BpWOX11 promotes adventitious root formation in Betula pendula.

Adventitious root formation is a key step in vegetative propagation via cuttings. It is crucial for establishing birch plantations and preserve birch varieties. Although previous studies have highlighted role of WOX11 in controlling adventitious root formation, no such study has been conducted in birch. Understanding the mechanism of adventitious root formation is essential for improvement of rooting or survival rate using stem cuttings in birch. In this study, we cloned BpWOX11 and produced BpWOX11 overexpression (OE) transgenic lines using the Agrobacterium-mediated plant transformation. OE lines exhibited early initiated adventitious root formation, leading to increase the rooting rate of stem cuttings plants. RNA sequencing analysis revealed that OE lines induced the gene expression related to expansin and cell division pathway, as well as defense and stress response genes. These may be important factors for the BpWOX11 gene to promote adventitious root formation in birch cuttings. The results of this study will help to further understand the molecular mechanisms controlling the formation of adventitious roots in birch.

Genome-wide identification of the TIFY family reveals JAZ subfamily function in response to hormone treatment in Betula platyphylla.

BACKGROUND: The TIFY family is a plant-specific gene family and plays an important role in plant growth and development. But few reports have been reported on the phylogenetic analysis and gene expression profiling of TIFY family genes in birch (Betula platyphylla). RESULTS: In this study, we characterized TIFY family and identified 12 TIFY genes and using phylogeny and chromosome mapping analysis in birch. TIFY family members were divided into JAZ, ZML, PPD and TIFY subfamilies. Phylogenetic analysis revealed that 12 TIFY genes were clustered into six evolutionary branches. The chromosome distribution showed that 12 TIFY genes were unevenly distributed on 5 chromosomes. Some TIFY family members were derived from gene duplication in birch. We found that six JAZ genes from JAZ subfamily played essential roles in response to Methyl jasmonate (MeJA), the JAZ genes were correlated with COI1 under MeJA. Co-expression and GO enrichment analysis further revealed that JAZ genes were related to hormone. JAZ proteins involved in the ABA and SA pathways. Subcellular localization experiments confirmed that the JAZ proteins were localized in the nucleus. Yeast two-hybrid assay showed that the JAZ proteins may form homologous or heterodimers to regulate hormones. CONCLUSION: Our results provided novel insights into biological function of TIFY family and JAZ subfamily in birch. It provides the theoretical reference for in-depth analysis of plant hormone and molecular breeding design for resistance.

Seasonal Transcriptome Profiling of Susceptible and Tolerant Citrus Cultivars to Citrus Huanglongbing.

Citrus huanglongbing (HLB) caused by 'Candidatus Liberibacter asiaticus' (CLas) is the most devastating citrus disease worldwide. Most commercial citrus cultivars are susceptible to HLB, with a few more tolerant exceptions such as 'LB8-9' Sugar Belle mandarin. Transcriptomic analyses have been widely used to investigate the potential mechanisms for disease susceptibility, resistance, or tolerance. Previous transcriptomic studies related to HLB mostly focused on single time point data collection. We hypothesize that changes in day length and temperature throughout the seasons have profound effects on citrus-CLas interactions. Here, we conducted RNA-seq analyses on HLB-susceptible Valencia sweet orange and HLB-tolerant mandarin 'LB8-9' in winter, spring, summer, and fall. Significant variations in differentially expressed genes (DEGs) related to HLB were observed among the four seasons. For both cultivars, the highest number of DEGs were found in the spring. CLas infection stimulates the expression of immune-related genes such as NBS-LRR, RLK, RLCK, CDPK, MAPK pathway, reactive oxygen species (ROS), and PR genes in both cultivars, consistent with the model that HLB is a pathogen-triggered immune disease. HLB-positive mandarin 'LB8-9' trees contained higher concentrations of maltose and sucrose, which are known to scavenge ROS. In addition, mandarin 'LB8-9' showed higher expression of genes involved in phloem regeneration, which might contribute to its HLB tolerance. This study shed light on the pathogenicity mechanism of the HLB pathosystem and the tolerance mechanism against HLB, providing valuable insights into HLB management.

Role of PsnWRKY70 in Regulatory Network Response to Infection with Alternaria alternata (Fr.) Keissl in Populus.

WRKY is an important complex family of transcription factors involved in plant immune responses. Among them, WRKY70 plays an important role in the process of the plant defense response to the invasion of pathogens. However, the defense mechanism of PsnWRKY70 is not clear in Populus nigra. In this study, we showed that PsnWRKY70-overexpression lines (OE) had fewer leaf blight symptoms than PsnWRKY70-repressing lines (RE). PsnWRKY70 activated MAP kinase cascade genes (PsnM2K4, PsnMPK3, PsnM3K18), calcium channel proteins-related genes (PsnCNG3, PsnCNGC1, PsnCNG4), and calcium-dependent protein kinases genes (PsnCDPKL, PsnCDPKW, PsnCDPKS, PsnCDPKQ). Furthermore, 129 genes of PsnWRKY70 putative genome-wide direct targets (DTGs) were identified by using transcriptome (RNA-seq) and DNA affinity purification sequencing (DAP-seq). PsnWRKY70 directly binds to the promoters of homologous genes and LRR domain proteins to promote the expression of WRKY6, WRKY18, WRKY22, and WRKY22-1, LRR domain proteins LRR8, LRR-RLK, ADR1-like 2, NB-ARC, etc. Our study suggests that PsnWRKY70 enhances the resistance of A. alternata in poplar by activating genes in both pathogen-associated molecular pattern-triggered immunity (PTI) and effector-triggered immunity (ETI).

A chromosome-scale reference genome of trifoliate orange (Poncirus trifoliata) provides insights into disease resistance, cold tolerance and genome evolution in Citrus.

Trifoliate orange (Poncirus trifoliata), a deciduous close relative of evergreen Citrus, has important traits for citrus production, including tolerance/resistance to citrus greening disease (Huanglongbing, HLB) and other major diseases, and cold tolerance. It has been one of the most important rootstocks, and one of the most valuable sources of resistance and tolerance genes for citrus. Here we present a high-quality, chromosome-scale genome assembly of P. trifoliata. The 264.9-Mb assembly contains nine chromosomal pseudomolecules with 25 538 protein-coding genes, covering 97.2% of the estimated gene space. Comparative analyses of P. trifoliata and nine Citrus genomes revealed 605 species-specific genes and six rapidly evolving gene families in the P. trifoliata genome. Poncirus trifoliata has evolved specific adaptation in the C-repeat/DREB binding factor (CBF)-dependent and CBF-independent cold signaling pathways to tolerate cold. We identified candidate genes within quantitative trait loci for HLB tolerance, and at the loci for resistance to citrus tristeza virus and citrus nematode. Genetic diversity analysis of Poncirus accessions and Poncirus/Citrus hybrids shows a narrow genetic base in the US germplasm collection, and points to the importance of collecting and preserving more natural genetic variation. Two phenotypically divergent Poncirus accessions are found to be clonally related, supporting a previous conjecture that dwarf Flying Dragon originated as a mutant of a non-dwarfing type. The high-quality genome reveals features and evolutionary insights of Poncirus, and it will serve as a valuable resource for genetic, genomic and molecular research and manipulation in citrus.

Deficiency of valencene in mandarin hybrids is associated with a deletion in the promoter region of the valencene synthase gene.

BACKGROUND: Valencene is a major sesquiterpene in citrus oil and biosynthesized by valencene synthase (Cstps1; EC: 4.2.3.73) from the 15-carbon substrate farnesyl diphosphate. It is abundant in juice of some mandarins (e.g. Citrus reticulata Blanco cv. Fortune), however, it is undetectable in others (e.g. C. reticulata Blanco cv. Murcott), We have discovered that the Murcott mandarin Cstps1 gene expression is severely reduced. A previous genetic mapping study using an F1 population of Fortune × Murcott found that the segregation of valencene production in fruit exhibited a Mendelian inheritance ratio of 1:1. There was only one dominant locus associated with valencene content detected on the mandarin genetic map. The goal of this study was to understand the molecular mechanism underlying the valencene deficiency observed in some citrus hybrids. RESULTS: There was a clear relationship between presence or absence of the valencene synthase gene (Cstps1) expression, and presence or absence of valencene among randomly selected mandarin hybrids. Cloning the coding regions of Cstps1 from Fortune and Murcott mandarin, and aligning with previous reported Valencia orange Cstps1 sequence, showed that they both exhibited extremely high similarity with the known Cstps1. By further cloning and analyzing the promoter region of Cstps1 from Valencia, Fortune and Murcott, a 12-nucleotide deletion at approximately - 270 bp from the Cstps1 coding region was only found in Murcott. Three binary vectors, designated as p1380-FortP-GUSin, p1380-MurcP-GUSin and p1380-MurcP(+ 12)-GUSin, were developed for promoter activity analysis. Transient over-expression of Fortune Cstps1 promoter in sweet orange showed notable GUS activity, but the Murcott Cstps1 promoter did not. In addition, by re-inserting the 12-nucleotide fragment, the activity of the Murcott Cstps1 promoter was mostly recovered. CONCLUSION: The deficiency of valencene production in some mandarins is probably due to a 12-nucleotide deletion in the promoter region of the Cstps1, which could be a crucial switch of Cstps1 transcription. Our results further enhanced the understanding of valencene biosynthesis in citrus.

Characterization and T-DNA insertion sites identification of a multiple-branches mutant br in Betula platyphylla × Betula pendula.

BACKGROUND: Plant architecture, which is mostly determined by shoot branching, plays an important role in plant growth and development. Thus, it is essential to explore the regulatory molecular mechanism of branching patterns based on the economic and ecological importance. In our previous work, a multiple-branches birch mutant br was identified from 19 CINNAMOYL-COENZYME A REDUCTASE 1 (CCR1)-overexpressed transgenic lines, and the expression patterns of differentially expressed genes in br were analyzed. In this study, we further explored some other characteristics of br, including plant architecture, wood properties, photosynthetic characteristics, and IAA and Zeatin contents. Meanwhile, the T-DNA insertion sites caused by the insertion of exogenous BpCCR1 in br were identified to explain the causes of the mutation phenotypes. RESULTS: The mutant br exhibited slower growth, more abundant and weaker branches, and lower wood basic density and lignin content than BpCCR1 transgenic line (OE2) and wild type (WT). Compared to WT and OE2, br had high stomatal conductance (Gs), transpiration rate (Tr), but a low non-photochemical quenching coefficient (NPQ) and chlorophyll content. In addition, br displayed an equal IAA and Zeatin content ratio of main branches' apical buds to lateral branches' apical buds and high ratio of Zeatin to IAA content. Two T-DNA insertion sites caused by the insertion of exogenous BpCCR1 in br genome were found. On one site, chromosome 2 (Chr2), no known gene was detected on the flanking sequence. The other site was on Chr5, with an insertion of 388 bp T-DNA sequence, resulting in deletion of 107 bp 5' untranslated region (UTR) and 264 bp coding sequence (CDS) on CORONATINE INSENSITIVE 1 (BpCOII). In comparison with OE2 and WT, BpCOI1 was down-regulated in br, and the sensitivity of br to Methyl Jasmonate (MeJA) was abnormal. CONCLUSIONS: Plant architecture, wood properties, photosynthetic characteristics, and IAA and Zeatin contents in main and lateral branches' apical buds changed in br over the study's time period. One T-DNA insertion was identified on the first exon of BpCOI1, which resulted in the reduction of BpCOI1 expression and abnormal perception to MeJA in br. These mutation phenotypes might be associated with a partial loss of BpCOI1 in birch.

Functional study of CHS gene family members in citrus revealed a novel CHS gene affecting the production of flavonoids.

BACKGROUND: Citrus flavonoids are considered as the important secondary metabolites because of their biological and pharmacological activities. Chalcone synthase (CHS) is a key enzyme that catalyses the first committed step in the flavonoid biosynthetic pathway. CHS genes have been isolated and characterized in many plants. Previous studies indicated that CHS is a gene superfamily. In citrus, the number of CHS members and their contribution to the production of flavonoids remains a mystery. In our previous study, the copies of CitCHS2 gene were found in different citrus species and the sequences are highly conserved, but the flavonoid content varied significantly among those species. RESULTS: From seventy-seven CHS and CHS-like gene sequences, ten CHS members were selected as candidates according to the features of their sequences. Among these candidates, expression was detected from only three genes. A predicted CHS sequence was identified as a novel CHS gene. The structure analysis showed that the gene structure of this novel CHS is very similar to other CHS genes. All three CHS genes were highly conserved and had a basic structure that included one intron and two exons, although they had different expression patterns in different tissues and developmental stages. These genes also presented different sensitivities to methyl jasmonate (MeJA) treatment. In transgenic plants, the expression of CHS genes was significantly correlated with the production of flavonoids. The three CHS genes contributed differently to the production of flavonoids. CONCLUSION: Our study indicated that CitCHS is a gene superfamily including at least three functional members. The expression levels of the CHS genes are highly correlated to the biosynthesis of flavonoids. The CHS enzyme is dynamically produced from several CHS genes, and the production of total flavonoids is regulated by the overall expression of CHS family genes.

Identification of QTLs controlling aroma volatiles using a 'Fortune' x 'Murcott' (Citrus reticulata) population.

BACKGROUND: Flavor is an important attribute of mandarin (Citrus reticulata Blanco), but flavor improvement via conventional breeding is very challenging largely due to the complexity of the flavor components and traits. Many aroma associated volatiles of citrus fruit have been identified, which are directly related to flavor, but knowledge of genetic linkages and relevant genes for these volatiles, along with applicable markers potentially for expeditious and economical marker-assisted selection (MAS), is very limited. The objective of this project was to identify single nucleotide polymorphism (SNP) markers associated with these volatile traits. RESULT: Aroma volatiles were investigated in two mandarin parents ('Fortune' and 'Murcott') and their 116 F1 progeny using gas chromatography mass spectrometry in 2012 and 2013. A total of 148 volatiles were detected, including one acid, 12 alcohols, 20 aldehydes, 14 esters, one furan, three aromatic hydrocarbons, 16 ketones, one phenol, 27 sesquiterpenes, 15 monoterpenes, and 38 unknowns. A total of 206 quantitative trait loci (QTLs) were identified for 94 volatile compounds using genotyping data generated from a 1536-SNP Illumina GoldenGate assay. In detail, 25 of the QTLs were consistent over more than two harvest times. Forty-one QTLs were identified for 17 aroma active compounds that included 18 sesquiterpenes and were mapped onto four genomic regions. Fifty QTLs were for 14 monoterpenes and mapped onto five genomic regions. Candidate genes for some QTLs were also identified. A QTL interval for monoterpenes and sesquiterpenes on linkage group 2 contained four genes: geranyl diphosphate synthase 1, terpene synthase 3, terpene synthase 4, and terpene synthase 14. CONCLUSIONS: Some fruit aroma QTLs were identified and the candidate genes in the terpenoid biosynthetic pathway were found within the QTL intervals. These QTLs could lead to an efficient and feasible MAS approach to mandarin flavor improvement.

Proteomic and metabolomic analyses provide insight into production of volatile and non-volatile flavor components in mandarin hybrid fruit.

BACKGROUND: Although many of the volatile constituents of flavor and aroma in citrus have been identified, the knowledge of molecular mechanisms and regulation of volatile production are very limited. Our aim was to understand mechanisms of flavor volatile production and regulation in mandarin fruit. RESULT: Fruits of two mandarin hybrids, Temple and Murcott with contrasting volatile and non- volatile profiles, were collected at three developmental stages. A combination of methods, including the isobaric tags for relative and absolute quantification (iTRAQ), quantitative real-time polymerase chain reaction, gas chromatography, and high-performance liquid chromatography, was used to identify proteins, measure gene expression levels, volatiles, sugars, organic acids and carotenoids. Two thirds of differentially expressed proteins were identified in the pathways of glycolysis, citric acid cycle, amino acid, sugar and starch metabolism. An enzyme encoding valencene synthase gene (Cstps1) was more abundant in Temple than in Murcott. Valencene accounted for 9.4% of total volatile content in Temple, whereas no valencene was detected in Murcott fruit. Murcott expression of Cstps1 is severely reduced. CONCLUSION: We showed that the diversion of valencene and other sesquiterpenes into the terpenoid pathway together with high production of apocarotenoid volatiles might have resulted in the lower concentration of carotenoids in Temple fruit.

Correlation between genetic polymorphism of matrix metalloproteinase-9 in patients with coronary artery disease and cardiac remodeling.

OBJECTIVE: To explore the correlation between genetic polymorphism of matrix metalloproteinase-9 (MMP-9) in patients with coronary artery disease (CAD) and cardiac remodeling. METHODS: A total of 272 subjects who received coronary angiography in our hospital from July 2008 to September 2013 were selected, including 172 CAD patients (CAD group) and another 100 ones (control group). Both groups were subjected to MMP-9 and ultrasonic detections to determine vascular remodeling and atherosclerotic plaques. C1562G polymorphism of MMP-9 gene was detected, and correlation with vascular remodeling and atherosclerotic plaque was analyzed. RESULTS: Serum MMP-9 level of CAD group (330.87±50.39 ng/ml) was significantly higher than that of control group (134.87±34.02 ng/ml) (P<0.05). Compared with control group, CAD group had significantly higher intima-media thickness, and significantly lower systolic peak velocity, mean systolic velocity and end-diastolic velocity (P<0.05). Total area of stenotic blood vessels was 67.34±22.98 mm(2), while that of control blood vessels was 64.00±20.83 mm(2). G/G, G/C and C/C genotype frequencies of MMP-9 differed significantly in the two groups (P<0.05). G and C allele frequencies of CAD group (70.9% and 29.1%) were significantly different from those of control group (50.0% and 50.0%) (P<0.05). G/G, G/C and C/C genotypes were manifested as lipid-rich, fibrous and calcified or ulcerated plaques respectively. Total area of stenotic blood vessels of G/G genotype significantly exceeded those of G/C and C/C genotypes (P<0.05), whereas the latter two had no significant differences. CONCLUSION: CAD promoted 1562C-G transformation of MMP-9 gene into genetic polymorphism, thus facilitating arterial remodeling and increasing unstable atherosclerotic plaques.

Identification of genes associated with low furanocoumarin content in grapefruit.

Some furanocoumarins in grapefruit (Citrus paradisi) are associated with the so-called grapefruit juice effect. Previous phytochemical quantification and genetic analysis suggested that the synthesis of these furanocoumarins may be controlled by a single gene in the pathway. In this study, cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis of fruit tissues was performed to identify the candidate gene(s) likely associated with low furanocoumarin content in grapefruit. Fifteen tentative differentially expressed fragments were cloned through the cDNA-AFLP analysis of the grapefruit variety Foster and its spontaneous low-furanocoumarin mutant Low Acid Foster. Sequence analysis revealed a cDNA-AFLP fragment, Contig 6, was homologous to a substrate-proved psoralen synthase gene, CYP71A22, and was part of citrus unigenes Cit.3003 and Csi.1332, and predicted genes Ciclev10004717m in mandarin and orange1.1g041507m in sweet orange. The two predicted genes contained the highly conserved motifs at one of the substrate recognition sites of CYP71A22. Digital gene expression profile showed the unigenes were expressed only in fruit and seed. Quantitative real-time PCR also proved Contig 6 was down-regulated in Low Acid Foster. These results showed the differentially expressed Contig 6 was related to the reduced furanocoumarin levels in the mutant. The identified fragment, homologs, unigenes, and genes may facilitate further furanocoumarin genetic study and grapefruit variety improvement.

Diversification in the genetic architecture of gene expression and transcriptional networks in organ differentiation of Populus.

A fundamental goal of systems biology is to identify genetic elements that contribute to complex phenotypes and to understand how they interact in networks predictive of system response to genetic variation. Few studies in plants have developed such networks, and none have examined their conservation among functionally specialized organs. Here we used genetical genomics in an interspecific hybrid population of the model hardwood plant Populus to uncover transcriptional networks in xylem, leaves, and roots. Pleiotropic eQTL hotspots were detected and used to construct coexpression networks a posteriori, for which regulators were predicted based on cis-acting expression regulation. Networks were shown to be enriched for groups of genes that function in biologically coherent processes and for cis-acting promoter motifs with known roles in regulating common groups of genes. When contrasted among xylem, leaves, and roots, transcriptional networks were frequently conserved in composition, but almost invariably regulated by different loci. Similarly, the genetic architecture of gene expression regulation is highly diversified among plant organs, with less than one-third of genes with eQTL detected in two organs being regulated by the same locus. However, colocalization in eQTL position increases to 50% when they are detected in all three organs, suggesting conservation in the genetic regulation is a function of ubiquitous expression. Genes conserved in their genetic regulation among all organs are primarily cis regulated (approximately 92%), whereas genes with eQTL in only one organ are largely trans regulated. Trans-acting regulation may therefore be the primary driver of differentiation in function between plant organs.

Construction of a high density genetic linkage map to define the locus conferring seedlessness from Mukaku Kishu mandarin.

Mukaku Kishu ('MK'), a small sized mandarin, is an important source of seedlessness in citrus breeding. Identification and mapping the gene(s) governing 'MK' seedlessness will expedite seedless cultivar development. In this study, two 'MK'-derived mapping populations- LB8-9 Sugar Belle® ('SB') × 'MK' (N=97) and Daisy ('D') × 'MK' (N=68) were genotyped using an Axiom_Citrus56 Array encompassing 58,433 SNP probe sets, and population specific male and female parent linkage maps were constructed. The parental maps of each population were integrated to produce sub-composite maps, which were further merged to develop a consensus linkage map. All the parental maps (except 'MK_D') had nine major linkage groups, and contained 930 ('SB'), 810 ('MK_SB'), 776 ('D') and 707 ('MK_D') SNPs. The linkage maps displayed 96.9 ('MK_D') to 98.5% ('SB') chromosomal synteny with the reference Clementine genome. The consensus map was comprised of 2588 markers including a phenotypic seedless (Fs)-locus and spanned a genetic distance of 1406.84 cM, with an average marker distance of 0.54 cM, which is substantially lower than the reference Clementine map. For the phenotypic Fs-locus, the distribution of seedy and seedless progenies in both 'SB' × 'MK' (55:42, χ2 = 1.74) and 'D' × 'MK' populations (33:35, χ2 = 0.06) followed a test cross pattern. The Fs-locus mapped on chromosome 5 with SNP marker 'AX-160417325' at 7.4 cM in 'MK_SB' map and between two SNP markers 'AX-160536283' and 'AX-160906995' at a distance of 2.4 and 4.9 cM, respectively in 'MK_D' map. The SNPs 'AX-160417325' and 'AX-160536283' correctly predicted seedlessness of 25-91.9% progenies in this study. Based on the alignment of flanking SNP markers to the Clementine reference genome, the candidate gene for seedlessness hovered in a ~ 6.0 Mb region between 3.97 Mb (AX-160906995) to 10.00 Mb (AX-160536283). This region has 131 genes of which 13 genes (belonging to seven gene families) reportedly express in seed coat or developing embryo. The findings of the study will prove helpful in directing future research for fine mapping this region and eventually underpinning the exact causative gene governing seedlessness in 'MK'.

A chromosome-level phased genome enabling allele-level studies in sweet orange: a case study on citrus Huanglongbing tolerance.

Sweet orange originated from the introgressive hybridizations of pummelo and mandarin resulting in a highly heterozygous genome. How alleles from the two species cooperate in shaping sweet orange phenotypes under distinct circumstances is unknown. Here, we assembled a chromosome-level phased diploid Valencia sweet orange (DVS) genome with over 99.999% base accuracy and 99.2% gene annotation BUSCO completeness. DVS enables allele-level studies for sweet orange and other hybrids between pummelo and mandarin. We first configured an allele-aware transcriptomic profiling pipeline and applied it to 740 sweet orange transcriptomes. On average, 32.5% of genes have a significantly biased allelic expression in the transcriptomes. Different cultivars, transgenic lineages, tissues, development stages, and disease status all impacted allelic expressions and resulted in diversified allelic expression patterns in sweet orange, but particularly citrus Huanglongbing (HLB) shifted the allelic expression of hundreds of genes in leaves and calyx abscission zones. In addition, we detected allelic structural mutations in an HLB-tolerant mutant (T19) and a more sensitive mutant (T78) through long-read sequencing. The irradiation-induced structural mutations mostly involved double-strand breaks, while most spontaneous structural mutations were transposon insertions. In the mutants, most genes with significant allelic expression ratio alterations (≥1.5-fold) were directly affected by those structural mutations. In T19, alleles located at a translocated segment terminal were upregulated, including CsDnaJ, CsHSP17.4B, and CsCEBPZ. Their upregulation is inferred to keep phloem protein homeostasis under the stress from HLB and enable subsequent stress responses observed in T19. DVS will advance allelic level studies in citrus.

Functional study of BpCOI1 reveals its role in affecting disease resistance in birch.

Plants interact with biotic and abiotic environments. Some of these interactions are detrimental including herbivory consumption and infections by microbial pathogens. The COI1 (coronatine insensitive 1) protein is the master controller of JA-regulated plant responses and plays a regulatory role in the plant defense response. However, there is little information on COI1 function in birch (Betula platyphylla × Betula pendula). Herein, we studied the F-box protein BpCOI1 which is located in the nucleus. To validate the function of this protein, we developed transgenic birch plants with overexpression or repression of BpCOI1 gene. Growth traits, such as tree height, ground diameter, number of lateral branches, did not change significantly among transgenic lines. Alternaria alternata treatment experiments indicated that low expression of BpCOI1 reduced disease resistance in birch. Furthermore, our results showed that low expression of BpCOI1 significantly reduced the sensitivity of plants to exogenous MeJA. Co-expression analysis showed gene expression patterns with similar characteristics. These genes may be closely related in function, or members involved in the same signaling pathway or physiological process with BpCOI 1. The results of transcriptome sequencing and co-expression analysis showed that BpCOI1 affects plant defense against Alternaria alternata by regulating jasmonates. This study reveals the role of BpCOI1 in disease resistance and proposes the possibility of controlling diseases through molecular breeding in birch.

Role of long non-coding RNA in regulatory network response to Candidatus Liberibacter asiaticus in citrus.

Long non-coding RNAs (lncRNAs) serve as crucial regulators in plant response to various diseases, while none have been systematically identified and characterized in response to citrus Huanglongbing (HLB) caused by Candidatus Liberibacter asiaticus (CLas) bacteria. Here, we comprehensively investigated the transcriptional and regulatory dynamics of the lncRNAs in response to CLas. Samples were collected from leaf midribs of CLas- and mock-inoculated HLB-tolerant rough lemon (Citrus jambhiri) and HLB-sensitive sweet orange (C. sinensis) at week 0, 7, 17, and 34 following inoculation using CLas+ budwood of three biological replicates in the greenhouse. A total of 8,742 lncRNAs, including 2,529 novel lncRNAs, were identified from RNA-seq data with rRNA-removed from strand-specific libraries. Genomic variation analyses of conserved lncRNAs from 38 citrus accessions showed that 26 single nucleotide polymorphisms (SNPs) were significantly correlated with HLB. In addition, lncRNA-mRNA weighted gene co-expression network analysis (WGCNA) showed a significant module correlated with CLas-inoculation in rough lemon. Notably, the most significant LNC_28805 and multiple co-expressed genes related to plant defense in the module were targeted by miRNA5021, suggesting that LNC28805 might compete with endogenous miR5021 to maintain the homeostasis of immune gene expression levels. Candidate WRKY33 and SYP121 genes targeted by miRNA5021 were identified as two key hub genes interacting with bacteria pathogen response genes based on the prediction of protein-protein interaction (PPI) network. These two genes were also found within HLB-associated QTL in linkage group 6. Overall, our findings provide a reference for a better understanding of the role of lncRNAs involved in citrus HLB regulation.

One-Stage Detection without Segmentation for Multi-Type Coronary Lesions in Angiography Images Using Deep Learning.

It is rare to use the one-stage model without segmentation for the automatic detection of coronary lesions. This study sequentially enrolled 200 patients with significant stenoses and occlusions of the right coronary and categorized their angiography images into two angle views: The CRA (cranial) view of 98 patients with 2453 images and the LAO (left anterior oblique) view of 176 patients with 3338 images. Randomization was performed at the patient level to the training set and test set using a 7:3 ratio. YOLOv5 was adopted as the key model for direct detection. Four types of lesions were studied: Local Stenosis (LS), Diffuse Stenosis (DS), Bifurcation Stenosis (BS), and Chronic Total Occlusion (CTO). At the image level, the precision, recall, mAP@0.1, and mAP@0.5 predicted by the model were 0.64, 0.68, 0.66, and 0.49 in the CRA view and 0.68, 0.73, 0.70, and 0.56 in the LAO view, respectively. At the patient level, the precision, recall, and F1scores predicted by the model were 0.52, 0.91, and 0.65 in the CRA view and 0.50, 0.94, and 0.64 in the LAO view, respectively. YOLOv5 performed the best for lesions of CTO and LS at both the image level and the patient level. In conclusion, the one-stage model without segmentation as YOLOv5 is feasible to be used in automatic coronary lesion detection, with the most suitable types of lesions as LS and CTO.

Heritable epigenetic modification of BpPIN1 is associated with leaf shapes in Betula pendula.

The new variety Betula pendula 'Dalecarlica', selected from Betula pendula, shows high ornamental value owing to its lobed leaf shape. In this study, to identify the genetic components of leaf shape formation, we performed bulked segregant analysis and molecular marker-based fine mapping to identify the causal gene responsible for lobed leaves in B. pendula 'Dalecarlica'. The most significant variations associated with leaf shape were identified within the gene BpPIN1 encoding a member of the PIN-FORMED family, responsible for the auxin efflux carrier. We further confirmed the hypomethylation at the promoter region promoting the expression level of BpPIN1, which causes stronger and longer veins and lobed leaf shape in B. pendula 'Dalecarlica'. These results indicated that DNA methylation at the BpPIN1 promoter region is associated with leaf shapes in B. pendula. Our findings revealed an epigenetic mechanism of BpPIN1 in the regulation of leaf shape in Betula  Linn. (birch), which could help in the molecular breeding of ornamental traits.

Comparative transcriptional and anatomical analyses of tolerant rough lemon and susceptible sweet orange in response to 'Candidatus Liberibacter asiaticus' infection.

Although there are no known sources of genetic resistance, some Citrus spp. are reportedly tolerant to huanglongbing (HLB), presumably caused by 'Candidatus Liberibacter asiaticus'. Time-course transcriptional analysis of tolerant rough lemon (Citrus jambhiri) and susceptible sweet orange (C. sinensis) in response to 'Ca. L. asiaticus' infection showed more genes differentially expressed in HLB-affected rough lemon than sweet orange at early stages but substantially fewer at late time points, possibly a critical factor underlying differences in sensitivity to 'Ca. L. asiaticus'. Pathway analysis revealed that stress responses were distinctively modulated in rough lemon and sweet orange. Although microscopic changes (e.g., callose deposition in sieve elements and phloem cell collapse) were found in both infected species, remarkably, phloem transport activity in midribs of source leaves in rough lemon was much less affected by HLB than in sweet orange. The difference in phloem cell transport activities is also implicated in the differential sensitivity to HLB between the two species. The results potentially lead to identification of key genes and the genetic mechanism in rough lemon to restrain disease development and maintain (or recover) phloem transport activity. These potential candidate genes may be used for improving citrus tolerance (or even resistance) to HLB by genetic engineering.