CmCNIH1 improves salt tolerance by influencing the trafficking of CmHKT1;1 in pumpkin.

Pumpkin is often used as a rootstock for other Cucurbitaceae crops due to its resistance to soil-borne diseases and abiotic stress. Pumpkin rootstocks use a sodium transporter (CmHKT1;1) to promote the transport of Na+ from the shoot to the root effectively and improve the salt tolerance of the scion. However, the molecular regulatory mechanisms that influence the activity of CmHKT1;1 during salt stress response remain unknown. In this study, CmCNIH1, a cornichon homolog, was identified as a potential cargo receptor for CmHKT1;1. Yeast two-hybrid, biomolecular fluorescence complementation and luciferase complementary assays demonstrated that CmCNIH1 and CmHKT1;1 could interact. CmCNIH1 was a key component of the cellular vesicle transport machinery located in the endoplasmic reticulum (ER), ER export site and Golgi apparatus. A CmCNIH1 knockout mutant was more sensitive to salt stress than the wild-type (WT). In addition, ion homeostasis was disrupted in cmcnih1 mutants, which had higher Na+ and lower K+ content in shoots and roots than the WT. Two-electrode voltage-clamp experiment displayed that CmCNIH1 could not influence the Na+ current that passed through the plasma membrane (PM) in CmHKT1;1-expressing Xenopus laevis oocytes. Data from co-localization assays indicated that intact CmCNIH1 protein could alter the subcellular localization of CmHKT1;1 in tobacco leaf, pumpkin root and yeast. In summary, CmCNIH1 may function as a cargo receptor that regulates the localization of CmHKT1;1 to the PM to improve salt tolerance in pumpkin.

CeO2 nanoparticles improved cucumber salt tolerance is associated with its induced early stimulation on antioxidant system.

Salinity is a global issue limiting efficient agricultural production. Nano-enabled plant salt tolerance is a hot topic. However, the role of nanoparticles induced possible early stimulation on antioxidant system in its improved plant salt tolerance is still largely unknown. Here, poly (acrylic) acid coated nanoceria (cerium oxide nanoparticles) (PNC, 7.8 nm, -31 mV) with potent ROS (reactive oxygen species) scavenging ability are used. Compared with control, no significant difference of H2O2 and O2•─ content, MDA (malondialdehyde) content, relative electric conductivity, and Fv/Fm was found in leaves and/or roots of cucumber before onset of salinity stress, regardless of leaf or root application of PNC. While, before onset of salinity stress, compared with control, the activities of SOD (superoxide dismutase, up to 1.8 folds change), POD (peroxidase, up to 2.5 folds change) and CAT (catalase, up to 2.3 folds change), and the content of GSH (glutathione, up to 3.0 folds change) and ASA (ascorbic acid, up to 2.4 folds change) in leaves and roots of cucumber with PNC leaf spray or root application were significantly increased. RNA seq analysis further confirmed that PNC foliar spray upregulates more genes in leaves over roots than the root application. These results showed that foliar sprayed PNC have stronger early stimulation effect on antioxidant system than the root applied one and leaf are more sensitive to PNC stimulation than root. After salt stress, cucumber plants with foliar sprayed PNC showed better improvement in salt tolerance than the root applied one. Also, plants with foliar sprayed PNC showed significant higher whole plant cerium content than the root applied one after salt stress. In summary, we showed that foliar spray of nanoceria is more optimal than root application in terms of improving cucumber salt tolerance, and this improvement is associated with better stimulation on antioxidant system in plants.

CmRCC1 Gene From Pumpkin Confers Cold Tolerance in Tobacco by Modulating Root Architecture and Photosynthetic Activity.

Low-temperature stress is the main limiting factor of cucurbit crop cultivation as it affects crop yield and quality. The identification of genes involved in cold tolerance is a crucial aspect of pumpkin rootstock breeding. Here, we examined the function of a pumpkin Regulator of Chromosome Condensation 1 (CmRCC1) gene in the root development and cold stress responses of tobacco (Nicotiana benthamiana). CmRCC1 expression was differentially induced in pumpkin root, stem, and leaf under cold stress. Transient transformation showed that CmRCC1 is located in the nucleus. CmRCC1 overexpression in tobacco increased the gravitropic set-point angle in lateral roots, as well as root diameter and volume. The expression of auxin polar transport factors, PIN1 and PIN3, decreased and increased in CmRCC1-overexpressed plants, respectively. Yeast two-hybrid verification and luciferase complementation imaging assay showed that CmRCC1 interacts with CmLAZY1. Furthermore, the decreases in maximum quantum yield of PS II, the effective quantum yield of PS II, and electron transfer rate and the increases in quantum yield of nonregulated energy dissipation and malondialdehyde content were compromised in transgenic plants compared with wild-type plants under cold stress. The results suggest that CmRCC1 plays an important role in the regulation of root architecture and positively modulates cold tolerance.

Comparative analysis of volatile compounds in thirty nine melon cultivars by headspace solid-phase microextraction and gas chromatography-mass spectrometry.

The types and amounts of volatiles in the fruits of 39 melon cultivars were determined. We identified 146 volatiles, including 55 esters, 23 aldehydes, 30 alcohols, 15 ketones, 6 acids and 17 others. Ethyl acetate, (Z)-6-nonenal and 3,6-(E,Z)-nonadien-1-ol were the most three abundant volatiles (average content > 50 µg/kg FW). Aroma profiles showed significant differences among cultivars. Zhongtian49 and Zhongtian20 had the most abundant aroma components (76) and Jinguniang exhibited the least (23). One non-climacteric inodorus cultivar (Xizhoumi25) had the highest content of total volatiles (1840 µg/kg FW). Principal component analysis clustered the 39 melon cultivars into five groups. This work describes the comparative diversity of melon fruit volatiles for a large number of cultivars. Furthermore, this study could support the selection of cultivars with a flavor that suits the public and also future breeding work towards the genetic improvement of melon flavor.

Pumpkin CmHKT1;1 Controls Shoot Na⁺ Accumulation via Limiting Na⁺ Transport from Rootstock to Scion in Grafted Cucumber.

Soil salinity adversely affects the growth and yield of crops, including cucumber, one of the most important vegetables in the world. Grafting with salt-tolerant pumpkin as the rootstock effectively improves the growth of cucumber under different salt conditions by limiting Na⁺ transport from the pumpkin rootstock to the cucumber scion. High-affinity potassium transporters (HKTs) are crucial for the long distance transport of Na⁺ in plants, but the function of pumpkin HKTs in this process of grafted cucumber plants remains unclear. In this work, we have characterized CmHKT1;1 as a member of the HKT gene family in Cucurbita moschata and observed an obvious upregulation of CmHKT1;1 in roots under NaCl stress conditions. Heterologous expression analyses in yeast mutants indicated that CmHKT1;1 is a Na⁺-selective transporter. The transient expression in tobacco epidermal cells and in situ hybridization showed CmHKT1;1 localization at plasma membrane, and preferential expression in root stele. Moreover, ectopic expression of CmHKT1;1 in cucumber decreased the Na⁺ accumulation in the plants shoots. Finally, the CmHKT1;1 transgenic line as the rootstock decreased the Na⁺ content in the wild type shoots. These findings suggest that CmHKT1;1 plays a key role in the salt tolerance of grafted cucumber by limiting Na⁺ transport from the rootstock to the scion and can further be useful for engineering salt tolerance in cucurbit crops.

Ectopic Expression of Pumpkin NAC Transcription Factor CmNAC1 Improves Multiple Abiotic Stress Tolerance in Arabidopsis.

Drought, cold and salinity are the major environmental stresses that limit agricultural productivity. NAC transcription factors regulate the stress response in plants. Pumpkin (Cucurbita moschata) is an important cucurbit vegetable crop and it has strong resistance to abiotic stress; however, the biological functions of stress-related NAC genes in this crop are largely unknown. This study reports the function of CmNAC1, a stress-responsive pumpkin NAC domain protein. The CmNAC1-GFP fusion protein was transiently expressed in tobacco leaves for subcellular localization analysis, and we found that CmNAC1 is localized in the nucleus. Transactivation assay in yeast cells revealed that CmNAC1 functions as a transcription activator, and its transactivation domain is located in the C-terminus. CmNAC1 was ubiquitously expressed in different organs, and its transcript was induced by salinity, cold, dehydration, H2O2, and abscisic acid (ABA) treatment. Furthermore, the ectopic expression (EE) of CmNAC1 in Arabidopsis led to ABA hypersensitivity and enhanced tolerance to salinity, drought and cold stress. In addition, five ABA-responsive elements were enriched in CmNAC1 promoter. The CmNAC1-EE plants exhibited different root architecture, leaf morphology, and significantly high concentration of ABA compared with WT Arabidopsis under normal conditions. Our results indicated that CmNAC1 is a critical factor in ABA signaling pathways and it can be utilized in transgenic breeding to improve the abiotic stress tolerance of crops.

Scanning ion-selective electrode technique and X-ray microanalysis provide direct evidence of contrasting Na+ transport ability from root to shoot in salt-sensitive cucumber and salt-tolerant pumpkin under NaCl stress.

Grafting onto salt-tolerant pumpkin rootstock can increase cucumber salt tolerance. Previous studies have suggested that this can be attributed to pumpkin roots with higher capacity to limit the transport of Na(+) to the shoot than cucumber roots. However, the mechanism remains unclear. This study investigated the transport of Na(+) in salt-tolerant pumpkin and salt-sensitive cucumber plants under high (200 mM) or moderate (90 mM) NaCl stress. Scanning ion-selective electrode technique showed that pumpkin roots exhibited a higher capacity to extrude Na(+), and a correspondingly increased H(+) influx under 200 or 90 mM NaCl stress. The 200 mM NaCl induced Na(+)/H(+) exchange in the root was inhibited by amiloride (a Na(+)/H(+) antiporter inhibitor) or vanadate [a plasma membrane (PM) H(+) -ATPase inhibitor], indicating that Na(+) exclusion in salt stressed pumpkin and cucumber roots was the result of an active Na(+)/H(+) antiporter across the PM, and the Na(+)/H(+) antiporter system in salt stressed pumpkin roots was sufficient to exclude Na(+) X-ray microanalysis showed higher Na(+) in the cortex, but lower Na(+) in the stele of pumpkin roots than that in cucumber roots under 90 mM NaCl stress, suggesting that the highly vacuolated root cortical cells of pumpkin roots could sequester more Na(+), limit the radial transport of Na(+) to the stele and thus restrict the transport of Na(+) to the shoot. These results provide direct evidence for pumpkin roots with higher capacity to limit the transport of Na(+) to the shoot than cucumber roots.

Cinnamic acid-inhibited ribulose-1,5-bisphosphate carboxylase activity is mediated through decreased spermine and changes in the ratio of polyamines in cowpea.

This study investigated the effects of cinnamic acid (CA) on ribulose-1,5-bisphosphate carboxylase (RuBPC) activity and the endogenous polyamine levels of cowpea leaves. The results show that 0.1 mM CA treatment decreased photosynthetic rate (P(n)) and RuBPC activity, but it did not affect the maximal photochemical efficiency of PSII (F(v)/F(m)), the actual photochemical efficiency of PSII (PhiPSII), intercellular CO(2) concentration (C(i)), and relative chlorophyll content. These suggest that the decrease in P(n) is at least partially attributed to a lowered RuBPC activity. In addition, 0.1 mM CA treatment increased the putrescine (Put) level, but decreased spermidine (Spd) and spermine (Spm) levels, thereby reducing the (Spd+Spm)/Put (PAs) ratio in the leaves. The exogenous application of 1 mM Spd markedly reversed these CA-induced effects for polyamine and partially restored the PAs ratio and RuBPC activity in leaves. Methylglyoxal-bis (guanylhydrazone) (MGBG), which is an inhibitor of S-adenosylmethionine decarboxylase (SAMDC), results in the inability of activated cells to synthesize Spd and exacerbates the negative effects induced by CA. The exogenous application of 1 mM D-arginine (D-Arg), which is an inhibitor of Put biosynthesis, decreased the levels of Put, but increased the PAs ratio and RuBPC activity in leaves. These results suggest that 0.1 mM CA inhibits RuBPC activity by decreasing the levels of endogenous free and perchloric acid soluble (PS) conjugated Spm, as well as the PAs ratio.