Development of a Nomogram Based on Diffusion-Weighted Imaging and Clinical Information to Predict Delayed Encephalopathy after Acute Carbon Monoxide Poisoning.

BACKGROUND: Delayed encephalopathy after acute carbon monoxide poisoning (DEACMP) is a severe complication that can arise from acute carbon monoxide poisoning (ACOP). This study aims to identify the independent risk factors associated with DEACMP and to develop a nomogram to predict the probability of developing DEACMP. METHODS: The data of patients diagnosed with ACOP between September 2015 and June 2021 were analyzed retrospectively. The patients were divided into the two groups: the DEACMP group and the non-DEACMP group. Univariate analysis and multivariate logistic regression analysis were conducted to identify the independent risk factors for DEACMP. Subsequently, a nomogram was constructed to predict the probability of DEACMP. RESULTS: The study included 122 patients, out of whom 30 (24.6%) developed DEACMP. The multivariate logistic regression analysis revealed that acute high-signal lesions on diffusion-weighted imaging (DWI), duration of carbon monoxide (CO) exposure, and Glasgow Coma Scale (GCS) score were independent risk factors for DEACMP (Odds Ratio = 6.230, 1.323, 0.714, p < 0.05). Based on these indicators, a predictive nomogram was constructed. CONCLUSIONS: This study constructed a nomogram for predicting DEACMP using high-signal lesions on DWI and clinical indicators. The nomogram may serve as a dependable tool to differentiate high-risk patients and enable the provision of personalized treatment to lower the incidence of DEACMP.

Photosynthetic contribution and characteristics of cucumber stems and petioles.

BACKGROUND: Photosynthesis in the green leafless blade tissues or organs of plants has been studied in some plants, but the photosynthetic characteristics of stems and petioles are poorly understood. Cucurbitaceous plants are climbing plants that have substantial stem and petiole biomass. Understanding the photosynthetic contribution of cucumber stems and petioles to their growth and the underlying molecular mechanisms are important for the regulating of growth in cucumber production. RESULTS: In this study, the photosynthetic capacity of cucumber stems and petioles were determined by 14CO2 uptake. The total carbon fixed by the stems and petioles was approximately 4% of that fixed by one leaf blade in the cucumber seedling stage, while the proportion of the carbon accumulated in the stems and petioles that redistributed to sink organs (roots and shoot apexes) obviously increased under leafless conditions. The photosynthetic properties of cucumber stems and petioles were studied using a combination of electron microscopy and isotope tracers to compare these properties of stems and petioles with those of leaf blade using two genotypes of cucumber (dark green and light green). Compared with those of the leaf blades, the chlorophyll contents of the cucumber stems and petioles were lower, and the stems and petioles had lower chloroplast numbers and lower stoma numbers but higher thylakoid grana lamella numbers and larger stoma sizes. The Chl a/b ratios were also decreased in the petioles and stems compared with those in the leaf blades. The total photosynthetic rates of the stems and petioles were equivalent to 6 ~ 8% of that of one leaf blade, but the respiration rates were similar in all the three organs, with an almost net 0 photosynthetic rate in the stems and petioles. Transcriptome analysis showed that compared with the leaf blades, the stems and petioles has significantly different gene expression levels in photosynthesis, porphyrin and chlorophyll metabolism; photosynthetic antenna proteins; and carbon fixation. PEPC enzyme activities were higher in the stems and petioles than in the leaf blades, suggesting that the photosynthetic and respiratory mechanisms in stems and petioles are different from those in leaf blade, and these results are consistent with the gene expression data. CONCLUSIONS: In this study, we confirmed the photosynthetic contribution to the growth of cucumber stems and petioles, and showed their similar photosynthetic patterns in the terms of anatomy, molecular biology and physiology, which were different from those of cucumber leaf blades.

Neuroprotective Effects of Long-Term Metformin Preconditioning on Rats with Ischemic Brain Injuries.

INTRODUCTION: This study is to analyze the neuroprotective effects of long-term metformin (Met) preconditioning on rats with ischemic brain injuries and the related mechanisms. METHODS: Twenty-five Sprague-Dawley rats were randomly divided into 5 groups: sham group, middle cerebral artery occlusion (MCAO) group, normal saline + MCAO group, pre- Met + MCAO group, and 3-MA + Met + MCAO group. Pathological changes of brain were observed by hematoxylin-eosin staining. Neurobehavior scores were calculated. Infarct area was assessed by 2,3,5-triphenyltetrazolium chloride staining. Apoptosis of neurons was detected by TdT-mediated dUTP Nick-End Labeling (TUNEL). Western blot tested the expression of LC3 (microtubule-associated protein 1 light chain 3), Beclin-1, adenosine 5'-monophosphate ([AMP]-activated protein kinase [AMPK]), and p-AMPK in hippocampal CA1 region. RESULTS: Compared with the sham group, the MCAO group induced severe pathological changes in the brain. The neurobehavior scores and infarct area in the brain were increased in the MCAO group than in the sham group. The apoptosis level in the MCAO group was also higher than in the sham group. However, after pretreatment with Met, the pathological changes in the brain were attenuated. Compared with the MCAO group, the pre-Met + MCAO group also had decreased neurobehavior scores and infarct area in the brain. Additionally, the apoptosis level in the pre-Met + MCAO group was lower than in the MCAO group. Moreover, the MCAO group had increased levels of LC3 and Beclin-1 than in the sham group. In the pre-Met + MCAO group, their levels were decreased than in the MCAO group. The p-AMPK level in the pre-Met + MCAO group was also increased than in the MCAO group, suggesting activation of p-AMPK by Met. CONCLUSION: Long-term Met pretreatment has neuroprotective effect on ischemic brain injury, which may be related to the regulation of autophagy-related protein expression and apoptosis.

Phloem loading in cucumber: combined symplastic and apoplastic strategies.

Phloem loading, as the first step of transporting photoassimilates from mesophyll cells to sieve element-companion cell complex, creates a driving force for long-distance nutrient transport. Three loading strategies have been proposed: passive symplastic loading, apoplastic loading and symplastic transfer followed by polymer-trapping of stachyose and raffinose. Although individual species are generally referred to as using a single phloem loading mechanism, it has been suggested that some plants may use more than one, i.e. 'mixed loading'. Here, by using a combination of electron microscopy, reverse genetics and 14 C labeling, loading strategies were studied in cucumber, a polymer-trapping loading species. The results indicate that intermediary cells (ICs), which mediate polymer-trapping, and ordinary companion cells, which mediate apoplastic loading, were mainly found in the fifth and third order veins, respectively. Accordingly, a cucumber galactinol synthase gene (CsGolS1) and a sucrose transporter gene (CsSUT2) were expressed mainly in the fifth/third and the third order veins, respectively. Immunolocalization analysis indicated that CsGolS1 was localized in companion cells (CCs) while CsSUT2 was in CCs and sieve elements (SEs). Suppressing CsGolS1 significantly decreased the stachyose level and increased sucrose content, while suppressing CsSUT2 decreased the sucrose level and increased the stachyose content in leaves. After 14 CO2 labeling, [14 C]sucrose export increased and [14 C]stachyose export reduced from petioles in CsGolS1i plants, but [14 C]sucrose export decreased and [14 C]stachyose export increased into petioles in CsSUT2i plants. Similar results were also observed after pre-treating the CsGolS1i leaves with PCMBS (transporter inhibitor). These results demonstrate that cucumber phloem loading depends on both polymer-trapping and apoplastic loading strategies.

Down-regulation of the Sucrose Transporter CsSUT1 Causes Male Sterility by Altering Carbohydrate Supply.

In plants, male sterility is an important agronomic trait, especially in hybrid crop production. Many factors are known to affect crop male sterility, but it remains unclear whether Suc transporters (SUTs) participate directly in this process. Here, we identified and functionally characterized the cucumber (Cucumis sativus) CsSUT1, a typical plasma membrane-localized energy-dependent high-affinity Suc-H+ symporter. CsSUT1 is expressed in male flowers and encodes a protein that is localized primarily in the tapetum, pollen, and companion cells of the phloem of sepals, petals, filaments, and pedicel. The male flowers of CsSUT1-RNA interference (RNAi) lines exhibited a decrease in Suc, hexose, and starch content, relative to those of the wild type, during the later stages of male flower development, a finding that was highly associated with male sterility. Transcriptomic analysis revealed that numerous genes associated with sugar metabolism, transport, and signaling, as well as with auxin signaling, were down-regulated, whereas most myeloblastosis (MYB) transcription factor genes were up-regulated in these CsSUT1-RNAi lines relative to wild type. Our findings demonstrate that male sterility can be induced by RNAi-mediated down-regulation of CsSUT1 expression, through the resultant perturbation in carbohydrate delivery and subsequent alteration in sugar and hormone signaling and up-regulation of specific MYB transcription factors. This knowledge provides a new approach for bioengineering male sterility in crop plants.

Transcriptomic and functional analysis of cucumber (Cucumis sativus L.) fruit phloem during early development.

The phloem of the Cucurbitaceae has long been a subject of interest due to its complex nature and the economic importance of the family. As in a limited number of other families, cucurbit phloem is bicollateral, i.e. with sieve tubes on both sides of the xylem. To date little is known about the specialized functions of the internal phloem (IP) and external phloem (EP). Here, a combination of microscopy, fluorescent dye transport analysis, micro-computed tomography, laser capture microdissection and RNA-sequencing (RNA-Seq) were used to study the functions of IP and EP in the vascular bundles (VBs) of cucumber fruit. There is one type of VB in the peduncle, but four in the fruit: peripheral (PeVB), main (MVB), carpel (CVB) and placental (PlVB). The VBs are bicollateral, except for the CVB and PlVB. Phloem mobile tracers and 14 C applied to leaves are transported primarily in the EP, and to a lesser extent in the IP. RNA-Seq data indicate preferential gene transcription in the IP related to differentiation/development, hormone transport, RNA or protein modification/processing/transport, and nitrogen compound metabolism and transport. The EP preferentially expresses genes for stimulus/stress, defense, ion transport and secondary metabolite biosynthesis. The MVB phloem is preferentially involved in photoassimilate transport, unloading and long-distance signaling, while the PeVB plays a more substantial role in morphogenesis and/or development and defense response. CVB and PlVB transcripts are biased toward development of reproductive organs. These findings provide an integrated view of the differentiated structure and function of the vascular tissue in cucumber fruit.

Down-Regulating Cucumber Sucrose Synthase 4 (CsSUS4) Suppresses the Growth and Development of Flowers and Fruits.

Sucrose synthase (SUS), which catalyzes the reversible conversion of sucrose and uridine diphosphate (UDP) into fructose and UDP-glucose, is a key enzyme in sucrose metabolism in higher plants. In this study, we used reverse genetic approaches and carbohydrate analysis to investigate the role of cucumber sucrose synthase gene 4 (CsSUS4) in the growth and development of sink organs. Transcript analyses showed that CsSUS4 was predominantly expressed in sink organs, particularly in flowers, fruits and roots, and that CsSUS4 protein was localized to companion cells and phloem parenchyma cells. Down-regulation of CsSUS4 expression resulted in a decrease in SUS activity in conjunction with lower hexose, starch and cellulose contents in fruits, and led to an overall reduction in the size and weight of flowers and fruits. Furthermore, CsSUS4 overexpression (OE) lines exhibited increased carbohydrate content, and larger and heavier flowers and fruits. The numbers of multi-petal flowers and multi-carpel fruits were greater in CsSUS4-OE plants compared with wild type and were regulated by MADS-box transcription factor. These results demonstrate that CsSUS4 plays important roles in the growth and development of cucumber flowers and fruits.

Fine mapping of a male sterility gene ms-3 in a novel cucumber (Cucumis sativus L.) mutant.

KEY MESSAGE: The cucumber male sterility gene ms - 3 was fine mapped in a 76 kb region harboring an MMD1 -like gene Csa3M006660 that may be responsible for the male sterile in cucumber. A cucumber (Cucumis sativus L.) male sterile mutant (ms-3) in an advanced-generation inbred line was identified, and genetic analysis revealed that the male sterility trait was controlled by a recessive nuclear gene, ms-3, which was stably inherited. Histological studies suggested that the main cause of the male sterility was defective microsporogenesis, resulting in no tetrad or microspores being formed. Bulked segregant analysis (BSA) and genotyping of an F2 population of 2553 individuals were employed used to fine map ms-3, which was delimited to a 76 Kb region. In this region, a single non-synonymous SNP was found in the Csa3M006660 gene locus, which was predicted to result in an amino acid change. Quantitative RT-PCR analysis of Csa3M006660 was consistent with the fact that it plays a role in the early development of cucumber pollen. The protein encoded by Csa3M006660 is predicted to be homeodomain (PHD) finger protein, and the high degree of sequence conservation with homologs from a range of plant species further suggested the importance of the ms-3 non-synonymous mutation. The data presented here provide support for Csa3M006660 as the most likely candidate gene for Ms-3.

Effects of generally recognized as safe (GRAS) and dietary compounds on phenylephrine metabolism in LS180 human intestinal cells.

Phenylephrine (PE) has low and variable oral bioavailability in humans, due in part to presystemic metabolism by sulfation. LS180 cells were used as a model of the human intestinal epithelium to examine phenylephrine metabolism and its inhibition by generally recognized as safe (GRAS) and dietary compounds. Curcumin, zingerone, resveratrol, guaiacol, pterostilbene and isoeugenol significantly inhibited phenylephrine disappearance, while vanillin, propylparaben and eugenol did not. However, when propylparaben was combined with either vanillin or eugenol, the phenylephrine disappearance was significantly inhibited. These data suggest that these compounds or combinations thereof may have potential to improve phenylephrine oral bioavailability.

Transcriptomic analysis of differentially expressed genes in flower-buds of genetic male sterile and wild type cucumber by RNA sequencing.

Cucumber (Cucumis sativus L.) pollen development involves a diverse range of gene interactions between sporophytic and gametophytic tissues. Previous studies in our laboratory showed that male sterility was controlled by a single recessive nuclear gene, and occurred in pollen mother cell meiophase. To fully explore the global gene expression and identify genes related to male sterility, a RNA-seq analysis was adopted in this study. Young male flower-buds (1-2 mm in length) from genetic male sterility (GMS) mutant and homozygous fertile cucumber (WT) were collected for two sequencing libraries. Total 545 differentially expressed genes (DEGs), including 142 up-regulated DEGs and 403 down-regulated DEGs, were detected in two libraries (Fold Change ≥ 2, FDR < 0.01). These genes were involved in a variety of metabolic pathways, like ethylene-activated signaling pathway, sporopollenin biosynthetic pathway, cell cycle and DNA damage repair pathway. qRT-PCR analysis was performed and showed that the correlation between RNA-Seq and qRT-PCR was 0.876. These findings contribute to a better understanding of the mechanism that leads to GMS in cucumber.

Suppression of cucumber stachyose synthase gene (CsSTS) inhibits phloem loading and reduces low temperature stress tolerance.

Stachyose is the main transporting sugar in phloem of Raffinose family oligosaccharides-transporting species. Stachyose synthase (STS) is a key enzyme for stachyose biosynthesis, but the gene encoding STS is poorly characterized in cucumber (Cucumis sativus L.), which is a model plant for studying stachyose metabolism and phloem function. In this research, stachyose synthase gene (CsSTS) from cucumber was isolated and its physiological functions were analyzed. CsSTS expressed mainly in the phloem of the minor veins in mature leaves and localized to companion cells. Reverse genetics with CsSTS RNAi lines revealed obviously reductions in STS activity and stachyose content along with a small amount of starch accumulation in leaves, suggesting that CsSTS is involved in phloem loading of cucumber leaves. After 6 °C low temperature stress, malondialdehyde content and electrical conductivity increased, especially in CsSTS-RNAi plants. But CsSTS expression was up-regulated, STS activity and stachyose level increased, the activities of reactive-oxygen-scavenging enzyme in cucumber seedlings improved significantly and starch accumulation reduced, especially in CsSTS-OE lines. These results demonstrate clearly that CsSTS is involved in phloem loading, carbohydrate distribution and tolerance of cucumber seedlings to low temperature stress.

The complex character of photosynthesis in cucumber fruit.

The surface area of a mature green cucumber (Cucumis sativa L.) fruit is comparable with that of a functional leaf, but the characteristics of fruit photosynthesis and its contribution to growth are poorly understood. Here, the photosynthetic properties of two genotypes of cucumber (dark green and light green fruits) were studied using a combination of electron microscopy, immunogold enzyme localization, chlorophyll fluorescence imaging, isotope tracer, and fruit darkening techniques. Chlorophyll content of the exocarp is similar to that of leaves, but there are no distinctive palisade and spongy tissues. The efficiency of PSII is similar to that in leaves, but with lower non-photochemical quenching (NPQ). Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is found mainly in the exocarp, while phosphoenolpyruvate carboxylase (PEPC) is primarily localized to vascular bundles and placenta tissue. Rubisco and PEPC expression at both transcriptional and translational levels increases concurrently during fruit growth. The contribution of fruit photosynthesis in exocarp to its own C accumulation is 9.4%, while ~88% of respiratory CO2 in fruit was captured and re-fixed. Photosynthesis by cucumber fruits, through direct fixation of atmospheric CO2 and recapture of respired CO2, as verified by 14CO2 uptake and gas exchange, makes an important contribution to fruit growth.

Down-Regulating CsHT1, a Cucumber Pollen-Specific Hexose Transporter, Inhibits Pollen Germination, Tube Growth, and Seed Development.

Efficient sugar transport is needed to support the high metabolic activity of pollen tubes as they grow through the pistil. Failure of transport results in male sterility. Although sucrose transporters have been shown to play a role in pollen tube development, the role of hexoses and hexose transporters is not as well established. The pollen of some species can grow in vitro on hexose as well as on sucrose, but knockouts of individual hexose transporters have not been shown to impair fertilization, possibly due to transporter redundancy. Here, the functions of CsHT1, a hexose transporter from cucumber (Cucumis sativus), are studied using a combination of heterologous expression in yeast (Saccharomyces cerevisiae), histochemical and immunohistochemical localization, and reverse genetics. The results indicate that CsHT1 is a plasma membrane-localized hexose transporter with high affinity for glucose, exclusively transcribed in pollen development and expressed both at the levels of transcription and translation during pollen grain germination and pollen tube growth. Overexpression of CsHT1 in cucumber pollen results in a higher pollen germination ratio and longer pollen tube growth than wild-type pollen in glucose- or galactose-containing medium. By contrast, antisense suppression of CsHT1 leads to inhibition of pollen germination and pollen tube elongation in the same medium and results in a decrease of seed number per fruit and seed size when antisense transgenic pollen is used to fertilize wild-type or transgenic cucumber plants. The important role of CsHT1 in pollen germination, pollen tube growth, and seed development is discussed.

Effect of Lixujieyu recipe in combination with Five Elements music therapy on chronic fatigue syndrome.

OBJECTIVE: To observe the clinical effects of the Lixujieyu recipe combined with Five Elements music therapy on chronic fatigue syndrome (CFS) identified as the symptom patterns of liver stagnation and spleen deficiency in terms of Traditional Chinese Medicine. METHODS: Patients with CFS were randomly divided into treatment group 1 (Lixujieyu recipe combined with Gong-Tune, n = 15); treatment group 2 (Lixujieyu recipe combined with Jiao-Tune, n = 15); treatment group 3 (Lixujieyu recipe combined with Yu-Tune, n = 15); treatment group 4 (Lixujieyu recipe combined with Shang-Tune, n = 15); treatment group 5 (Lixuiievu recipe combined with Zhi-Tune, n = 15); and the control group (Lixujieyu recipe alone, n = 15). Chinese medicine was given twice daily, and music was listened to for 45 minutes daily, 5 days a week. All patients were treated for 4 weeks. Patients were assessed via the Fatigue Scale, the Hamilton Depression Rating Scale, and the Hamilton Anxiety Rating Scale before and after treatment. RESULTS: Treatment groups 1 and 2 had better effects on relieving the symptoms of physical fatigue related to anxiety and depression than the control group (P < 0.05). CONCLUSION: Lixujieyu recipe combined with Gong-Tune or Jiao-Tune significantly relieved the symptoms of CFS.

Antisense suppression of cucumber (Cucumis sativus†L.) sucrose synthase 3 (CsSUS3) reduces hypoxic stress tolerance.

Sucrose synthase (SUS; EC 2.4.1.13) plays important roles in sugar metabolism and abiotic stress response. But the genes encoding SUS in cucumber (Cucumis sativus L.) have not been well studied. Here, we isolated four cucumber sucrose synthase genes (CsSUS). Among them, CsSUS3, which highly expressed in the roots, was chosen for further study. Immunolocalization and subcellular localization analysis indicated that CsSUS3 localized in the cytosol and the plasma membrane, and mainly existed in the companion cells of phloem in the roots. When suffering hypoxia stress from flooding, CsSUS3 expression and SUS activity in roots increased, especially in the lateral roots; moreover, the soluble SUS activity increased clearly, but the membrane fraction hardly changed. Compared with the wild-type cucumbers, the transgenic lines with antisense expression of CsSUS3 were more sensitive to flooding. After 6 d of flooding, the SUS activity, soluble sugar and uridine 5'-diphosphate glucose (UDPG) content and the ratio of ATP/ADP in the roots of transgenic plants were significantly lower than that in wild-type plants. Moreover, the transgenic lines grew more slowly with more yellow necrosis in the leaves. These findings suggested CsSUS3 participated in resisting hypoxic stress. Furthermore, the mechanism of CsSUS3 in resisting hypoxic stress was also discussed.

A Prospective, Multicentered, Randomized, Double-Blind, Placebo-Controlled Clinical Trial of Keluoxin Capsules in the Treatment of Microalbuminuria in Patients with Type 2 Early Diabetic Kidney Disease.

Background: To evaluate the efficacy and safety of Keluoxin (KLX) capsules and provide validated evidence for the application of KLX in the treatment of diabetic kidney disease (DKD). Methods: A multicenter, randomized, double-blind, placebo-controlled trial design was used to screen 129 patients with DKD (urinary albumin-to-creatinine ratio [UACR]: male, 2.5-30 mg/mmol; female, 3.5-30 mg/mmol) and with Qi and Yin deficiency and blood stasis symptoms. Written informed consent was obtained from all patients. The patients were randomly divided into KLX and control groups. The KLX group was orally administered KLX (6 g/day) and irbesartan tablets (150 mg/day), whereas the control group was administered KLX placebo (6 g/day) and irbesartan tablets (150 mg/day). Patients were observed for 24 weeks to evaluate the natural logarithm of the UACR (log-UACR), the odds ratio (OR) for a sustained increase in the UACR of at least 30% and 40%, estimated glomerular filtration rate (eGFR), changes in symptoms and quality-of-life scores, and adverse events. Results: The changes of the natural log-UACR during the 24 weeks compared with baseline in the KLX group were better than those in the control group (LS mean ± standard error, -0.26 ± 0.10 vs. 0.01 ± 0.09, p = 0.0292). The incidence of a sustained increase in the UACR of at least 30% and 40% was found to be significantly lower in the KLX group (OR, 0.26; 95% confidence interval [CI], 0.09-0.75; OR, 0.29; 95% CI, 0.10-0.82). Changes in symptoms and quality-of-life scores in the KLX group were better than those in the control group. There was no statistically significant difference in eGFR or the incidence of adverse events between the groups. Conclusions: Overall, these results suggest that KLX capsules combined with irbesartan can reduce microalbuminuria, relieve the symptoms, and improve the quality of life for patients with type 2 early DKD compared with the use of irbesartan alone. Trial registration: Chinese Clinical Trial Registry, registration number: ChiCTR2100052764.

Molecular cloning and characterization of the light-regulation and circadian-rhythm of the VDE gene promoter from Zingiber officinale.

Ginger (Zingiber officinale Rosc.) is prone to photoinhibition under intense sunlight. Excessive light can be dissipated by the xanthophyll cycle, where violaxanthin de-epoxidase (VDE) plays a critical role in protecting the photosynthesis apparatus from the damage of excessive light. We isolated ~2.0 kb of ginger VDE (GVDE) gene promoter, which contained the circadian box, I-box, G-box and GT-1 motif. Histochemical staining of Arabidopsis indicated the GVDE promoter was active in almost all organs, especially green tissues. ß-glucuronidase (GUS) activity driven by GVDE promoter was repressed rather than activated by high light. GUS activity was altered by hormones, growth regulators and abiotic stresses, which increased with 2,4-dichlorophenoxyacetic acid and decreased with abscisic acid, salicylic acid, zeatin, salt (sodium chloride) and polyethylene glycol. Interestingly, GUS activities with gibberellin or indole-3-acetic acid increased in the short-term (24 h) and decreased in the long-term (48 and 72 h). Analysis of 5' flank deletion found two crucial functional regions residing in -679 to -833 and -63 to -210. Northern blotting analysis found transcription to be regulated by the endogenous circadian clock. Finally, we found a region necessary for regulating the circadian rhythm and another for the basic promoter activity. Key message A novel promoter, named GVDE promoter, was first isolated and analyzed in this study. We have determined one region crucial for promoter activity and another responsible for keeping circadian rhythms.

Phloem unloading follows an extensive apoplasmic pathway in cucumber (Cucumis sativus L.) fruit from anthesis to marketable maturing stage.

The phloem unloading pathway remains unclear in fruits of Cucurbitaceae, a classical stachyose-transporting species with bicollateral phloem. Using a combination of electron microscopy, transport of phloem-mobile symplasmic tracer carboxyfluorescein, assays of acid invertase and sucrose transporter, and [(14)C]sugar uptake, the phloem unloading pathway was studied in cucumber (Cucumis sativus) fruit from anthesis to the marketable maturing stage. Structural investigations showed that the sieve element-companion cell (SE-CC) complex of the vascular bundles feeding fruit flesh is apparently symplasmically restricted. Imaging of carboxyfluorescein unloading showed that the dye remained confined to the phloem strands of the vascular bundles in the whole fruit throughout the stages examined. A 37 kDa acid invertase was located predominantly in the cell walls of SE-CC complexes and parenchyma cells. Studies of [(14)C]sugar uptake suggested that energy-driven transporters may be functional in sugar trans-membrane transport within symplasmically restricted SE-CC complex, which was further confirmed by the existence of a functional plasma membrane sucrose transporter (CsSUT4) in cucumber fruit. These data provide a clear evidence for an apoplasmic phloem unloading pathway in cucumber fruit. A presumption that putative raffinose or stachyose transporters may be involved in soluble sugars unloading was discussed.

The predominance of the apoplasmic phloem-unloading pathway is interrupted by a symplasmic pathway during Chinese jujube fruit development.

Phloem unloading plays a pivotal part in photoassimilate transport and partitioning into sink organs. However, it remains unclear whether the unloading pathway alters to adapt to developmental transitions in sinks, especially in fleshy fruits accumulating a high level of soluble sugars. Using a combination of electron microscopy, transport of the phloem-mobile symplasmic tracer carboxyfluorescein and assays of acid invertase, the pathway of unloading was investigated in different varieties of Chinese jujube fruit (Zizyphus jujuba Mill. cv Dongzao and Lizao). Structural investigation showed that the sieve element-companion-cell complex of bundles feeding the fruit flesh is symplasmically connected with surrounding parenchyma cells at the middle stage, and isolated during the early and late stages. Numerous plasmodesmata are present between phloem parenchyma cells and flesh cells throughout fruit development. Confocal laser scanning images of carboxyfluorescein unloading showed that the dye remained confined to the phloem strands during the early and late stages of fruit development, whereas it was released from the functional phloem at the middle stage. The levels of both the expression and activities of cell wall acid invertase and soluble acid invertase varied in an inverse pattern relative to fruit development. These results provided clear evidence for the predominance of an apoplasmic phloem unloading pathway interrupted with a symplasmic pathway and simultaneous symplasmic and apoplasmic unloading pathways in post-phloem transport during fruit development. Similar unloading pathways were obtained in different varieties of jujube fruit. The mechanisms and significance of the adaptive switch in the phloem-unloading pathway during fruit development were discussed.