Genome-Wide Identification and Expression Pattern of Sugar Transporter Genes in the Brown Planthopper, Nilaparvata lugens (Stål).

Sugar transporters play important roles in controlling carbohydrate transport and are responsible for mediating the movement of sugars into cells in numerous organisms. In insects, sugar transporters not only play a role in sugar transport but may also act as receptors for virus entry and the accumulation of plant defense compounds. The brown planthopper, Nilaparvata lugens, inflicts damage on rice plants by feeding on their phloem sap, which is rich in sugars. In the present study, we identified 34 sugar transporters in N. lugens, which were classified into three subfamilies based on phylogenetic analysis. The motif numbers varied from seven to eleven, and motifs 2, 3, and 4 were identified in the functional domains of all 34 NlST proteins. Chromosome 1 was found to possess the highest number of NlST genes, harboring 15. The gut, salivary glands, fat body, and ovary were the different tissues enriched with NlST gene expression. The expression levels of NlST2, 3, 4, 7, 20, 27, 28, and 31 were higher in the gut than in the other tissues. When expressed in a Saccharomyces cerevisiae hexose transporter deletion mutant (strain EBY.VW4000), only ApST4 (previously characterized) and NlST4, 28, and 31 were found to transport glucose and fructose, resulting in functional rescue of the yeast mutant. These results provide valuable data for further studies on sugar transporters in N. lugens and lay a foundation for finding potential targets to control N. lugens.

A secure cross-domain interaction scheme for blockchain-based intelligent transportation systems.

In the intelligent transportation system (ITS), secure and efficient data communication among vehicles, road testing equipment, computing nodes, and transportation agencies is important for building a smart city-integrated transportation system. However, the traditional centralized processing approach may face threats in terms of data leakage and trust. The use of distributed, tamper-proof blockchain technology can improve the decentralized storage and security of data in the ITS network. However, the cross-trust domain devices, terminals, and transportation agencies in the heterogeneous blockchain network of the ITS still face great challenges in trusted data communication and interoperability. In this article, we propose a heterogeneous cross-chain interaction mechanism based on relay nodes and identity encryption to solve the problem of data cross-domain interaction between devices and agencies in the ITS. First, we propose the ITS cross-chain communication framework and improve the cross-chain interaction model. The relay nodes are interconnected through libP2P to form a relay node chain, which is used for cross-chain information verification and transmission. Secondly, we propose a relay node secure access scheme based on identity-based encryption to provide reliable identity authentication for relay nodes. Finally, we build a standard cross-chain communication protocol and cross-chain transaction lifecycle for this mechanism. We use Hyperledger Fabric and FISCO BCOS blockchain to design and implement this solution, and verify the feasibility of this cross-chain interaction mechanism. The experimental results show that the mechanism can achieve a stable data cross-chain read throughput of 2,000 transactions per second, which can meet the requirements of secure and efficient cross-chain communication and interaction among heterogeneous blockchains in the ITS, and has high application value.

SlbHLH152, a bHLH transcription factor positively regulates iron homeostasis in tomato.

The maintain of iron (Fe) homeostasis is essential for plant survival. In tomato, few transcription factors have been identified as regulators of Fe homeostasis, among which SlbHLH068 induced by iron deficiency, plays an important role. However, the upstream regulator(s) responsible for activating the expression of SlbHLH068 remain(s) unknown. In this study, the bHLH (basic helix-loop-helix) transcription factor SlbHLH152 was identified as an upstream regulator of SlbHLH068 using yeast one-hybrid screening. Deletion of SlbHLH152 led to a significant decline in Fe concentration, which was accompanied by reduced expression of Fe-deficiency-responsive genes. In contrast, SlbHLH152 overexpression plants displayed tolerance to iron deficiency, increased Fe accumulation, and elevated expression of Fe-deficiency-responsive genes. Further analysis indicated that SlbHLH152 directly activates the transcription of SlbHLH068. Taken together, our results suggest that SlbHLH152 may be involved in the regulation of iron homeostasis by directly activating the transcription of SlbHLH068 in tomato.

Co-silencing of ABA receptors (SlRCAR) reveals interactions between ABA and ethylene signaling during tomato fruit ripening.

The ripening of fleshy fruits is highly dependent on the regulation of endogenous hormones, including ethylene, abscisic acid (ABA) and other phytohormones. However, the regulatory mechanism of ABA signaling and its interaction with ethylene signaling in fruit ripening are still unclear. In this study, multi-gene interference (RNAi) was applied to silence the ABA receptor genes in tomato for screening the specific receptors that mediate ABA signaling during fruit ripening. The results indicated that the ABA receptors, including SlRCAR9, SlRCAR12, SlRCAR11, and SlRCAR13, participate in the regulation of tomato fruit ripening. Comparative analysis showed that SlRCAR11 and SlRCAR13 play more important roles in mediating ABA signaling during tomato fruit ripening. Co-silencing of the four genes encoding these receptors could weaken the ethylene biosynthesis and signaling pathway at the early stage of tomato fruit ripening, leading to delayed fruit ripening. Meanwhile, co-silencing enhanced fruit firmness, and altered the shelf-life and susceptibility to Botrytis cinerea of the transgenic fruits. Furthermore, blocking ABA signaling did not affect the ability of ethylene to induce fruit ripening, whereas the block may inhibit the effectiveness of ABA in promoting fruit ripening. These results suggested that ABA signaling may be located upstream of ethylene signaling in regulating fruit ripening. Our findings provide a new insight into the complex regulatory network of phytohormones in regulating fruit ripening in tomato.

SlNAC6, A NAC transcription factor, is involved in drought stress response and reproductive process in tomato.

Tomato plants are susceptible to drought stress, but the mechanism involved in this process still remains poorly understood. In the present study, we demonstrated that SlNAC6, a nuclear-localized protein induced by exogenous abscisic acid (ABA) or polyethylene glycol (PEG) stress treatment, plays a positive role in tomato plant response to PEG stress. Down-regulation of SlNAC6 (SlNAC6-RNAi) resulted in a semidwarf phenotype, and the SlNAC6-RNAi lines showed reduced tolerance to PEG stress, exhibiting a higher water loss rate and degree of oxidative damage, as well as lower values of proline content and antioxidant enzyme activity, when compared with those in wild type (WT). In contrast, overexpression of SlNAC6 (SlNAC6-OE) leads to a significant delay of growth, and the SlNAC6-OE lines showed greatly enhanced tolerance to PEG stress concomitant with a lower water loss rate and degree of oxidative damage, as well as higher values of proline content and antioxidant enzyme activity. Further study showed that the transcription level of ABA signaling-related genes and the ABA content are respectively decreased or increased in SlNAC6-RNAi and SlNAC6-OE seedlings, as verified by multiple physiological parameters, such as stomatal conductance, water loss rate, seed germination, and root length. Moreover, overexpression of SlNAC6 can accelerate tomato fruit ripening. Collectively, this study demonstrates SlNAC6 exerts important roles in tomato development, drought stress response, and fruit ripening processes, some of them perhaps partly through modulating an ABA-mediated pathway, which implies SlNAC6 may hold the potential applications in improving agronomic traits of tomato or other Solanaceae crops.

Genome-Wide Identification and Expression Analysis of Tomato ADK Gene Family during Development and Stress.

Adenylate kinase (ADK) is widely distributed in organisms and plays an important role in cellular energy homeostasis. In plants, ADK has important functions in plant growth and development regulation as well as in adaptation to the environment. However, little information is available about the ADK genes in tomato (Solanum lycopersicum), an important economic crop. To investigate the characteristics and functions of ADK genes in tomato, a total of 11 ADK genes were identified and named according to their chromosomal locations. The ADK family in Arabidopsis, tomato, potato, and rice was divided into six groups, and motif analysis revealed that each SlADK protein contained five to eight conserved motifs. A total of 4 to 19 exons were identified in tomato ADK gene family members, and interestingly, most members possessed 4 exons. Several stress response elements were identified in the promoter regions of SlADKs. The 11 SlADKs were randomly distributed on 9 of the 12 tomato chromosomes. Three duplication events were observed between tomato chromosomes, and a high degree of conservation of synteny was demonstrated between tomato and potato. The online TomExpress platform prediction revealed that SlADKs were expressed in various tissues and organs, basically consistent with the data obtained from real-time quantitative PCR (qPCR). The qPCR verification was also performed to determine the expression level of SlADKs and demonstrated that the genes responded to multiple abiotic stresses, such as drought, salt, and cold. Besides, the qPCR results showed that SlADK transcription was responsive to most of the applied hormone treatment. For correlation network analysis under 44 global conditions, the results showed that the number of 17, 3, 4, and 6 coexpressed genes matched with SlADK5, 8, 9, and 11, respectively. For specific gene function analysis, expression of SlADK10 was inhibited using virus-induced gene silencing (VIGS). Compared to wild-type plants, plants with silenced SlADK10 gene had poor drought resistance, indicating SlADK10 regulated drought tolerance of tomato positively. In summary, the information provided in the present study will be helpful to understand the evolutionary relationship and their roles of tomato ADK gene family in further research.

Tomato calmodulin-like protein SlCML37 is a calcium (Ca2+) sensor that interacts with proteasome maturation factor SlUMP1 and plays a role in tomato fruit chilling stress tolerance.

Calmodulin-like proteins (CMLs), as well as their targets, play significant roles in various key developmental and stress responses in the plant. In tomato (Solanum lycopersicum), there are at least 52 CML genes in its genome. However, most of their functions are not well known, especially in response to cold stress. Here, we investigated SlCML37 biochemical and structural characteristics, including a typical α-helical secondary structure and exposing its hydrophobic regions after binding to Ca2+. Then we certificated that SlCML37 protein could physically interact with SlUMP1 by using yeast two-hybrid, bimolecular florescence complementation (BiFC) and GST pull-down assays. Further analysis showed that SlCML37-transgenic tomato fruit conferred significantly improved tolerance to chilling stress. This study indicates a possible role of calmodulin-like protein-mediated proteasome assemble in the regulation of plant cold response.

Tomato transcriptional repressor MYB70 directly regulates ethylene-dependent fruit ripening.

Ethylene is a key plant hormone controlling the ripening of climacteric fruits, and several transcription factors acting as important regulators of fruit ripening have been identified in tomato (Solanum lycopersicum), a model for climacteric fruits. The vast majority of these transcription factors are transcriptional activators, however, and the associated transcriptional regulatory mechanisms of most regulators are unclear. Here, we report on a tomato transcriptional repressor (termed SlMYB70) that negatively regulates fruit ripening by directly modulating ethylene biosynthesis. As an EAR motif-containing MYB transcription factor-encoding gene, SlMYB70 displayed a ripening-associated expression pattern and was responsive to ethylene. RNA interference (RNAi)-mediated repression of SlMYB70 accelerated fruit ripening, but overexpression of SlMYB70 delayed fruit ripening. Ethylene production was noticeably increased and decreased in SlMYB70-RNAi and SlMYB70-overexpressing lines, respectively, compared with wild-type tomatoes. SlMYB70 was proven to be a transcriptional repressor, dependent on the EAR repression motif, and to repress the transcription of two ethylene biosynthesis genes in fruit ripening, namely SlACS2 and SlACO3. The promoters of SlACS2 and SlACO3 are directly bound by SlMYB70, which was verified using a combination of yeast one-hybrid chromatin immunoprecipitation quantitative polymerase chain reaction and electrophoretic mobility shift assays. These results suggest that SlMYB70 negatively regulates fruit ripening via the direct transcriptional repression of ethylene biosynthesis genes, which provides insights into the ethylene-mediated key regulatory hierarchy in climacteric fruit ripening, and also highlights different types of transcriptional regulation of fruit ripening.

SlMYB72 Regulates the Metabolism of Chlorophylls, Carotenoids, and Flavonoids in Tomato Fruit.

Tomato (Solanum lycopersicum) fruit ripening is accompanied by the degradation of chlorophylls and the accumulation of carotenoids and flavonoids. Tomato SlMYB72 belongs to the R2R3 MYB subfamily, is located in the nucleus, and possesses transcriptional activator activity. Down-regulation of the SlMYB72 gene produced uneven-colored fruits; that is, dark green spots appeared on immature and mature green fruits, whereas yellow spots appeared on red fruits. Down-regulation of SlMYB72 increased chlorophyll accumulation, chloroplast biogenesis and development, and photosynthesis rate in fruits. This down-regulation decreased lycopene content, promoted ß-carotene production and chromoplast development, and increased flavonoid accumulation in fruits. RNA sequencing analysis revealed that down-regulation of SlMYB72 altered the expression levels of genes involved in the biosynthesis of chlorophylls, carotenoids, and flavonoids. SlMYB72 protein interacted with the auxin response factor SlARF4. SlMYB72 directly targeted protochlorophyllide reductase, Mg-chelatase H subunit, and knotted1-like homeobox2 genes and regulated chlorophyll biosynthesis and chloroplast development. SlMYB72 directly bound to phytoene synthase, ζ-carotene isomerase, and lycopene ß-cyclase genes and regulated carotenoid biosynthesis. SlMYB72 directly targeted 4-coumarate-coenzyme A ligase and chalcone synthase genes and regulated the biosynthesis of flavonoids and phenolic acid. The uneven color phenotype in RNA interference-SlMYB72 fruits was due to uneven silencing of SlMYB72 and uneven expression of chlorophyll, carotenoid, and flavonoid biosynthesis genes. In summary, this study identified important roles for SlMYB72 in the regulation of chlorophyll, carotenoid, and flavonoid metabolism and provided a potential target to improve fruit nutrition in horticultural crops.

Citrus Postharvest Green Mold: Recent Advances in Fungal Pathogenicity and Fruit Resistance.

As the major postharvest disease of citrus fruit, postharvest green mold is caused by the necrotrophic fungus Penicillium digitatum (Pd), which leads to huge economic losses worldwide. Fungicides are still the main method currently used to control postharvest green mold in citrus fruit storage. Investigating molecular mechanisms of plant-pathogen interactions, including pathogenicity and plant resistance, is crucial for developing novel and safer strategies for effectively controlling plant diseases. Despite fruit-pathogen interactions remaining relatively unexplored compared with well-studied leaf-pathogen interactions, progress has occurred in the citrus fruit-Pd interaction in recent years, mainly due to their genome sequencing and establishment or optimization of their genetic transformation systems. Recent advances in Pd pathogenicity on citrus fruit and fruit resistance against Pd infection are summarized in this review.

MiR-539-5p alleviates sepsis-induced acute lung injury by targeting ROCK1.

INTRODUCTION: Sepsis-induced acute lung injury (ALI) is an inflammatory process involved with simultaneous production of inflammatory cytokines and chemokines. In this study, we investigated the regulatory role of miR-539-5p in sepsis-induced ALI using a mouse model of cecal ligation puncture (CLP) and an in vitro model of primary murine pulmonary microvascular endothelial cells (MPVECs). MATERIAL AND METHODS: Adult male C57BL/6 mice were intravenously injected with or without miR-539-5p agomir or scrambled control one week before CLP operation. MPVECs were transfected with miR-539-5p mimics or control mimics, followed by lipopolysaccharide (LPS) stimulation. ROCK1 was predicted and confirmed as a direct target of miR-539-5p using dual-luciferase reporter assay. In rescue experiment, MPVECs were co-transfected with lentiviral vector expressing ROCK1 (or empty vector) and miR-539-5p mimics 24 h before LPS treatment. The transcriptional activity of caspase-3, the apoptosis ratio, the levels of miR-539-5p, interleukin-1b (IL-1b), interleukin-6 (IL-6), and ROCK1 were assessed. RESULTS: Compared to sham group, mice following CLP showed pulmonary morphological abnormalities, elevated production of IL-1b and IL-6, and increased caspase-3 activity and apoptosis ratio in the lung. In MPVECs, LPS stimulation resulted in a significant induction of inflammatory cytokine levels and apoptosis compared to untreated cells. The overexpression of miR-539-5p in septic mice alleviated sepsis-induced pulmonary injury, apoptosis, and inflammation. MiR-539-5p also demonstrated anti-apoptotic and anti-inflammatory effect in LPS-treated MPVECs. The upregulation of ROCK1 in MPVECs recovered miR-539-5p-suppressed caspase-3 activity and proinflammatory cytokine production. CONCLUSION: In conclusion, miR-539-5p alleviated sepsis-induced ALI via suppressing its downstream target ROCK1, suggesting a therapeutic potential of miR-539-5p for the management of sepsis-induced ALI.

SlMYB75, an MYB-type transcription factor, promotes anthocyanin accumulation and enhances volatile aroma production in tomato fruits.

Genetic manipulation of genes to upregulate specific branches of metabolic pathways is a method that is commonly used to improve fruit quality. However, the use of a single gene to impact several metabolic pathways is difficult. Here, we show that overexpression of the single gene SlMYB75 (SlMYB75-OE) is effective at improving multiple fruit quality traits. In these engineered fruits, the anthocyanin content reached 1.86 mg g-1 fresh weight at the red-ripe stage, and these SlMYB75-OE tomatoes displayed a series of physiological changes, including delayed ripening and increased ethylene production. In addition to anthocyanin, the total contents of phenolics, flavonoids and soluble solids in SlMYB75-OE fruits were enhanced by 2.6, 4, and 1.2 times, respectively, compared to those of wild-type (WT) fruits. Interestingly, a number of aroma volatiles, such as aldehyde, phenylpropanoid-derived and terpene volatiles, were significantly increased in SlMYB75-OE fruits, with some terpene volatiles showing more than 10 times higher levels than those in WT fruits. Consistent with the metabolic assessment, transcriptomic profiling indicated that the genes involved in the ethylene signaling, phenylpropanoid and isoprenoid pathways were greatly upregulated in SlMYB75-OE fruits. Yeast one-hybrid and transactivation assays revealed that SlMYB75 is able to directly bind to the MYBPLANT and MYBPZM cis-regulatory elements and to activate the promoters of the LOXC, AADC2 and TPS genes. The identification of SlMYB75 as a key regulator of fruit quality attributes through the transcriptional regulation of downstream genes involved in several metabolic pathways opens new avenues towards engineering fruits with a higher sensory and nutritional quality.

The MADS-Box Gene SlMBP21 Regulates Sepal Size Mediated by Ethylene and Auxin in Tomato.

Normal organ size is achieved by successful co-ordination of cell proliferation and cell expansion, which are modulated by multiple factors such as ethylene and auxin. In our work, SlMBP21-RNAi (RNA interference) tomato exhibited longer sepals and improved fruit set. Histological analysis indicated that longer sepals were attributed to cell expansion. To explore how SlMBP21 regulates sepal size, we compared the transcriptomes of sepals between SlMBP21-RNAi and the wild type by RNA sequencing and found that the differentially expressed genes were dominantly related to cell expansion, ethylene and auxin, and photosynthesis. Down-regulation of SlMBP21 affected ethylene production and the free IAA and IAA-Val intensity in sepals. Hormone treatment further indicated that SlMBP21 was involved in the ethylene and auxin pathways. As reported, ethylene and auxin were important factors for cell expansion. Hence, SlMBP21 negatively regulated cell expansion to control sepal size, and ethylene and auxin may mediate this process. Additionally, the contents of Chl and the activity of ribulose-1, 5-bisphosphate carboxylase/oxygenase, the key photosynthetic enzyme, were both increased in SlMBP21-RNAi sepals, which indicated that photosynthesis might be enhanced in transgenic longer sepals. Therefore, the longer sepal, with better protection and enhanced photosynthesis, may contribute to improve fruit set. Altogether, these results suggested that SlMBP21 was a novel factor involved in organ size control. Moreover, our study provided potential application value for improving fruit set.