytokinin levels respond to grafting, above and below the graft junction, respectively, to promote the formation of xylem and phloem patterning. In addition, the accumulation of auxin above the graft junction induced cells to prepare for mitosis and promoted the formation of callus. In short, our work provides an important reference for theoretical research and production application of tomato grafting in the future.

Salicylic acid regulates adventitious root formation via competitive inhibition of the auxin conjugation enzyme CsGH3.5 in cucumber hypocotyls.

MAIN CONCLUSION: Exogenous SA treatment at appropriate concentrations promotes adventitious root formation in cucumber hypocotyls, via competitive inhibiting the IAA-Asp synthetase activity of CsGH3.5, and increasing the local free IAA level. Adventitious root formation is critical for the cutting propagation of horticultural plants. Indole-3-acetic acid (IAA) has been shown to play a central role in regulating this process, while for salicylic acid (SA), its exact effects and regulatory mechanism have not been elucidated. In this study, we showed that exogenous SA treatment at the concentrations of both 50 and 100 µM promoted adventitious root formation at the base of the hypocotyl of cucumber seedlings. At these concentrations, SA could induce the expression of CYCLIN and Cyclin-dependent Kinase (CDK) genes during adventitious rooting. IAA was shown to be involved in SA-induced adventitious root formation in cucumber hypocotyls. Exposure to exogenous SA led to a slight increase in the free IAA content, and pre-treatment with the auxin transport inhibitor 1-naphthylphthalamic acid (NPA) almost completely abolished the inducible effects of SA on adventitious root number. SA-induced IAA accumulation was also associated with the enhanced expression of Gretchen Hagen3.5 (CsGH3.5). The in vitro enzymatic assay indicated that CsGH3.5 has both IAA- and SA-amido synthetase activity and prefers aspartate (Asp) as the amino acid conjugate. The Asp concentration dictated the functional activity of CsGH3.5 on IAA. Both affinity and catalytic efficiency (Kcat/Km) increased when the Asp concentration increased from 0.3 to 1 mM. In contrast, CsGH3.5 showed equal catalytic efficiency for SA at low and high Asp concentrations. Furthermore, SA functioned as a competitive inhibitor of the IAA-Asp synthetase activity of CsGH3.5. During adventitious formation, SA application indeed repressed the IAA-Asp levels in the rooting zone. These data show that SA plays an inducible role in adventitious root formation in cucumber through competitive inhibition of the auxin conjugation enzyme CsGH3.5. SA reduces the IAA conjugate levels, thereby increasing the local free IAA level and ultimately enhancing adventitious root formation.

Dynamic changes in bacterial communities in the recirculating nutrient solution of cucumber plug seedlings cultivated in an ebb-and-flow subirrigation system.

Ebb-and-flow subirrigation systems are highly efficient, water-saving and environmentally friendly. However, one concern with these recirculating systems is the possible transmission of plant pathogens. Here, through 16S rRNA-targeted Illumina sequencing, the bacterial dynamics in a recirculating nutrient solution were characterized for cucumber plug seedlings cultivated in an ebb-and-flow system in summer and winter. Both the bacterial number and diversity in the nutrient solution increased immediately after the first irrigation cycle; then, these values were gradually stable with recirculating irrigation. In summer and winter, different bacterial compositions and changing patterns were observed. In summer, the predominant genera in the nutrient solution included Comamonas, Pseudomonas, Acinetobacter, Reyranella, Sphingobium, Bradyrhizobium, Sphingomonas, and Acidovorax. Of those genera, during recirculating irrigation, the relative abundance of Bradyrhizobium gradually decreased, whereas those of Pseudomonas, Reyranella, Sphingobium, Sphingomonas, and Acidovorax gradually increased. In winter, the bacterial communities were mainly composed of Nevskia, Bosea, Sphingobium, Acidovorax, Pseudomonas, and Hydrocarboniphaga. Of those genera, the relative abundance of Bosea, Sphingobium, and Acidovorax showed an increasing trend, whereas those of Nevskia and Hydrocarboniphaga decreased overall. Furthermore, in both summer and winter, no plant pathogenic bacteria on cucumber could be detected; however, some potentially beneficial bacteria, including Comamonas testosteroni, Acinetobacter baumannii, Pseudomonas aeruginosa, P. koreensis and Sphingobium yanoikuyae, colonized the nutrient solution and exhibited increased relative abundances during irrigation. The colonization of these bacteria might facilitate the plant growth promotion. Inoculation of the microbes from the effluent nutrient solution also promoted the growth of cucumber seedlings, but did not lead to any disease. The present data elucidate the bacterial dynamics in a cucumber cultivation ebb-and-flow system and provide useful information for biological control during cucumber seedling production.

Bacterial communities in the rhizosphere, phyllosphere and endosphere of tomato plants.

Plants harbor diverse bacterial communities, which play crucial roles in plant health and growth, in their rhizosphere, phyllosphere and endosphere. Tomato is an important model for studying plant-microbe interactions, but comparison of its associated bacterial community is still lacking. In this study, using Illumina sequencing of 16S rRNA amplicons, we characterized and compared the bacterial size and community from rootzone soil as well as the rhizosphere, phyllosphere and endosphere of roots, stems, leaves, fruits and seeds of tomato plants that were grown in greenhouse conditions. Habitat (soil, phyllospheric, and endophytic) structured the community. The bacterial communities from the soil-type samples (rootzone soil and rhizosphere) showed the highest richness and diversity. The lowest bacterial diversity occurred in the phyllospheric samples, while the lowest richness occurred in the endosphere. Among the endophytic samples, both bacterial diversity and richness varied in different tissues, with the highest values in roots. The most abundant phyla in the tomato-associated community was Proteobacteria, with the exception of the seeds and jelly, where both Proteobacteria and Firmicutes were dominant. At the genus level, the sequences of Pseudomonas and Acinetobacter were prevalent in the rhizosphere, and in the phyllosphere, more than 97% of the sequences were assigned to Acinetobacter. For the endophytes, Acinetobacter, Enterobacter, and Pseudomonas were the abundant genera in the roots, stems and leaves. In the fruits, the bacterial endophytes varied in different compartments, with Enterobacter being enriched in the pericarp and seeds, Acinetobacter in the placenta, and Weissella in the jelly. The present data provide a comprehensive description of the tomato-associated bacterial community and will be useful for better understanding plant-microbe interactions and selecting suitable bacterial taxa for tomato production.

Gene co-expression network analysis reveals pathways associated with graft healing by asymmetric profiling in tomato.

BACKGROUND: The ability of severed rootstocks and shoots to re-establish vascular connections is used to generate grafted plants that combine desirable traits from both scions and rootstocks. Clarifying the mechanisms of graft healing is essential for its further application. We performed RNA sequencing of internodes near the cut position, making a distinction between separated or grafted tissues above and below the cut, in order to obtain a genetic description of graft union formation. RESULTS: Using weighted gene co-expression analysis, variable transcripts were clustered into 10 distinct co-expression networks (modules) based on expression profiles, and genes with the most "hubness" ("hub" genes show the most connections in a network) within each module were predicted. A large proportion of modules were related to Position, and represent asymmetric expression networks from different pathways. Expression of genes involved in auxin and sugar transport and signaling, and brassinosteroid biosynthesis was increased above the cut, while stress response genes were up-regulated below the cut. Some modules were related to graft union formation, among which oxidative detoxification genes were co-expressed along with both wounding response and cell wall organization genes. CONCLUSIONS: The present work provides a comprehensive understanding of graft healing-related gene networks in tomato. Also, the candidate pathways and hub genes identified here will be valuable for future studies of grafting in tomato.

GhMCS1, the Cotton Orthologue of Human GRIM-19, Is a Subunit of Mitochondrial Complex I and Associated with Cotton Fibre Growth.

GRIM-19 (Gene associated with Retinoid-Interferon-induced Mortality 19) is a subunit of mitochondrial respiratory complex I in mammalian systems, and it has been demonstrated to be a multifunctional protein involved in the cell cycle, cell motility and innate immunity. However, little is known about the molecular functions of its homologues in plants. Here, we characterised GhMCS1, an orthologue of human GRIM-19 from cotton (Gossypium hirsutum L.), and found that it was essential for maintaining complex integrity and mitochondrial function in cotton. GhMCS1 was detected in various cotton tissues, with high levels expressed in developing fibres and flowers and lower levels in leaves, roots and ovules. In fibres at different developmental stages, GhMCS1 expression peaked at 5-15 days post anthesis (dpa) and then decreased at 20 dpa and diminished at 25 dpa. By Western blot analysis, GhMCS1 was observed to be localised to the mitochondria of cotton leaves and to colocalise with complex I. In Arabidopsis, GhMCS1 overexpression enhanced the assembly of complex I and thus respiratory activity, whereas the GhMCS1 homologue (At1g04630) knockdown mutants showed significantly decreased respiratory activities. Furthermore, the mutants presented with some phenotypic changes, such as smaller whole-plant architecture, poorly developed seeds and fewer trichomes. More importantly, in the cotton fibres, both the GhMCS1 transcript and protein levels were correlated with respiratory activity and fibre developmental phase. Our results suggest that GhMCS1, a functional ortholog of the human GRIM-19, is an essential subunit of mitochondrial complex I and is involved in cotton fibre development. The present data may deepen our knowledge on the potential roles of mitochondria in fibre morphogenesis.

Quantitative Proteomic Profiling of Early and Late Responses to Salicylic Acid in Cucumber Leaves.

Salicylic acid (SA) is an important phytohormone that plays vital regulatory roles in plant growth, development, and stress responses. However, studies on the molecular mechanism of SA, especially during the early SA responses, are lagging behind. In this study, we initiated a comprehensive isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic analysis to explore the early and late SA-responsive proteins in leaves of cucumber (Cucumis sativus L.) seedlings. Upon SA application through the roots, endogenous SA accumulated in cucumber leaves. By assaying the changes in marker gene expression and photosynthetic rate, we collected samples at 12 h and 72 h post treatment (hpt) to profile the early and late SA responsiveness, respectively. The iTRAQ assay followed by tandem mass spectrometry revealed 135 differentially expressed proteins (DEPs) at 12 hpt and 301 DEPs at 72 hpt. The functional categories for these SA-responsive proteins included in a variety of biochemical processes, including photosynthesis, redox homeostasis, carbohydrate and energy metabolism, lipid metabolism, transport, protein folding and modification, proteolysis, cell wall organization, and the secondary phenylpropanoid pathway. Conclusively, based on the abundant changes of these DEPs, together with their putative functions, we proposed a possible SA-responsive protein network. It appears that SA could elicit reactive oxygen species (ROS) production via enhancing the photosynthetic electron transferring, and then confer some growth-promoting and stress-priming effects on cells during the late phase, including enhanced photosynthesis and ROS scavenging, altered carbon metabolic flux for the biosynthesis of amino acids and nucleotides, and cell wall reorganization. Overall, the present iTRAQ assay provides higher proteome coverage and deepened our understanding of the molecular basis of SA-responses.

Comparative proteomic analysis reveals intracellular targets for bacillomycin L to induce Rhizoctonia solani Kühn hyphal cell death.

Bacillomycin L, a natural iturinic lipopeptide produced by Bacillus amyloliquefaciens, is characterized by strong antifungal activity against a variety of agronomically important filamentous fungi including Rhizoctonia solani Kühn. To further understand its antifungal actions, proteomes were comparatively studied within R. solani hyphal cells treated with or without bacillomycin L. The results show that 39 proteins were alternatively expressed within cells in response to this lipopeptide, which are involved in stress response, carbohydrate, amino acid and nucleotide metabolism, cellular component organization, calcium homeostasis, protein degradation, RNA processing, gene transcription, and others, suggesting that, in addition to inducing cell membrane permeabilization, iturin exhibits antibiotic activities by targeting intracellular molecules. Based on these results, a model of action of bacillomycin L against R. solani hyphal cells was proposed. Our study provides new insight into the antibiotic mechanisms of iturins.

Genome-Wide Comparative Analysis of the Phospholipase D Gene Families among Allotetraploid Cotton and Its Diploid Progenitors.

In this study, 40 phospholipase D (PLD) genes were identified from allotetraploid cotton Gossypium hirsutum, and 20 PLD genes were examined in diploid cotton Gossypium raimondii. Combining with 19 previously identified Gossypium arboreum PLD genes, a comparative analysis was performed among the PLD gene families among allotetraploid and two diploid cottons. Based on the orthologous relationships, we found that almost each G. hirsutum PLD had a corresponding homolog in the G. arboreum and G. raimondii genomes, except for GhPLDß3A, whose homolog GaPLDß3 may have been lost during the evolution of G. arboreum after the interspecific hybridization. Phylogenetic analysis showed that all of the cotton PLDs were unevenly classified into six numbered subgroups: α, ß/γ, δ, ε, ζ and φ. An N-terminal C2 domain was found in the α, ß/γ, δ and ε subgroups, while phox homology (PX) and pleckstrin homology (PH) domains were identified in the ζ subgroup. The subgroup φ possessed a single peptide instead of a functional domain. In each phylogenetic subgroup, the PLDs showed high conservation in gene structure and amino acid sequences in functional domains. The expansion of GhPLD and GrPLD gene families were mainly attributed to segmental duplication and partly attributed to tandem duplication. Furthermore, purifying selection played a critical role in the evolution of PLD genes in cotton. Quantitative RT-PCR documented that allotetraploid cotton PLD genes were broadly expressed and each had a unique spatial and developmental expression pattern, indicating their functional diversification in cotton growth and development. Further analysis of cis-regulatory elements elucidated transcriptional regulations and potential functions. Our comparative analysis provided valuable information for understanding the putative functions of the PLD genes in cotton fiber.

Characterization of the Fatty Acid Desaturase Genes in Cucumber: Structure, Phylogeny, and Expression Patterns.

Fatty acid desaturases (FADs) introduce double bonds into the hydrocarbon chains of fatty acids to produce unsaturated fatty acids, and therefore play a critical role in plant development and acclimation to environmental stresses. In this study, 23 full-length FAD genes in cucumber (Cucumis sativus L.) were identified through database searches, including three CsFAB2 genes, two CsFAD2 genes, fourteen CsFAD5 genes, and one gene each for CsFAD3, CsFAD4, CsFAD6 and CsFAD7. These cucumber FAD genes were distributed on all seven chromosomes and two additional scaffolds. Based on a phylogenetic analysis, the cucumber FAD proteins were clustered into five subfamilies with their counterparts from other plants. Gene structures and protein sequences were considerably conserved in each subfamily. All three CsFAB2 proteins shared conserved structure with the known plant soluble FAD proteins. The other cucumber FADs belonged to the membrane-bound FADs and contained three highly conserved histidine boxes. Additionally, the putative endoplasmic reticulum retention signal was found at the C-termini of the CsFAD2 and CsFAD3 proteins, while the N-termini of CsFAD4, CsFAD5, CsFAD6, CsFAD7 and three CsFAB2s contained a predicted chloroplast signal peptide, which was consistent with their associated metabolic pathways. Furthermore, a gene expression analysis showed that CsFAD2 and CsFAD3 were universally expressed in all tested tissues, whereas the other cucumber FAD genes were preferentially expressed in the cotyledons or leaves. The tissue-specific expression patterns of cucumber FAD genes were correlated well with the differences in the fatty acid compositions ofroots and leaves. Finally, the cucumber FAD genes showed a cold-induced and heat-repressed expression pattern, although with distinct regulatory time courses among the different CsFAD members, which indicates the potential roles of the FADs in temperature stress resistance in cucumber.

Genome-wide analysis and expression profiling of the phospholipase D gene family in Gossypium arboreum.

The plant phospholipase D (PLD) plays versatile functions in multiple aspects of plant growth, development, and stress responses. However, until now, our knowledge concerning the PLD gene family members and their expression patterns in cotton has been limited. In this study, we performed for the first time the genome-wide analysis and expression profiling of PLD gene family in Gossypium arboretum, and finally, a total of 19 non-redundant PLD genes (GaPLDs) were identified. Based on the phylogenetic analysis, they were divided into six well-supported clades (α, ß/γ, δ, ε, ζ and φ). Most of the GaPLD genes within the same clade showed the similar exon-intron organization and highly conserved motif structures. Additionally, the chromosomal distribution pattern revealed that GaPLD genes were unevenly distributed across 10 of the 13 cotton chromosomes. Segmental duplication is the major contributor to the expansion of GaPLD gene family and estimated to have occurred from 19.61 to 20.44 million years ago when a recent large-scale genome duplication occurred in cotton. Moreover, the expression profiling provides the functional divergence of GaPLD genes in cotton and provides some new light on the molecular mechanisms of GaPLDα1 and GaPLDδ2 in fiber development.

[Roles of phenylalanine ammonia-lyase in low temperature tolerance in cucumber seedlings].

To reveal the roles of phenylalanine ammonia-lyase (PAL) in low temperature tolerance in cucumber seedlings, a specific PAL inhibitor (AOPP) was sprayed to the seedlings, and then the stress tolerance was determined. The results suggested that both gene expression and enzymatic activity of PAL in cucumber leaves were induced under low temperature. The seedlings pretreated with AOPP showed lower PAL activity and less accumulation of phenolics and flavonoids. Low temperature caused damages in cucumber seedlings, and pretreatment of AOPP aggravated these damages. Compared to the control, the seedlings pretreated with AOPP showed significantly higher relative electrolyte leakage and MDA production, lower maximum photochemical efficiency of PSII (Fv/Fm) but higher photo-chemical quenching coefficient Y(NO), and reduced expression of low temperature-responsive genes (PR1-la, COR47, P5CS and HSP70). In cucumber seedlings, low temperature stress induced the accumulation of H2O2, increased the contents of ascobate (AsA) but decreased the contents of dehydroascobate (DHA), and thus reduced the value of AsA: DHA. In the AOPP-pretreated seedlings, the activities of antioxidant enzymes (CAT and APX) were significantly repressed, and excess H2O2 accumulated. The value of AsA: DHA was also lower than the control. Furthermore, co-application of H2O2 scavenger alleviated the low temperature-induced damages in the AOPP-pretreated seedlings, while coapplication of a CAT inhibitor made the seedlings more sensitive to low temperature stress. These results indicated that under low temperature stress, the enhanced activities of PAL could increase the biosynthesis of phenylpropanoid compounds and activate the cellular antioxidant enzymes, which could scavenge the excess ROS and maintain the cellular redox status, and thereby reduce the photo- and oxidative damages caused by low temperature stress.

Comparative proteomic and biochemical analyses reveal different molecular events occurring in the process of fiber initiation between wild-type allotetraploid cotton and its fuzzless-lintless mutant.

To explore lint fiber initiation-related proteins in allotetraploid cotton (Gossypium hirsutum L.), a comparative proteomic analysis was performed between wild-type cotton (Xu-142) and its fuzzless-lintless mutant (Xu-142-fl) at five developmental time points for lint fiber initiation from -3 to +3 days post-anthesis (dpa). Using two-dimensional gel electrophoresis (2-DE) combined with mass spectrometry (MS) analyses, 91 differentially accumulated protein (DAP) species that are related to fiber initiation were successfully identified, of which 58 preferentially accumulated in the wild-type and 33 species in the fl mutant. These DAPs are involved in various cellular and metabolic processes, mainly including important energy/carbohydrate metabolism, redox homeostasis, amino acid and fatty acid biosynthesis, protein quality control, cytoskeleton dynamics, and anthocyanidin metabolism. Further physiological and biochemical experiments revealed dynamic changes in the carbohydrate flux and H2O2 levels in the cotton fiber initiation process. Compared with those in the fl mutant, the contents of glucose and fructose in wild-type ovules sharply increased after anthesis with a relatively higher rate of amino acid biosynthesis. The relative sugar starvation and lower rate of amino acid biosynthesis in the fl mutant ovules may impede the carbohydrate/energy supply and cell wall synthesis, which is consistent with the proteomic results. However, the H2O2 burst was only observed in the wild-type ovules on the day of anthesis. Cotton boll injection experiments in combination with electron microscope observation collectively indicated that H2O2 burst, which is negatively regulated by ascorbate peroxidases (APx), plays an important role in the fiber initiation process. Taken together, our study demonstrates a putative network of DAP species related to fiber initiation in cotton ovules and provides a foundation for future studies on the specific functions of these proteins in fiber development.