miR171-targeted SCARECROW-LIKE genes CsSCL2 and CsSCL3 regulate somatic embryogenesis in citrus.

Somatic embryogenesis (SE) is a key regeneration pathway in various biotechnology approaches to crop improvement, especially for economically important perennial woody crops like citrus. However, maintenance of SE capability has long been a challenge and becomes a bottleneck in biotechnology-facilitated plant improvement. In the embryogenic callus (EC) of citrus, we identified 2 csi-miR171c-targeted SCARECROW-LIKE genes CsSCL2 and CsSCL3 (CsSCL2/3), which exert positive feedback regulation on csi-miR171c expression. Suppression of CsSCL2 expression by RNA interference (RNAi) enhanced SE in citrus callus. A thioredoxin superfamily protein CsClot was identified as an interactive protein of CsSCL2/3. Overexpression of CsClot disturbed reactive oxygen species (ROS) homeostasis in EC and enhanced SE. Chromatin immunoprecipitation sequencing (ChIP-Seq) and RNA-Seq identified 660 genes directly suppressed by CsSCL2 that were enriched in biological processes including development-related processes, auxin signaling pathway, and cell wall organization. CsSCL2/3 bound to the promoters of regeneration-related genes, such as WUSCHEL-RELATED HOMEOBOX 2 (CsWOX2), CsWOX13, and Lateral Organ Boundaries Domain 40 (LBD40), and repressed their expression. Overall, CsSCL2/3 modulate ROS homeostasis through the interactive protein CsClot and directly suppress the expression of regeneration-related genes, thus regulating SE in citrus. We uncovered a regulatory pathway of miR171c-targeted CsSCL2/3 in SE, which shed light on the mechanism of SE and regeneration capability maintenance in citrus.

Chromosome-level assembly of the Neolamarckia cadamba genome provides insights into the evolution of cadambine biosynthesis.

Neolamarckia cadamba (Roxb.), a close relative of Coffea canephora and Ophiorrhiza pumila, is an important traditional medicine in Southeast Asia. Three major glycosidic monoterpenoid indole alkaloids (MIAs), cadambine and its derivatives 3ß-isodihydrocadambine and 3ß-dihydrocadambine, accumulate in the bark and leaves, and exhibit antimalarial, antiproliferative, antioxidant, anticancer and anti-inflammatory activities. Here, we report a chromosome-scale N. cadamba genome, with 744.5 Mb assembled into 22 pseudochromosomes with contig N50 and scaffold N50 of 824.14 Kb and 29.20 Mb, respectively. Comparative genomic analysis of N. cadamba with Co. canephora revealed that N. cadamba underwent a relatively recent whole-genome duplication (WGD) event after diverging from Co. canephora, which contributed to the evolution of the MIA biosynthetic pathway. We determined the key intermediates of the cadambine biosynthetic pathway and further showed that NcSTR1 catalyzed the synthesis of strictosidine in N. cadamba. A new component, epoxystrictosidine (C27H34N2O10, m/z 547.2285), was identified in the cadambine biosynthetic pathway. Combining genome-wide association study (GWAS), population analysis, multi-omics analysis and metabolic gene cluster prediction, this study will shed light on the evolution of MIA biosynthetic pathway genes. This N. cadamba reference sequence will accelerate the understanding of the evolutionary history of specific metabolic pathways and facilitate the development of tools for enhancing bioactive productivity by metabolic engineering in microbes or by molecular breeding in plants.

Genome-Wide Identification and Expression Analysis of WRKY Gene Family in Neolamarckia cadamba.

The WRKY transcription factor family plays important regulatory roles in multiple biological processes in higher plants. They have been identified and functionally characterized in a number of plant species, but very little is known in Neolamarckia cadamba, a 'miracle tree' for its fast growth and potential medicinal resource in Southeast Asia. In this study, a total of 85 WRKY genes were identified in the genome of N. cadamba. They were divided into three groups according to their phylogenetic features, with the support of the characteristics of gene structures and conserved motifs of protein. The NcWRKY genes were unevenly distributed on 22 chromosomes, and there were two pairs of segmentally duplicated events. In addition, a number of putative cis-elements were identified in the promoter regions, of which hormone- and stress-related elements were shared in many NcWRKYs. The transcript levels of NcWRKY were analyzed using the RNA-seq data, revealing distinct expression patterns in various tissues and at different stages of vascular development. Furthermore, 16 and 12 NcWRKY genes were confirmed to respond to various hormone treatments and two different abiotic stress treatments, respectively. Moreover, the content of cadambine, the active metabolite used for the various pharmacological activities found in N. cadamba, significantly increased after Methyl jasmonate treatment. In addition, expression of NcWRKY64/74 was obviously upregulated, suggesting that they may have a potential function of regulating the biosynthesis of cadambine in response to MeJA. Taken together, this study provides clues into the regulatory roles of the WRKY gene family in N. cadamba.

miR156 regulates somatic embryogenesis by modulating starch accumulation in citrus.

Somatic embryogenesis (SE) is a major regeneration approach for in vitro cultured tissues of plants, including citrus. However, SE capability is difficult to maintain, and recalcitrance to SE has become a major obstacle to plant biotechnology. We previously reported that miR156-SPL modules regulate SE in citrus callus. However, the downstream regulatory pathway of the miR156-SPL module in SE remains unclear. In this study, we found that transcription factors CsAGL15 and CsFUS3 bind to the CsMIR156A promoter and activate its expression. Suppression of csi-miR156a function leads to up-regulation of four target genes, SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (CsSPL) genes, and reduction of SE efficiency. In the short tandem target mimic (STTM)-miR156a overexpression callus (MIM156), the number of amyloplasts and starch content were significantly reduced, and genes involved in starch synthesis and transport were down-regulated. csi-miR172d was down-regulated, whereas the target genes, CsTOE1.1 and CsTOE1.2, which inhibit the expression of starch biosynthesis genes, were up-regulated. In our working model, CsAGL15 and CsFUS3 activate csi-miR156a, which represses CsSPLs and further regulates csi-miR172d and CsTOEs, thus altering starch accumulation in callus cells and regulating SE in citrus. This study elucidates the pathway of miR156-SPLs and miR172-TOEs-mediated regulation of SE, and provides new insights into enhancing SE capability in citrus.

miR171 modulates induction of somatic embryogenesis in citrus callus.

KEY MESSAGE: Overexpression of miR171 restored SE competence in the recalcitrant citrus callus, and inhibition of miR171 function weakened SE competence in the strongly embryogenic citrus callus. Somatic embryogenesis (SE) is an important way of in vitro regeneration for plants. For perennial woody crops such as citrus, embryogenic callus is usually induced from unfertilized aborted ovules and widely used in biotechnology aided breeding. However, SE capacity always declines in callus during subculture, which makes regeneration difficult and hinders the application of biotechnology. We previously found that miR171 may be a regulator of SE in citrus, based on the abundant expression of csi-miR171c in the embryogenic callus and during SE of citrus. Here, we report that miR171 promotes SE and is required for SE in citrus. Overexpression of miR171 restored SE competence in the recalcitrant callus of 'Guoqing No.1' Satsuma mandarin (G1), whereas inhibition of miR171 function by Short Tandem Target Mimic (STTM) weakened SE competence in the strongly embryogenic callus of 'Valencia' sweet orange (V). The comparative transcriptomic analysis in miR171 overexpressed callus line (OE) and the wild type callus (WT) indicated that overexpression of miR171 decreased the expression level of its SCARECROW-LIKE (CsSCL) targets, and activated stress response related biological processes and metabolic processes that are required for cell differentiation. However, CsSCLs were up-regulated in the OE callus during SE induction process, which activated the cell division and developmental processes that are required for embryogenesis progress. Our results validate the function of miR171 in regulation of SE and reveal the biological responses provoked by miR171 in citrus that may promote SE.

Loss of Wnt16 Leads to Skeletal Deformities and Downregulation of Bone Developmental Pathway in Zebrafish.

Wingless-type MMTV integration site family, member 16 (wnt16), is a wnt ligand that participates in the regulation of vertebrate skeletal development. Studies have shown that wnt16 can regulate bone metabolism, but its molecular mechanism remains largely undefined. We obtained the wnt16-/- zebrafish model using the CRISPR-Cas9-mediated gene knockout screen with 11 bp deletion in wnt16, which led to the premature termination of amino acid translation and significantly reduced wnt16 expression, thus obtaining the wnt16-/- zebrafish model. The expression of wnt16 in bone-related parts was detected via in situ hybridization. The head, spine, and tail exhibited significant deformities, and the bone mineral density and trabecular bone decreased in wnt16-/- using light microscopy and micro-CT analysis. RNA sequencing was performed to explore the differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis found that the down-regulated DEGs are mainly concentrated in mTOR, FoxO, and VEGF pathways. Protein-protein interaction (PPI) network analysis was performed with the detected DEGs. Eight down-regulated DEGs including akt1, bnip4, ptena, vegfaa, twsg1b, prkab1a, prkab1b, and pla2g4f.2 were validated by qRT-PCR and the results were consistent with the RNA-seq data. Overall, our work provides key insights into the influence of wnt16 gene on skeletal development.

Maternal immune activation alters visual acuity and retinogeniculate axon pruning in offspring mice.

Individuals with autism spectrum disorder (ASD) have been found to have a variety of sensory processing deficits. Here we report that maternal immune activation, a known factor for ASD, alters visual acuity in the offspring mice. By intraperitoneally injecting polyinosinic-polycytidylic acid (polyI:C) to induce maternal immune activation during embryonic days 10 to 14, we found that polyI:C treatment impairs visual acuity in young adult offspring mice as examined by their optomotor responses. Concurrently, polyI:C treatment suppresses retinogeniculate axon elimination, resulting in a high fraction of weak optical fibers innervating the relay neurons in the visual thalamus. The results link in-utero maternal inflammation to defective optical fiber pruning and arrested developmental strengthening of single optic fibers which may underlie impaired visual acuity.

miR156-SPL modules regulate induction of somatic embryogenesis in citrus callus.

miR156 is a highly conserved plant miRNA and has been extensively studied because of its versatile roles in plant development. Here, we report a novel role of miR156 in regulating somatic embryogenesis (SE) in citrus, one of the most widely cultivated fruit crops in the world. SE is an important means of in vitro regeneration, but over the course of long-term sub-culturing there is always a decline in the SE potential of the preserved citrus embryogenic callus, and this represents a key obstacle for citrus biotechnology. In this study, the SE competence of citrus callus of wild kumquat (Fortunella hindsii) was significantly enhanced by either overexpression of csi-miR156a or by individual knock-down of the two target genes, CsSPL3 and CsSPL14, indicating that the effect of miR156-SPL modules was established during the initial phases of SE induction. Biological processes that might promote SE in response to miR156 overexpression were explored using RNA-seq, and mainly included hormone signaling pathways, stress responses, DNA methylation, and the cell cycle. CsAKIN10 was identified as interacting protein of CsSPL14. Our results provide insights into the regulatory pathway through which miR156-SPL modules enhance the SE potential of citrus callus, and provide a theoretical basis for improvement of plant SE competence.

Genome-wide organization and expression profiling of the R2R3-MYB transcription factor family in pineapple (Ananas comosus).

BACKGROUND: The MYB proteins comprise one of the largest families of plant transcription factors, which are involved in various plant physiological and biochemical processes. Pineapple (Ananas comosus) is one of three most important tropical fruits worldwide. The completion of pineapple genome sequencing provides a great opportunity to investigate the organization and evolutionary traits of pineapple MYB genes at the genome-wide level. RESULTS: In the present study, a total of 94 pineapple R2R3-MYB genes were identified and further phylogenetically classified into 26 subfamilies, as supported by the conserved gene structures and motif composition. Collinearity analysis indicated that the segmental duplication events played a crucial role in the expansion of pineapple MYB gene family. Further comparative phylogenetic analysis suggested that there have been functional divergences of MYB gene family during plant evolution. RNA-seq data from different tissues and developmental stages revealed distinct temporal and spatial expression profiles of the AcMYB genes. Further quantitative expression analysis showed the specific expression patterns of the selected putative stress-related AcMYB genes in response to distinct abiotic stress and hormonal treatments. The comprehensive expression analysis of the pineapple MYB genes, especially the tissue-preferential and stress-responsive genes, could provide valuable clues for further function characterization. CONCLUSIONS: In this work, we systematically identified AcMYB genes by analyzing the pineapple genome sequence using a set of bioinformatics approaches. Our findings provide a global insight into the organization, phylogeny and expression patterns of the pineapple R2R3-MYB genes, and hence contribute to the greater understanding of their biological roles in pineapple.

Genome-scale mRNA and small RNA transcriptomic insights into initiation of citrus apomixis.

Nucellar embryony (NE) is an adventitious form of apomixis common in citrus, wherein asexual embryos initiate directly from nucellar cells surrounding the embryo sac. NE enables the fixation of desirable agronomic traits and the production of clonal offspring of virus-free rootstock, but impedes progress in hybrid breeding. In spite of the great importance of NE in citrus breeding and commercial production, little is understood about the underlying molecular mechanisms. In this study, the stages of nucellar embryo initiation (NEI) were determined for two polyembryonic citrus cultivars via histological observation. To explore the genes and regulatory pathways involved in NEI, we performed mRNA-seq and sRNA-seq analyses of ovules immediately prior to and at stages during NEI in the two pairs of cultivars. A total of 305 differentially expressed genes (DEGs) were identified between the poly- and monoembryonic ovules. Gene ontology (GO) analysis revealed that several processes are significantly enriched based on DEGs. In particular, response to stress, and especially response to oxidative stress, was over-represented in polyembryonic ovules. Nearly 150 miRNAs, comprising ~90 conserved and ~60 novel miRNAs, were identified in the ovules of either cultivar pair. Only two differentially expressed miRNAs (DEMs) were identified, of which the novel miRN23-5p was repressed whereas the targets accumulated in the polyembryonic ovules. This integrated study on the transcriptional and post-transcriptional regulatory profiles between poly- and monoembryonic citrus ovules provides new insights into the mechanism of NE, which should contribute to revealing the regulatory mechanisms of plant apomixis.

Comparative metabolic and transcriptional analysis of a doubled diploid and its diploid citrus rootstock (C. junos cv. Ziyang xiangcheng) suggests its potential value for stress resistance improvement.

BACKGROUND: Polyploidy has often been considered to confer plants a better adaptation to environmental stresses. Tetraploid citrus rootstocks are expected to have stronger stress tolerance than diploid. Plenty of doubled diploid citrus plants were exploited from diploid species for citrus rootstock improvement. However, limited metabolic and molecular information related to tetraploidization is currently available at a systemic biological level. This study aimed to evaluate the occurrence and extent of metabolic and transcriptional changes induced by tetraploidization in Ziyang xiangcheng (Citrus junos Sieb. ex Tanaka), which is a special citrus germplasm native to China and widely used as an iron deficiency tolerant citrus rootstock. RESULTS: Doubled diploid Ziyang xiangcheng has typical morphological and anatomical features such as shorter plant height, larger and thicker leaves, bigger stomata and lower stomatal density, compared to its diploid parent. GC-MS (Gas chromatography coupled to mass spectrometry) analysis revealed that tetraploidization has an activation effect on the accumulation of primary metabolites in leaves; many stress-related metabolites such as sucrose, proline and γ-aminobutyric acid (GABA) was remarkably up-regulated in doubled diploid. However, LC-QTOF-MS (Liquid chromatography quadrupole time-of-flight mass spectrometry) analysis demonstrated that tetraploidization has an inhibition effect on the accumulation of secondary metabolites in leaves; all the 33 flavones were down-regulated while all the 6 flavanones were up-regulated in 4x. By RNA-seq analysis, only 212 genes (0.8% of detected genes) are found significantly differentially expressed between 2x and 4x leaves. Notably, those genes were highly related to stress-response functions, including responses to salt stress, water and abscisic acid. Interestingly, the transcriptional divergence could not explain the metabolic changes, probably due to post-transcriptional regulation. CONCLUSION: Taken together, tetraploidization induced considerable changes in leaf primary and secondary metabolite accumulation in Ziyang xiangcheng. However, the effect of tetraploidization on transcriptome is limited. Compared to diploid, higher expression level of stress related genes and higher content of stress related metabolites in doubled diploid could be beneficial for its stress tolerance.

Knowledge, attitudes, practices, and self-efficacy of the Chinese public regarding cardiopulmonary resuscitation: an online cross-sectional survey.

Objective: To evaluate the current status of Chinese public's knowledge, attitudes, practices (KAP) and self-efficacy regarding cardiopulmonary resuscitation (CPR), and to analyze the factors that influence KAP and self-efficacy. Methods: An online cross-sectional survey was conducted from February to June 2022 in Mainland China via a self-designed self-filled questionnaire. Potential participants were recruited through WeChat by convenience sampling and snowball sampling methods. Descriptive and quantitative analyses were used for statistical analysis. Results: The survey included 4,450 participants from 31 provinces, autonomous regions, or municipalities across Mainland China, aged 18 or above. The public's average understanding (clear and very clear) of the knowledge regarding CPR was 67.4% (3,000/4,450), with an average proportion of positive attitudes at 96.8% (4,308/4,450). In practice, the average proportion of good practices was 92.8% (4,130/4,450), while the percentage of good self-efficacy averaged at 58.9% (2,621/4,450), only 42.4% (1,885/4,450) of the participants had confidence in the correct use of automated external defibrillator (AED). Pearson correlation analysis showed a significantly positive correlation among knowledge, attitude, practice, and self-efficacy (p < 0.01). Multiple linear regression analysis revealed that several factors have a significant influence on the public's CPR KAP and self-efficacy, including ever having received CPR training (p < 0.001), hearing about AED (p < 0.001), performing CPR on others (p < 0.001), hearing about CPR (p < 0.001), occupation (p < 0.001), personal health status (p < 0.001), education level (p < 0.001), gender (p < 0.001), and encountering someone in need of CPR (p = 0.021). Conclusion: The Chinese public demonstrates good knowledge of CPR, positive attitude, and high willingness to perform CPR. However, there is still room for improvement in the mastery of some professional knowledge points related to CPR and AED. It should be noted that knowledge, attitude, practice, and self-efficacy are interrelated and influence each other. Factors such as prior CPR training, hearing about AED, having performed CPR before, hearing about CPR, occupation, personal health status, education level, gender, and having encountered someone in need of CPR have a significant impact on the public's KAP and self-efficacy.

Application of the whole-course care model (IWF/C Care) for postintensive care syndrome based on an early warning system in critically ill patients: a randomised controlled trial study protocol.

INTRODUCTION: Critically ill patients are at risk of developing postintensive care syndrome (PICS), which is manifested by physical, psychological and cognitive impairment. Currently, there are no programmes that combine early warning systems with interventions for PICS. We hypothesise that a comprehensive care model for PICS based on an early warning system would reduce medical costs and the incidence of PICS. METHODS AND ANALYSIS: The Intensive Care Unit (ICU) -Ward-Family/Community whole-course care (IWF/C Care) trial will be a unicentric, randomised, controlled trial. A total of 138 ICU patients from two ICUs at a university hospital in Guizhou province, China, will be enrolled in February 2023. The inclusion criteria are an age of 18 years or older, an ICU stay of more than 48 hours, provide informed consent and the ability to communicate normally. Patients will be followed for 12 months and randomised in a 1:1:1 ratio to three groups. INTERVENTIONS: Patients in intervention group 1 will be assessed by the PICS early warning system within 24 hours of ICU discharge, and precise interventions will be carried out according to the results; that is, high-risk patients will receive care based on the IWF/C Care model and low-risk patients will receive routine care. All patients in intervention group 2 will receive care based on the IWF/C Care model. The control group will receive routine care. The primary endpoints are the incidence of PICS and quality of life. The secondary endpoints include the incidence of adverse events: the unplanned readmission rate, cost-effectiveness, and the experiences and feelings of patients receiving care based on the IWF/C Care model. The incidence of PICS will be measured at ICU discharge, general ward discharge, the home/community stage and 1 month and 3, 6, 9, and 12 months after discharge. ETHICS AND DISSEMINATION: Ethics approval was obtained from Biomedical Research Ethics Committee of the Affiliated Hospital of Zunyi Medical University (approval number: KLL-2022-780). The results of this study will be distributed through peer-reviewed journals. TRIAL REGISTRATION NUMBER: ChiCTR2300068135.

Diet-induced inflammation in the anterior paraventricular thalamus induces compulsive sucrose-seeking.

Overconsumption of palatable food may initiate neuroadaptive responses in brain reward circuitry that may contribute to eating disorders. Here we report that high-fat diet (HFD) consumption impedes threat-cue-induced suppression of sucrose-seeking in mice. This compulsive sucrose-seeking was due to enhanced cue-triggered neuronal activity in the anterior paraventricular thalamus (aPVT) resulting from HFD-induced microglia activation. Thus, metabolic inflammation in the aPVT produces an adaptive response to threat cues, leading to compulsive food-seeking.

Selection and Validation of Reference Genes for mRNA Expression by Quantitative Real-Time PCR Analysis in Neolamarckia cadamba.

Neolamarckia cadamba is an economically-important fast-growing tree species in South China and Southeast Asia. As a prerequisite first step for future gene expression studies, we have identified and characterized a series of stable reference genes that can be used as controls for quantitative real time PCR (qRT-PCR) expression analysis in this study. The expression stability of 15 candidate reference genes in various tissues and mature leaves under different conditions was evaluated using four different algorithms, i.e., geNorm, NormFinder, BestKeeper and RefFinder. Our results showed that SAMDC was the most stable of the selected reference genes across the set of all samples, mature leaves at different photosynthetic cycles and under drought stress, whereas RPL10A had the most stable expression in various tissues. PGK and RPS25 were considered the most suitable reference for mature leaves at different developmental stages and under cold treatment, respectively. Additionally, the gene expression profiles of sucrose transporter 4 (NcSUT4), and 9-cis-epoxycarotenoid dioxygenase 3 (NcNCED3) were used to confirm the validity of candidate reference genes. Collectively, our study is the first report to validate the optimal reference genes for normalization under various conditions in N. cadamba and will benefit the future discovery of gene function in this species.

Overexpression of the CsFUS3 gene encoding a B3 transcription factor promotes somatic embryogenesis in Citrus.

In citrus, genetic improvement via biotechnology is challenging due to insufficient understanding of molecular barriers that prevent regeneration by somatic embryogenesis (SE). Our previous study indicated that LEC genes were involved in SE in citrus, but their regulatory roles remain to be elucidated. Here, we cloned one of the LEC genes, CsFUS3, and show that it is preferentially expressed during SE and in the embryogenic callus (EC) derived from citrus varieties with strong embryogenic competence. The overexpression of CsFUS3 in recalcitrant citrus callus restored embryogenic competence. Complementation of the loss-of-function Arabidopsis fus3 mutant with the CsFUS3 gene restored normal late embryogenesis, which is consistent with the CsFUS3 and AtFUS3 proteins contributing to the same regulatory network in Arabidopsis. Transcriptome profiling revealed that the expression of particular TFs that promote SE was up-regulated in the citrus overexpression (OE) line. The 104 differentially expressed genes associated with hormone biosynthesis, catabolism, and signaling are particularly noteworthy. The dynamic change in the ratio of ABA to GA during SE in wild-type callus mirrored the expression pattern of CsFUS3. In contrast, in the OE line, the ratio of ABA to GA was higher and the capacity for SE was greater when the OE line was separately treated with ABA and GA biosynthesis inhibitors. Taken together, our results demonstrate that the overexpression of CsFUS3 appears to establish a cellular environment favorable to SE, at least in part by promoting a high ABA to GA ratio and by regulating the expression of TFs that promote SE.

Characterization of a Citrus R2R3-MYB Transcription Factor that Regulates the Flavonol and Hydroxycinnamic Acid Biosynthesis.

Flavonols and hydroxycinnamic acids are important phenylpropanoid metabolites in plants. In this study, we isolated and characterized a citrus R2R3-MYB transcription factor CsMYBF1, encoding a protein belonging to the flavonol-specific MYB subgroup. Ectopic expression of CsMYBF1 in tomato led to an up-regulation of a series of genes involved in primary metabolism and the phenylpropanoid pathway, and induced a strong accumulation of hydroxycinnamic acid compounds but not the flavonols. The RNAi suppression of CsMYBF1 in citrus callus caused a down-regulation of many phenylpropanoid pathway genes and reduced the contents of hydroxycinnamic acids and flavonols. Transactivation assays indicated that CsMYBF1 activated several promoters of phenylpropanoid pathway genes in tomato and citrus. Interestingly, CsMYBF1 could activate the CHS gene promoter in citrus, but not in tomato. Further examinations revealed that the MYBPLANT cis-elements were essential for CsMYBF1 in activating phenylpropanoid pathway genes. In summary, our data indicated that CsMYBF1 possessed the function in controlling the flavonol and hydroxycinnamic acid biosynthesis, and the regulatory differences in the target metabolite accumulation between two species may be due to the differential activation of CHS promoters by CsMYBF1. Therefore, CsMYBF1 constitutes an important gene source for the engineering of specific phenylpropanoid components.