Optimization of loop-mediated isothermal amplification (LAMP) assay for robust visualization in SARS-CoV-2 and emerging variants diagnosis.

Loop-mediated isothermal amplification (LAMP) is widely used in detection of pathogenic microorganisms including SARS-CoV-2. However, the performance of LAMP assay needs further exploration in the emerging SARS-CoV-2 variants test. Here, we design serials of primers and select an optimal set for LAMP-based on SARS-CoV-2 N gene for a robust and visual assay in SARS-CoV-2 diagnosis. The limit of detectable template reaches 10 copies of N gene per 25 µL reaction at isothermal 58℃ within 40 min. Importantly, the primers for LAMP assay locate at 12 to 213 nt of N gene, a highly conservative region, which serves as a compatible test in emerging SARS-CoV-2 variants. Comparison to a commercial qPCR assay, this LAMP assay exerts the high viability in diagnosis of 41 clinical samples. Our study optimizes an advantageous LAMP assay for colorimetric detection of SARS-CoV-2 and emerging variants, which is hopeful to be a promising test in COVID-19 surveillance.

Cullin1 binds and promotes NLRP3 ubiquitination to repress systematic inflammasome activation.

Activation of the NACHT, leucine-rich repeat, and pyrin domains-containing protein 3 (collectively known as NLRP3) inflammasome plays a key role in host immune response, which is the first line of defense against cellular stresses and pathogen infections. However, excessive inflammasome activation damages host cells, and therefore it must be precisely controlled. Here, we discover that Cullin1 (CUL1), a key component of the Skp1-Cullin1-F-box E3 ligase, plays a critical role in controlling the NLRP3 inflammasome. CUL1 represses inflammasome assembly in cultured cells, suppresses NLRP3 function in human monocytic cell line macrophages, and attenuates inflammatory responses in mouse model. Detailed studies demonstrate that CUL1 interacts with NLRP3 and promotes NLRP3 ubiquitination, but not protein degradation, to repress the NLRP3 inflammasome activation. Moreover, upon inflammatory stimuli, including ATP and nigericin treatments, CUL1 disassociates from NLRP3 to release the repression of the NLRP3 inflammasome. Thus, this study reveals a distinct and unique mechanism underlying the control of systematic activation of the NLRP3 inflammasome.-Wan, P., Zhang, Q., Liu, W., Jia, Y., Ai, S., Wang, T., Wang, W., Pan, P., Yang, G., Xiang, Q., Huang, S., Yang, Q., Zhang, W., Liu, F., Tan, Q., Zhang, W., Wu, K., Liu, Y., Wu, J. Cullin1 binds and promotes NLRP3 ubiquitination to repress systematic inflammasome activation.

Genome-wide identification of WRKY family genes in peach and analysis of WRKY expression during bud dormancy.

Bud dormancy in deciduous fruit trees is an important adaptive mechanism for their survival in cold climates. The WRKY genes participate in several developmental and physiological processes, including dormancy. However, the dormancy mechanisms of WRKY genes have not been studied in detail. We conducted a genome-wide analysis and identified 58 WRKY genes in peach. These putative genes were located on all eight chromosomes. In bioinformatics analyses, we compared the sequences of WRKY genes from peach, rice, and Arabidopsis. In a cluster analysis, the gene sequences formed three groups, of which group II was further divided into five subgroups. Gene structure was highly conserved within each group, especially in groups IId and III. Gene expression analyses by qRT-PCR showed that WRKY genes showed different expression patterns in peach buds during dormancy. The mean expression levels of six WRKY genes (Prupe.6G286000, Prupe.1G393000, Prupe.1G114800, Prupe.1G071400, Prupe.2G185100, and Prupe.2G307400) increased during endodormancy and decreased during ecodormancy, indicating that these six WRKY genes may play a role in dormancy in a perennial fruit tree. This information will be useful for selecting fruit trees with desirable dormancy characteristics or for manipulating dormancy in genetic engineering programs.

Umbilical Cord Mesenchymal-Stem-Cell-Derived Exosomes Exhibit Anti-Oxidant and Antiviral Effects as Cell-Free Therapies.

The oxidative stress induced by the accumulation of reactive oxygen species (ROS) can lead to cell aging and death. Equally, the skeletal muscle usually hosts enteroviral persistent infection in inflammatory muscle diseases. As excellent bioactive products, the exosomes derived from umbilical cord mesenchymal stem cells (ucMSCs) have been proven to be safe and have low immunogenicity with a potential cell-free therapeutic function. Here, exosomes derived from ucMSCs (ucMSC-EXO) were extracted and characterized. In a model of oxidative damage to skin fibroblasts (HSFs) under exposure to H2O2, ucMSC-EXO had an observable repairing effect for the HSFs suffering from oxidative damage. Furthermore, ucMSC-EXO inhibited mitogen-activated protein kinases (MAPK), c-Jun N-terminal kinase (JNK), and nuclear factor kappa-B (NF-κB) signaling pathways, thereby promoting p21 protein expression while decreasing lamin B1 protein expression, and finally alleviated oxidative stress-induced cell damage and aging. In a model of rhabdomyosarcoma (RD) cells being infected by enterovirus 71 (EV71) and coxsackievirus B3 (CVB3), the ucMSC-EXO enhanced the expression of interferon-stimulated gene 15 (ISG15) and ISG56 to inhibit enteroviral replication, whereafter reducing the virus-induced proinflammatory factor production. This study provides a promising therapeutic strategy for ucMSC-EXO in anti-oxidative stress and antiviral effects, which provides insight into extending the function of ucMSC-EXO in cell-free therapy.

The NAC transcription factor MdNAC29 negatively regulates drought tolerance in apple.

Drought stress is an adverse stimulus that affects agricultural production worldwide. NAC transcription factors are involved in plant development and growth but also play different roles in the abiotic stress response. Here, we isolated the apple MdNAC29 gene and investigated its role in regulating drought tolerance. Subcellular localization experiments showed that MdNAC29 was localized to the nucleus and transcription was induced by the PEG treatment. Over-expression of MdNAC29 reduced drought tolerance in apple plants, calli, and tobacco, and exhibited higher relative conductivity, malondialdehyde (MDA) content, and lower chlorophyll content under drought stress. The transcriptomic analyses revealed that MdNAC29 reduced drought resistance by modulating the expression of photosynthesis and leaf senescence-related genes. The qRT-PCR results showed that overexpression of MdNAC29 repressed the expression of drought-resistance genes. Yeast one-hybrid and dual-luciferase assays demonstrated that MdNAC29 directly repressed MdDREB2A expression. Moreover, the yeast two-hybrid and bimolecular fluorescence complementation assays demonstrated that MdNAC29 interacted with the MdPP2-B10 (F-box protein), which responded to drought stress, and MdPP2-B10 enhanced the repressive effect of MdNAC29 on the transcriptional activity of the MdDREB2A. Taken together, our results indicate that MdNAC29 is a negative regulator of drought resistance, and provide a theoretical basis for further molecular mechanism research.

The apple GARP family gene MdHHO3 regulates the nitrate response and leaf senescence.

The regulation of plant gene expression by nitrate is a complex regulatory process. Here, we identified 90 GARP family genes in apples by genome-wide analysis. As a member of the GARP gene family, the expression of MdHHO3 (Malus domestica HYPERSENSITIVITY TO LOW PHOSPHATE-ELICITED PRIMARY ROOT SHORTENING1 HOMOLOG 3) is upregulated under N (nitrogen) supply. The results of DNA-binding site analysis and electrophoretic mobility shift assays (EMSA) showed that MdHHO3 binds to the motif-containing GAATC. Furthermore, MdHHO3 binds to its promoter sequence and inhibits its activity. In addition, the overexpression of MdHHO3 in apple calli resulted in less accumulation of nitrate in 35S:MdHHO3-GFP calli and downregulated the expression of the nitrate transport-related genes but upregulated the expression of the nitrate assimilation-related genes. Similarly, the expression of the nitrate transport-related genes was downregulated and the expression of the nitrate assimilation-related genes was upregulated in MdHHO3 overexpression Arabidopsis and tobacco plants. Interaction experiments showed that MdHHO3 could bind to the promoter MdNRT2.1 (NITRATE TRANSPORTER 2.1) and negatively regulate its expression. Moreover, the exposure of MdHHO3-overexpressing Arabidopsis and tobacco to nitrate deficiency resulted in an early senescence phenotype as compared to the WT plants. These results show that MdHHO3 can not only negatively regulate nitrate accumulation in response to nitrate but also promote early leaf senescence under nitrate deficiency. This information may be useful to further reveal the mechanism of the nitrate response and demonstrates that nitrate deficiency induces leaf senescence in apples.

Transcriptome analysis reveals candidate genes involved in nitrogen deficiency stress in apples.

Nitrogen is one of the macroelements required for plant growth and development and the identification of candidate genes involved in nitrogen deficiency stress is of great importance to the sustainable development of agriculture. Here, we found that the color of apple leaves changed from dark green to yellow-green, the malondialdehyde (MDA) content, soluble protein content, and proline content significantly increased, the chlorophyll content significantly decreased in response to nitrate deficiency stress. According to the physiological and biochemical changes of apple leaves during nitrate deficiency stress, nitrogen deficiency stress was divided into two stages: early nitrogen deficiency stage (ES) and late nitrogen deficiency stage (LS). Transcriptome sequencing was performed in these two stress stages. 5773 differential expression genes (DEGs) were identified in the early nitrogen deficiency stress stage and 6130 DEGs were identified in the late nitrogen deficiency stress stage. Functional analysis of these DEGs revealed that a large number of DEGs were enriched in 'porphyrin and chlorophyll metabolic' pathways, the 'photosynthesis' pathway, the 'photosynthesis-antenna protein' pathway, and the 'ABA', 'ETH', and 'JA' signal transduction pathways, and the metabolic networks of these pathways were constructed. In addition, overexpression of MdNAC4 weakened the tolerance of apple calli to nitrogen deficiency stress. Taken together, our results reveal possible pathways for apple adaptation to nitrogen deficiency stress and identify the function of MdNAC4, a key transcription factor regulating nitrogen deficiency stress, which enriches the molecular mechanism of apple adapting to a nitrogen deficiency environment.

MdNAC4 Interacts With MdAPRR2 to Regulate Nitrogen Deficiency-Induced Leaf Senescence in Apple (Malus domestica).

Nitrogen (N) is one of the important macronutrients in plants, and N deficiency induces leaf senescence. However, the molecular mechanism underlying how N deficiency affects leaf senescence is unclear. Here, we report an apple NAC TF, MdNAC4, that participates in N deficiency-induced leaf senescence. The senescence phenotype of apple leaves overexpressing MdNAC4 was enhanced after N deficiency. Consistently, the chlorophyll content of transgenic leaves was significantly lower than that in the WT control leaves, the expression of chlorophyll catabolism-related genes (MdNYC1, MdPAO, and MdSGR1) was significantly higher than that in the WT controls, and the expression of chlorophyll synthesis-related genes (MdHEMA, MdCHLI, and MdCHLM) was significantly lower than that in the WT control leaves. Furthermore, MdNAC4 was found to directly activate the transcription of the chlorophyll catabolism-related genes MdNYC1 and MdPAO. Additionally, MdNAC4 was proven to interact with MdAPRR2 proteins both in vitro and in vivo, and overexpression of MdAPRR2 seemed to delay N deficiency-induced leaf senescence. Correspondingly, the chlorophyll loss of MdAPRR2-overexpressing (MdAPRR2-OE) lines was significantly lower than in WT control plants. Although downregulated, the expression of the chlorophyll synthesis-related genes MdHEMA, MdCHLI, and MdCHLM in the transgenic plants was more than twice that in the WT control plants. Taken together, our results enrich the regulatory network of leaf senescence induced by N deficiency through the interaction between MdNAC4 and MdAPRR2.

PpMYB52 negatively regulates peach bud break through the gibberellin pathway and through interactions with PpMIEL1.

Bud dormancy, which enables damage from cold temperatures to be avoided during winter and early spring, is an important adaptive mechanism of deciduous fruit trees to cope with seasonal environmental changes and temperate climates. Understanding the regulatory mechanism of bud break in fruit trees is highly important for the artificial control of bud break and the prevention of spring frost damage. However, the molecular mechanism underlying the involvement of MYB TFs during the bud break of peach is still unclear. In this study, we isolated and identified the PpMYB52 (Prupe.5G240000.1) gene from peach; this gene is downregulated in the process of bud break, upregulated in response to ABA and downregulated in response to GA. Overexpression of PpMYB52 suppresses the germination of transgenic tomato seeds. In addition, Y2H, Bimolecular fluorescence complementation (BiFC) assays verified that PpMYB52 interacts with a RING-type E3 ubiquitin ligase, PpMIEL1, which is upregulated during bud break may positively regulate peach bud break by ubiquitination-mediated degradation of PpMYB52. Our findings are the first to characterize the molecular mechanisms underlying the involvement of MYB TFs in peach bud break, increasing awareness of dormancy-related molecules to avoid bud damage in perennial deciduous fruit trees.

Prunus persica transcription factor PpNAC56 enhances heat resistance in transgenic tomatoes.

Members of the NAC (NAM, ATAF1,2 and CUC2) transcription factor family are involved in numerous processes of plant growth and development and play an important role in the response to abiotic stresses such as salinity, drought and heat, but little research on this topic has been done in peach. In this study, we analyzed the expression patterns of PpNAC56 under abiotic stress and found that PpNAC56 responded to high-temperature stress. To verify the function of PpNAC56, we overexpressed this gene in tomato plants and found that, compared with WT plants, the transgenic tomato plants could accumulate more osmoregulatory substances after high-temperature treatment and thus were more heat resistance. Then, using Y2H, BIFC, and pull-down assays, we found that PpNAC56 could interact with PpMIEL1. In addition, Y1H and dual-luciferase assays verified that PpNAC56 could activate the expression of PpHSP17.4 and PpSnRK2D. The above experimental results demonstrate that PpNAC56 plays an important role in the plant response to heat stress.

Prunus persica Terpene Synthase PpTPS1 Interacts with PpABI5 to Enhance Salt Resistance in Transgenic Tomatoes.

Terpene synthase (TPS) is related to the production of aromatic substances, but there are few studies on the impact of abiotic stress on TPS and its molecular mechanism, especially in peaches. This study found that salt resistance and abscisic acid (ABA) sensitivity of transgenic tomatoes were enhanced by overexpression of PpTPS1. Moreover, it was found that PpTPS1 interacted with and antagonized the expression of the bZIP transcription factor ABA INSENSITIVE 5 (PpABI5), which is thought to play an important role in salt suitability. In addition, PpTCP1, PpTCP13, and PpTCP15 were found to activate the expression of PpTPS1 by yeast one-hybrid (Y1H) and dual-luciferase assays, and they could also be induced by ABA. In summary, PpTPS1 may be involved in the ABA signaling regulatory pathway and play an important role in salt acclimation, providing a new reference gene for the improvement of salt resistance in peaches.

ASB17 Facilitates the Burst of LPS-Induced Inflammation Through Maintaining TRAF6 Stability.

ASB17, a member of the ankyrin repeat and SOCS box-containing protein (ASB) family, has been supposed to act as an E3 ubiquitin ligase. Actually, little is known about its biological function. In this study, we found that ASB17 knocking-out impaired the expression of the pro-inflammatory cytokines CCL2 and IL-6 in bone marrow-derived dendritic cells (BMDCs) stimulated by lipopolysaccharide (LPS), indicating an inflammation-promoting role of this gene. We reveal that ASB17 promotes LPS-induced nuclear factor kappa B (NF-κB) signal activation through interacting with TNF receptor-associated factor 6 (TRAF6) which is a crucial adaptor protein downstream of toll-like receptors (TLR). ASB17 via its aa177-250 segment interacts with the Zn finger domain of TRAF6. The interaction of ASB17 stabilizes TRAF6 protein through inhibiting K48-linked TRAF6 polyubiquitination. Therefore, we suggest that ASB17 facilitates LPS-induced NF-κB activation by maintaining TRAF6 protein stability. The inflammation enhancer role of ASB17 is recognized here, which provides new understanding of the activation process of inflammation and immune response.

Overexpression of the Peach Transcription Factor Early Bud-Break 1 Leads to More Branches in Poplar.

Shoot branching is an important adaptive trait that determines plant architecture. In a previous study, the Early bud-break 1 (EBB1) gene in peach (Prunus persica var. nectarina) cultivar Zhongyou 4 was transformed into poplar (Populus trichocarpa). PpEBB1-oe poplar showed a more branched phenotype. To understand the potential mechanisms underlying the EBB1-mediated branching, transcriptomic and proteomics analyses were used. The results showed that a large number of differentially expressed genes (DEGs)/differentially expressed proteins (DEPs) associated with light response, sugars, brassinosteroids (BR), and nitrogen metabolism were significantly enriched in PpEBB1-oe poplar. In addition, contents of sugars, BR, and amino acids were measured. Results showed that PpEBB1 significantly promoted the accumulation of fructose, glucose, sucrose, trehalose, and starch. Contents of brassinolide (BL), castasterone (CS), and 6-deoxocathasterone (6-deoxoCS) were all significantly changed with overexpressing PpEBB1. Various types of amino acids were measured and four of them were significantly improved in PpEBB1-oe poplar, including aspartic acid (Asp), arginine (Arg), cysteine (Cys), and tryptohpan (Trp). Taken together, shoot branching is a process controlled by a complex regulatory network, and PpEBB1 may play important roles in this process through the coordinating multiple metabolic pathways involved in shoot branching, including light response, phytohormones, sugars, and nitrogen.

Endodormancy Release Can Be Modulated by the GA4-GID1c-DELLA2 Module in Peach Leaf Buds.

Gibberellin (GA) plays a key role in the release of bud dormancy and the GA receptor GID1 (GIBBERELLIN INSENSITIVE DWARF1) and DELLA protein are the GA signaling parts, but the molecular mechanism of GA-GID1-DELLA module regulating leaf bud dormancy in peach (Prunus persica) is still not very clear. In this study, we isolated and characterized the GID1 gene PpGID1c from the peach cultivar "Zhong you No.4." Overexpressing PpGID1c in Arabidopsis promoted seed germination, which indicated that PpGID1c has an important function in dormancy. The expression level of PpGID1c in peach leaf buds during endodormancy release was higher than that during ecodormancy and was positively correlated with GA4 levels. Our study also found that GA4 had the most obvious effect on promoting the bud break, indicating that GA4 may be the key gibberellin to promoting peach leaf bud endodormancy release. Moreover, a quantitative real-time PCR (qRT-PCR) found that GA4 could increase the expression of the gibberellin signaling gene PpDELLA2. A yeast two-hybrid (Y2H) assay suggested that the PpGID1c interaction with the PpDELLA1 protein was not dependent on gibberellin, while the PpGID1c interaction with PpDELLA2 required GA4 or another gibberellin. These findings suggested that the GA4-GID1c-DELLA2 module regulates peach leaf bud endodormancy release, with this finding significantly enhancing our comprehensive understanding of bud endodormancy release and revealing a new mechanism for regulating leaf bud endodormancy release in peach.

BTB-TAZ Domain Protein PpBT3 modulates peach bud endodormancy by interacting with PpDAM5.

The dormancy-associated MADS-box (DAM) gene DAM5 has crucial roles in bud endodormancy; however, the molecular regulatory mechanism of PpDAM5 in peach (Prunus persica) has not been elucidated. In this study, using yeast two-hybrid screening, we isolated a BTB-TAZ Domain Protein PpBT3, which interacts with PpDAM5 protein, in the peach cultivar 'Chun xue'. As expected, we found that abscisic acid (ABA) maintained bud endodormancy and induced expression of the PpDAM5 gene, and that over-expressing PpDAM5 in Arabidopsis thaliana repressed seed germination. In contrast, over-expressing PpBT3 in A. thaliana promoted seed germination, and conferred resistance to ABA-mediated germination inhibition. Additionally, a qRT-PCR (quantitative real-time polymerase chain reaction) experiment suggested that the transcript level of PpBT3 gradually increased towards the endodormancy release period, which is the opposite trend of the expression pattern of PpDAM5. Our results suggest that PpBT3 modulates peach bud endodormancy by interacting with PpDAM5, thus revealing a new mechanism for regulating bud dormancy of perennial deciduous trees.

PpNUDX8, a Peach NUDIX Hydrolase, Plays a Negative Regulator in Response to Drought Stress.

Drought stress is a serious abiotic stress source that affects the growth and fruit quality of peach trees. However, the molecular mechanism of the NUDIX hydrolase family in peaches in response to drought stress is still unclear. Here, we isolated and identified the PpNUDX8 (Prupe.5G062300.1) gene from the peach NUDIX hydrolase family, and found that PpNUDX8 has a typical NUDIX hydrolase domain. In this study, we performed 15% PEG6000 drought treatment on peach seedlings, and qRT-PCR analysis showed that 15% PEG6000 induced the transcription level of PpNUDX8. Overexpression of PpNUDX8 reduced the tolerance of calli to 4% PEG6000 treatment. Compared with wild-type apple calli, PpNUDX8 transgenic apple calli had a lower fresh weight and higher MDA content. After 15% PEG6000 drought treatment, PpNUDX8 transgenic tobacco had a greater degree of wilting and shorter primary roots than Under control conditions. The chlorophyll, soluble protein, and proline contents in the transgenic tobacco decreased, and the MDA content and relative conductivity increased. At the same time, PpNUDX8 negatively regulated ABA signal transduction and reduced the transcriptional expression of stress response genes. In addition, PpNUDX8 was not sensitive to ABA, overexpression of PpNUDX8 reduced the expression of the ABA synthesis-related gene NCED6 and increases the expression of the ABA decomposition-related gene CYP1 in tobacco, which in turn leads to a decrease in the ABA content in tobacco. In addition, Under control conditions, overexpression of PpNUDX8 destroyed the homeostasis of NAD and reduced nicotinamide adenine dinucleotide (NADH) in tobacco. After 15% PEG6000 drought treatment, the changes in NAD and NADH in PpNUDX8 transgenic tobacco were more severe than those in WT tobacco. In addition, PpNUDX8 also interacted with PpSnRk1γ (Prupe.6G323700.1).

OVATE Family Protein PpOFP1 Physically Interacts With PpZFHD1 and Confers Salt Tolerance to Tomato and Yeast.

The OVATE family protein (OFP) genes (OFPs) have been shown to respond to salt stress in plants. However, the regulatory mechanism for salt tolerance of the peach (Prunus persica) OFP gene PpOFP1 has not been elucidated. In this study, using yeast two-hybrid screening, we isolated a nucleus-localized ZF-HD_dimer domain protein PpZFHD1, which interacts with the PpOFP1 protein in the peach cultivar "Zhongnongpan No.10". A segmentation experiment further suggested that the interaction happens more specifically between the N-terminal, contains ZF-HD_dimer domain, of PpZFHD1 and the C-terminal, consists of OVATE domain, of PpOFP1. Additionally, quantitative real-time polymerase chain reaction (qRT-PCR) experiments indicate that transcription of these two genes are induced by 200 mmol/L (mM) NaCl treatment. Heterogeneous transformation experiments suggested that the growth status of transformed yeast strain over-expressing each of these two genes was more robust than that of control (CK). Furthermore, transgenic tomato plants over-expressing PpOFP1 were also more robust. They had a higher content of chlorophyll, soluble proteins, soluble sugars, and proline. Activities of the superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in these plants were higher, and tissues from these plants exhibited a lower relative conductivity and malondialdehyde (MDA) content. These results suggest that PpOFP1 physically interacts with PpZFHD1 and confers salt tolerance to tomato and yeast, thus revealing a novel mechanism for regulating salt tolerance in peach and other perennial deciduous trees.

Chromosome-level genome assemblies of five Prunus species and genome-wide association studies for key agronomic traits in peach.

Prunus species include many important perennial fruit crops, such as peach, plum, apricot, and related wild species. Here, we report de novo genome assemblies for five species, including the cultivated species peach (Prunus persica), plum (Prunus salicina), and apricot (Prunus armeniaca), and the wild peach species Tibetan peach (Prunus mira) and Chinese wild peach (Prunus davidiana). The genomes ranged from 240 to 276 Mb in size, with contig N50 values of 2.27-8.30 Mb and 25,333-27,826 protein-coding gene models. As the phylogenetic tree shows, plum diverged from its common ancestor with peach, wild peach species, and apricot ~7 million years ago (MYA). We analyzed whole-genome resequencing data of 417 peach accessions, called 3,749,618 high-quality SNPs, 577,154 small indels, 31,800 deletions, duplications, and inversions, and 32,338 insertions, and performed a structural variant-based genome-wide association study (GWAS) of key agricultural traits. From our GWAS data, we identified a locus associated with a fruit shape corresponding to the OVATE transcription factor, where a large inversion event correlates with higher OVATE expression in flat-shaped accessions. Furthermore, a GWAS revealed a NAC transcription factor associated with fruit developmental timing that is linked to a tandem repeat variant and elevated NAC expression in early-ripening accessions. We also identified a locus encoding microRNA172d, where insertion of a transposable element into its promoter was found in double-flower accessions. Thus, our efforts have suggested roles for OVATE, a NAC transcription factor, and microRNA172d in fruit shape, fruit development period, and floral morphology, respectively, that can be connected to traits in other crops, thereby demonstrating the importance of parallel evolution in the diversification of several commercially important domesticated species. In general, these genomic resources will facilitate functional genomics, evolutionary research, and agronomic improvement of these five and other Prunus species. We believe that structural variant-based GWASs can also be used in other plants, animal species, and humans and be combined with deep sequencing GWASs to precisely identify candidate genes and genetic architecture components.

NS5 Conservative Site Is Required for Zika Virus to Restrict the RIG-I Signaling.

During host-virus co-evolution, cells develop innate immune systems to inhibit virus invasion, while viruses employ strategies to suppress immune responses and maintain infection. Here, we reveal that Zika virus (ZIKV), a re-emerging arbovirus causing public concerns and devastating complications, restricts host immune responses through a distinct mechanism. ZIKV nonstructural protein 5 (NS5) interacts with the host retinoic acid-inducible gene I (RIG-I), an essential signaling molecule for defending pathogen infections. NS5 subsequently represses K63-linked polyubiquitination of RIG-I, attenuates the phosphorylation and nuclear translocation of interferon regulatory factor 3 (IRF3), and inhibits the expression and production of interferon-ß (IFN-ß), thereby restricting the RIG-I signaling pathway. Interestingly, we demonstrate that the methyltransferase (MTase) domain of NS5 is required for the repression of RIG-I ubiquitination, IRF3 activation, and IFN-ß production. Detailed studies further reveal that the conservative active site D146 of NS5 is critical for the suppression of the RIG-I signaling. Therefore, we uncover an essential role of NS5 conservative site D146 in ZIKV-mediated repression of innate immune system, illustrate a distinct mechanism by which ZIKV evades host immune responses, and discover a potential target for anti-viral infection.

Comparison Between Flat and Round Peaches, Genomic Evidences of Heterozygosity Events.

Bud sports occur in many plant species, including fruit trees. Although they are correlated with genetic variance in somatic cells, the mechanisms responsible for bud sports are mostly unknown. In this study, a peach bud sport whose fruit shape was transformed to round from flat was identified by next generation sequencing (NGS), and we provide evidence that a long loss of heterozygosity (LOH) event may be responsible for this alteration in fruit shape. Moreover, compared to the reference genome, we identified 237,476 high quality single nucleotide polymorphisms (SNPs) in the wild-type and bud sport genomes. Using this SNP set, a long LOH event was identified at the distal end of scaffold Pp06 of the bud sport genome. Haplotypes from 155 additional peach accessions were phased, suggesting that the homozygous distal end of scaffold Pp06 of the bud sport was likely derived from only one haplotype of the wild-type flat peach. A genome-wide association study (GWAS) of 127 peach accessions was conducted to associate a SNP found at 26,924,482 bp of scaffold Pp06 to differences in fruit shape. All accessions with round-shaped fruit were found to have an A/A genotype, while those with A/T, or T/T genotypes had flat-shaped fruits. Finally, we also found that 236 peach accessions and 141 Prunus species with round-type fruit were found to have an A/A genotype at this SNP, while 22 flat peach accessions had an A/T genotype. Taken together, our results suggest that genes flanking this A/T polymorphism, and haplotyped carrying the T allele may determine flat fruit shape in this population. Furthermore, the LOH event resulting in the loss of the haplotype carrying the T allele may therefore be responsible for fruit shape alteration in wild-type flat peach.