Overexpression of AtMYB2 Promotes Tolerance to Salt Stress and Accumulations of Tanshinones and Phenolic Acid in Salvia miltiorrhiza.

Salvia miltiorrhiza is a prized traditional Chinese medicinal plant species. Its red storage roots are primarily used for the treatment of cardiovascular and cerebrovascular diseases. In this study, a transcription factor gene AtMYB2 was cloned and introduced into Salvia miltiorrhiza for ectopic expression. Overexpression of AtMYB2 enhanced salt stress resistance in S. miltiorrhiza, leading to a more resilient phenotype in transgenic plants exposed to high-salinity conditions. Physiological experiments have revealed that overexpression of AtMYB2 can decrease the accumulation of reactive oxygen species (ROS) during salt stress, boost the activity of antioxidant enzymes, and mitigate oxidative damage to cell membranes. In addition, overexpression of AtMYB2 promotes the synthesis of tanshinones and phenolic acids by upregulating the expression of biosynthetic pathway genes, resulting in increased levels of these secondary metabolites. In summary, our findings demonstrate that AtMYB2 not only enhances plant tolerance to salt stress, but also increases the accumulation of secondary metabolites in S. miltiorrhiza. Our study lays a solid foundation for uncovering the molecular mechanisms governed by AtMYB2 and holds significant implications for the molecular breeding of high-quality S. miltiorrhiza varieties.

Simultaneous Promotion of Salt Tolerance and Phenolic Acid Biosynthesis in Salvia miltiorrhiza via Overexpression of Arabidopsis MYB12.

Transcription factors play crucial roles in regulating plant abiotic stress responses and physiological metabolic processes, which can be used for plant molecular breeding. In this study, an R2R3-MYB transcription factor gene, AtMYB12, was isolated from Arabidopsis thaliana and introduced into Salvia miltiorrhiza under the regulation of the CaMV35S promoter. The ectopic expression of AtMYB12 resulted in improved salt tolerance in S. miltiorrhiza; transgenic plants showed a more resistant phenotype under high-salinity conditions. Physiological experiments showed that transgenic plants exhibited higher chlorophyll contents, and decreased electrolyte leakage and O2- and H2O2 accumulation when subjected to salt stress. Moreover, the activity of reactive oxygen species (ROS)-scavenging enzymes was enhanced in S. miltiorrhiza via the overexpression of AtMYB12, and transgenic plants showed higher superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) activities compared with those of the wild type (WT) under salt stress, coupled with lower malondialdehyde (MDA) levels. In addition, the amount of salvianolic acid B was significantly elevated in all AtMYB12 transgenic hair roots and transgenic plants, and qRT-PCR analysis revealed that most genes in the phenolic acid biosynthetic pathway were up-regulated. In conclusion, these results demonstrated that AtMYB12 can significantly improve the resistance of plants to salt stress and promote the biosynthesis of phenolic acids by regulating genes involved in the biosynthetic pathway.

Optimal Frequency of Hepatitis C Virus (HCV) RNA Testing for Detection of Acute HCV Infection Among At-risk People With Human Immunodeficiency Virus: A Multicenter Study.

Using 3-stage pooled-plasma hepatitis C virus (HCV) RNA testing performed quarterly among at-risk people with human immunodeficiency virus (PWH), we found that if testing had been performed every 6 or 12 months, 58.6%-91.7% of PWH who recently acquired HCV would be delayed for diagnosis and might contribute to onward HCV transmission with longer durations.

Knocking Out the Transcription Factor OsNAC092 Promoted Rice Drought Tolerance.

Environmental drought stress threatens rice production. Previous studies have reported that related NAC (NAM, ATAF1/2, and CUC) transcription factors play an important role in drought stress. Herein, we identified and characterized OsNAC092, encoding an NAC transcription factor that is highly expressed and induced during drought tolerance. OsNAC092 knockout lines created using the clustered regularly interspaced palindromic repeats (CRISPR)-associated protein 9 (Cas9) system exhibited increased drought resistance in rice. RNA sequencing showed that the knockout of OsNAC092 caused a global expression change, and differential gene expression is chiefly associated with "response to light stimulus," "MAPK signaling pathway," "plant hormone signal transduction," "response to oxidative stress," "photosynthesis," and "water deprivation." In addition, the antioxidants and enzyme activities of the redox response were significantly increased. OsNAC092 mutant rice exhibited a higher ability to scavenge more ROS and maintained a high GSH/GSSG ratio and redox level under drought stress, which could protect cells from oxidant stress, revealing the importance of OsNAC092 in the rice's response to abiotic stress. Functional analysis of OsNAC092 will be useful to explore many rice resistance genes in molecular breeding to aid in the development of modern agriculture.

BolTLP1, a Thaumatin-like Protein Gene, Confers Tolerance to Salt and Drought Stresses in Broccoli (Brassica oleracea L. var. Italica).

Plant thaumatin-like proteins (TLPs) play pleiotropic roles in defending against biotic and abiotic stresses. However, the functions of TLPs in broccoli, which is one of the major vegetables among the B. oleracea varieties, remain largely unknown. In the present study, bolTLP1 was identified in broccoli, and displayed remarkably inducible expression patterns by abiotic stress. The ectopic overexpression of bolTLP1 conferred increased tolerance to high salt and drought conditions in Arabidopsis. Similarly, bolTLP1-overexpressing broccoli transgenic lines significantly improved tolerance to salt and drought stresses. These results demonstrated that bolTLP1 positively regulates drought and salt tolerance. Transcriptome data displayed that bolTLP1 may function by regulating phytohormone (ABA, ethylene and auxin)-mediated signaling pathways, hydrolase and oxidoreductase activity, sulfur compound synthesis, and the differential expression of histone variants. Further studies confirmed that RESPONSE TO DESICCATION 2 (RD2), RESPONSIVE TO DEHYDRATION 22 (RD22), VASCULAR PLANT ONE-ZINC FINGER 2 (VOZ2), SM-LIKE 1B (LSM1B) and MALATE DEHYDROGENASE (MDH) physically interacted with bolTLP1, which implied that bolTLP1 could directly interact with these proteins to confer abiotic stress tolerance in broccoli. These findings provide new insights into the function and regulation of bolTLP1, and suggest potential applications for bolTLP1 in breeding broccoli and other crops with increased tolerance to salt and drought stresses.

Obtaining World Coordinate Information of UAV in GNSS Denied Environments.

GNSS information is vulnerable to external interference and causes failure when unmanned aerial vehicles (UAVs) are in a fully autonomous flight in complex environments such as high-rise parks and dense forests. This paper presents a pan-tilt-based visual servoing (PBVS) method for obtaining world coordinate information. The system is equipped with an inertial measurement unit (IMU), an air pressure sensor, a magnetometer, and a pan-tilt-zoom (PTZ) camera. In this paper, we explain the physical model and the application method of the PBVS system, which can be briefly summarized as follows. We track the operation target with a UAV carrying a camera and output the information about the UAV's position and the angle between the PTZ and the anchor point. In this way, we can obtain the current absolute position information of the UAV with its absolute altitude collected by the height sensing unit and absolute geographic coordinate information and altitude information of the tracked target. We set up an actual UAV experimental environment. To meet the calculation requirements, some sensor data will be sent to the cloud through the network. Through the field tests, it can be concluded that the systematic deviation of the overall solution is less than the error of GNSS sensor equipment, and it can provide navigation coordinate information for the UAV in complex environments. Compared with traditional visual navigation systems, our scheme has the advantage of obtaining absolute, continuous, accurate, and efficient navigation information at a short distance (within 15 m from the target). This system can be used in scenarios that require autonomous cruise, such as self-powered inspections of UAVs, patrols in parks, etc.

Knockout of the OsNAC006 Transcription Factor Causes Drought and Heat Sensitivity in Rice.

Rice (Oryza sativa) responds to various abiotic stresses during growth. Plant-specific NAM, ATAF1/2, and CUC2 (NAC) transcription factors (TFs) play an important role in controlling numerous vital growth and developmental processes. To date, 170 NAC TFs have been reported in rice, but their roles remain largely unknown. Herein, we discovered that the TF OsNAC006 is constitutively expressed in rice, and regulated by H2O2, cold, heat, abscisic acid (ABA), indole-3-acetic acid (IAA), gibberellin (GA), NaCl, and polyethylene glycol (PEG) 6000 treatments. Furthermore, knockout of OsNAC006 using the CRISPR-Cas9 system resulted in drought and heat sensitivity. RNA sequencing (RNA-seq) transcriptome analysis revealed that OsNAC006 regulates the expression of genes mainly involved in response to stimuli, oxidoreductase activity, cofactor binding, and membrane-related pathways. Our findings elucidate the important role of OsNAC006 in drought responses, and provide valuable information for genetic manipulation to enhance stress tolerance in future plant breeding programs.

Enhancement of tanshinone production in Salvia miltiorrhiza hairy root cultures by metabolic engineering.

BACKGROUND: Tanshinones are diterpenoid compounds that are used to treat cardiovascular diseases. As current extraction methods for tanshinones are inefficient, there is a pressing need to improve the production of these bioactive compounds to meet increasing demand. RESULTS: Overexpression of SmMDS (2-c-methyl-d-erythritol 2,4-cyclodiphosphate synthase, a tanshinone biosynthesis gene) in transgenic Salvia miltiorrhiza hairy roots significantly increased the tanshinone yield compared to the control, and total tanshinone content in SmMDS-overexpressing lines increased after elicitor treatment. Total tanshinones increased to 2.5, 2.3, and 3.2 mg/g DW (dry weight) following treatment with Ag+, YE (yeast extract), and MJ (methyl jasmonate), respectively, compared with the non-induced transgenic line (1.7 mg/g DW). Also, qRT-PCR analysis showed that the expression levels of two pathway genes was positively correlated with increased accumulation of tanshinone. CONCLUSIONS: Our study provides an effective strategy for increasing the content of tanshinones and other natural compounds using a combination of genetic engineering and elicitor treatment.

Parallel Speciation of Wild Rice Associated with Habitat Shifts.

The occurrence of parallel speciation strongly implies the action of natural selection. However, it is unclear how general a phenomena parallel speciation is since it was only shown in a small number of animal species. In particular, the adaptive process and mechanisms underlying the process of parallel speciation remain elusive. Here, we used an integrative approach incorporating population genomics, common garden, and crossing experiments to investigate parallel speciation of the wild rice species Oryza nivara from O. rufipogon. We demonstrated that O. nivara originated multiple times from different O. rufipogon populations and revealed that different O. nivara populations have evolved similar phenotypes under divergent selection, a reflection of recurrent local adaptation of ancient O. rufipogon populations to dry habitats. Almost completed premating isolation was detected between O. nivara and O. rufipogon in the absence of any postmating barriers between and within these species. These results suggest that flowering time is a "magic" trait that contributes to both local adaptation and reproductive isolation in the origin of wild rice species. Our study thus demonstrates a convincing case of parallel ecological speciation as a consequence of adaptation to new environments.

Identification of Glutathione Peroxidase (GPX) Gene Family in Rhodiola crenulata and Gene Expression Analysis under Stress Conditions.

Glutathione peroxidases (GPXs) are important enzymes in the glutathione-ascorbate cycle for catalyzing the reduction of H2O2 or organic hydroperoxides to water. GPXs play an essential role in plant growth and development by participating in photosynthesis, respiration, and stress tolerance. Rhodiola crenulata is a popular traditional Chinese medicinal plant which displays an extreme energy of tolerance to harsh alpine climate. The GPXs gene family might provide R. crenulata for extensively tolerance to environment stimulus. In this study, five GPX genes were isolated from R. crenulata. The protein amino acid sequences were analyzed by bioinformation softwares with the results that RcGPXs gene sequences contained three conserve cysteine residues, and the subcellular location predication were in the chloroplast, endoplasmic reticulum, or cytoplasm. Five RcGPXs members presented spatial and temporal specific expression with higher levels in young and green organs. And the expression patterns of RcGPXs in response to stresses or plant hormones were investigated by quantitative real-time PCR. In addition, the putative interaction proteins of RcGPXs were obtained by yeast two-hybrid with the results that RcGPXs could physically interact with specific proteins of multiple pathways like transcription factor, calmodulin, thioredoxin, and abscisic acid signal pathway. These results showed the regulation mechanism of RcGPXs were complicated and they were necessary for R. crenulata to adapt to the treacherous weather in highland.

Transcriptome and DNA methylome reveal insights into yield heterosis in the curds of broccoli (Brassica oleracea L var. italic).

BACKGROUND: Curds are the main edible organs, which exhibit remarkable yield heterosis in F1 hybrid broccoli. However, the molecular basis underlying heterosis in broccoli remains elusive. RESULTS: In the present study, transcriptome profiles revealed that the hybridization made most genes show additive expression patterns in hybrid broccoli. The differentially expressed genes including the non-additively expressed genes detected in the hybrid broccoli and its parents were mainly involved in light, hormone and hydrogen peroxide-mediated signaling pathways, responses to stresses, and regulation of floral development, which suggested that these biological processes should play crucial roles in the yield heterosis of broccoli. Among them, light and hydrogen peroxide-mediated signaling pathways represent two novel classes of regulatory processes that could function in yield or biomass heterosis of plants. Totally, 53 candidate genes closely involved in curd yield heterosis were identified. Methylome data indicated that the DNA methylation ratio of the hybrids was higher than that of their parents. However, the DNA methylation levels of most sites also displayed additive expression patterns. These sites with differential methylation levels were predominant in the intergenic regions. In most cases, the changes of DNA methylation levels in gene regions did not significantly affect their expression levels. CONCLUSIONS: The differentially expressed genes, the regulatory processes and the possible roles of DNA methylation modification in the formation of curd yield heterotic trait were discovered. These findings provided comprehensive insights into the curd yield heterosis in broccoli, and were significant for breeding high-yield broccoli varieties.

Ectopic Overexpression of bol-miR171b Increases Chlorophyll Content and Results in Sterility in Broccoli ( Brassica oleracea L var. italica).

MiR171 plays pleiotropic roles in the growth and development of several plant species. However, the mechanism underlying the miR171-mediated regulation of organ development in broccoli remains unknown. In this study, bol-miR171b was characterized and found to be differentially expressed in various broccoli organs. The ectopic overexpression of bol-miR171b in Arabidopsis affected the leaf and silique development of transgenic lines. In particular, the chlorophyll content of leaves from overexpressed bol-miR171b transgenic Arabidopsis was higher than that of the vector controls. The fertility and seed yield of Arabidopsis with overexpressed bol-miR171b were markedly lower than those of the vector controls. Similarly, overexpressed bol-miR171b transgenic broccoli exhibited dark green leaves with high chlorophyll content, and nearly all of the flowers were sterile. These results demonstrated that overexpression of bol-miR171b could increase the chlorophyll content of transgenic plants. Degradome sequencing was conducted to identify the targets of bol-miR171b. Two members of the GRAS gene family, BolSCL6 and BolSCL27, were cleaved by bol-miR171b-3p in broccoli. In addition to the genes targeted by bol-miR171b-3p, adenylylsulfate reductase 3 ( APSR3), which played important roles in plant sulfate assimilation and reduction, was speculated to be cleaved by bol-miR171b-5p, suggesting that the star sequence of bol-miR171b may also have functions in broccoli. Comparative transcriptome analysis further revealed that the genes involved in chloroplast development and sulfate homeostasis should participate in the bol-miR171b -mediated regulatory network. Taken together, these findings provided new insights into the function and regulation of bol-miR171b in broccoli and indicated the potential of bol-miR171b as a small RNA molecule that increased leaf chlorophyll in plants by genetic engineering.

Transcriptome Analyses from Mutant Salvia miltiorrhiza Reveals Important Roles for SmGASA4 during Plant Development.

Salvia miltiorrhiza (S. miltiorrhiza) is an important Chinese herb that is derived from the perennial plant of Lamiaceae, which has been used to treat neurasthenic insomnia and cardiovascular disease. We produced a mutant S. miltiorrhiza (MT), from breeding experiments, that possessed a large taproot, reduced lateral roots, and defective flowering. We performed transcriptome profiling of wild type (WT) and MT S. miltiorrhiza using second-generation Illumina sequencing to identify differentially expressed genes (DEGs) that could account for these phenotypical differences. Of the DEGs identified, we investigated the role of SmGASA4, the expression of which was down-regulated in MT plants. SmGASA4 was introduced into Arobidopsis and S. militiorrhiza under the control of a CaMV35S promoter to verify its influence on abiotic stress and S. miltiorrhiza secondary metabolism biosynthesis. SmGASA4 was found to promote flower and root development in Arobidopsis. SmGASA4 was also found to be positively regulated by Gibberellin (GA) and significantly enhanced plant resistance to salt, drought, and paclobutrazol (PBZ) stress. SmGASA4 also led to the up-regulation of the genes involved in salvianolic acid biosynthesis, but inhibited the expression of the genes involved in tanshinone biosynthesis. Taken together, our results reveal SmGASA4 as a promising candidate gene to promote S. miltiorrhiza development.

Comparative Transcriptome Analyses Reveal Potential Mechanisms of Enhanced Drought Tolerance in Transgenic Salvia Miltiorrhiza Plants Expressing AtDREB1A from Arabidopsis.

In our previous study, drought-resistant transgenic plants of Salvia miltiorrhiza were produced via overexpression of the transcription factor AtDREB1A. To unravel the molecular mechanisms underpinning elevated drought tolerance in transgenic plants, in the present study we compared the global transcriptional profiles of wild-type (WT) and AtDREB1A-expressing transgenic plants using RNA-sequencing (RNA-seq). Using cluster analysis, we identified 3904 differentially expressed genes (DEGs). Compared with WT plants, 423 unigenes were up-regulated in pRD29A::AtDREB1A-31 before drought treatment, while 936 were down-regulated and 1580 and 1313 unigenes were up- and down-regulated after six days of drought. COG analysis revealed that the 'signal transduction mechanisms' category was highly enriched among these DEGs both before and after drought stress. Based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation, DEGs associated with "ribosome", "plant hormone signal transduction", photosynthesis", "plant-pathogen interaction", "glycolysis/gluconeogenesis" and "carbon fixation" are hypothesized to perform major functions in drought resistance in AtDREB1A-expressing transgenic plants. Furthermore, the number of DEGs associated with different transcription factors increased significantly after drought stress, especially the AP2/ERF, bZIP and MYB protein families. Taken together, this study substantially expands the transcriptomic information for S. miltiorrhiza and provides valuable clues for elucidating the mechanism of AtDREB1A-mediated drought tolerance in transgenic plants.

Overexpression of the Glutathione Peroxidase 5 (RcGPX5) Gene From Rhodiola crenulata Increases Drought Tolerance in Salvia miltiorrhiza.

Excessive cellular accumulation of reactive oxygen species (ROS) due to environmental stresses can critically disrupt plant development and negatively affect productivity. Plant glutathione peroxidases (GPXs) play an important role in ROS scavenging by catalyzing the reduction of H2O2 and other organic hydroperoxides to protect plant cells from oxidative stress damage. RcGPX5, a member of the GPX gene family, was isolated from a traditional medicinal plant Rhodiola crenulata and constitutively expressed in Salvia miltiorrhiza under control of the CaMV 35S promoter. Transgenic plants showed increased tolerance to oxidative stress caused by application of H2O2 and drought, and had reduced production of malondialdehyde (MDA) compared with the wild type. Under drought stress, seedlings of the transgenic lines wilted later than the wild type and recovered growth 1 day after re-watering. In addition, the reduced glutathione (GSH) and total glutathione (T-GSH) contents were higher in the transgenic lines, with increased enzyme activities including glutathione reductase (GR), ascorbate peroxidase (APX), and GPX. These changes prevent H2O2 and O2 - accumulation in cells of the transgenic lines compared with wild type. Overexpression of RcGPX5 alters the relative expression levels of multiple endogenous genes in S. miltiorrhiza, including transcription factor genes and genes in the ROS and ABA pathways. In particular, RcGPX5 expression increases the mass of S. miltiorrhiza roots while reducing the concentration of the active ingredients. These results show that heterologous expression of RcGPX5 in S. miltiorrhiza can affect the regulation of multiple biochemical pathways to confer tolerance to drought stress, and RcGPX5 might act as a competitor with secondary metabolites in the S. miltiorrhiza response to environmental stimuli.

Genome-Wide Identification of AP2/ERF Transcription Factors in Cauliflower and Expression Profiling of the ERF Family under Salt and Drought Stresses.

The AP2/ERF transcription factors (TFs) comprise one of the largest gene superfamilies in plants. These TFs perform vital roles in plant growth, development, and responses to biotic and abiotic stresses. In this study, 171 AP2/ERF TFs were identified in cauliflower (Brassica oleracea L. var. botrytis), one of the most important horticultural crops in Brassica. Among these TFs, 15, 9, and 1 TFs were classified into the AP2, RAV, and Soloist family, respectively. The other 146 TFs belong to ERF family, which were further divided into the ERF and DREB subfamilies. The ERF subfamily contained 91 TFs, while the DREB subfamily contained 55 TFs. Phylogenetic analysis results indicated that the AP2/ERF TFs can be classified into 13 groups, in which 25 conserved motifs were confirmed. Some motifs were group- or subgroup- specific, implying that they are significant to the functions of the AP2/ERF TFs of these clades. In addition, 35 AP2/ERF TFs from the 13 groups were selected randomly and then used for expression pattern analysis under salt and drought stresses. The majority of these AP2/ERF TFs exhibited positive responses to these stress conditions. In specific, Bra-botrytis-ERF054a, Bra-botrytis-ERF056, and Bra-botrytis-CRF2a demonstrated rapid responses. By contrast, six AP2/ERF TFs were showed to delay responses to both stresses. The AP2/ERF TFs exhibiting specific expression patterns under salt or drought stresses were also confirmed. Further functional analysis indicated that ectopic overexpression of Bra-botrytis-ERF056 could increase tolerance to both salt and drought treatments. These findings provide new insights into the AP2/ERF TFs present in cauliflower, and offer candidate AP2/ERF TFs for further studies on their roles in salt and drought stress tolerance.

Small RNA Sequencing Reveals Differential miRNA Expression in the Early Development of Broccoli (Brassica oleracea var. italica) Pollen.

Pollen development is an important and complex biological process in the sexual reproduction of flowering plants. Although the cytological characteristics of pollen development are well defined, the regulation of its early stages remains largely unknown. In the present study, miRNAs were explored in the early development of broccoli (Brassica oleracea var. italica) pollen. A total of 333 known miRNAs that originated from 235 miRNA families were detected. Fifty-five novel miRNA candidates were identified. Sixty of the 333 known miRNAs and 49 of the 55 predicted novel miRNAs exhibited significantly differential expression profiling in the three distinct developmental stages of broccoli pollen. Among these differentially expressed miRNAs, miRNAs that would be involved in the developmental phase transition from uninucleate microspores to binucleate pollen grains or from binucleate to trinucleate pollen grains were identified. miRNAs that showed significantly enriched expression in a specific early stage of broccoli pollen development were also observed. In addition, 552 targets for 127 known miRNAs and 69 targets for 40 predicted novel miRNAs were bioinformatically identified. Functional annotation and GO (Gene Ontology) analysis indicated that the putative miRNA targets showed significant enrichment in GO terms that were related to plant organ formation and morphogenesis. Some of enriched GO terms were detected for the targets directly involved in plant male reproduction development. These findings provided new insights into the functions of miRNA-mediated regulatory networks in broccoli pollen development.

Modulating AtDREB1C Expression Improves Drought Tolerance in Salvia miltiorrhiza.

Dehydration responsive element binding proteins are transcription factors of the plant-specific AP2 family, many of which contribute to abiotic stress responses in several plant species. We investigated the possibility of increasing drought tolerance in the traditional Chinese medicinal herb, Salvia miltiorrhiza, through modulating the transcriptional regulation of AtDREB1C in transgenic plants under the control of a constitutive (35S) or drought-inducible (RD29A) promoter. AtDREB1C transgenic S. miltiorrhiza plants showed increased survival under severe drought conditions compared to the non-transgenic wild-type (WT) control. However, transgenic plants with constitutive overexpression of AtDREB1C showed considerable dwarfing relative to WT. Physiological tests suggested that the higher chlorophyll content, photosynthetic capacity, and superoxide dismutase, peroxidase, and catalase activity in the transgenic plants enhanced plant drought stress resistance compared to WT. Transcriptome analysis of S. miltiorrhiza following drought stress identified a number of differentially expressed genes (DEGs) between the AtDREB1C transgenic lines and WT. These DEGs are involved in photosynthesis, plant hormone signal transduction, phenylpropanoid biosynthesis, ribosome, starch and sucrose metabolism, and other metabolic pathways. The modified pathways involved in plant hormone signaling are thought to be one of the main causes of the increased drought tolerance of AtDREB1C transgenic S. miltiorrhiza plants.

Curd development associated gene (CDAG1) in cauliflower (Brassica oleracea L. var. botrytis) could result in enlarged organ size and increased biomass.

The curd is a specialized organ and the most important product organ of cauliflower (Brassica oleracea L. var. botrytis). However, the mechanism underlying the regulation of curd formation and development remains largely unknown. In the present study, a novel homologous gene containing the Organ Size Related (OSR) domain, namely, CDAG1 (Curd Development Associated Gene 1) was identified in cauliflower. Quantitative analysis indicated that CDAG1 showed significantly higher transcript levels in young tissues. Functional analysis demonstrated that the ectopic overexpression of CDAG1 in Arabidopsis and cauliflower could significantly promote organ growth and result in larger organ size and increased biomass. Organ enlargement was predominantly due to increased cell number. In addition, 228 genes involved in the CDAG1-mediated regulatory network were discovered by transcriptome analysis. Among these genes, CDAG1 was confirmed to inhibit the transcriptional expression of the endogenous OSR genes, ARGOS and ARL, while a series of ethylene-responsive transcription factors (ERFs) were found to increased expression in 35S:CDAG1 transgenic Arabidopsis plants. This implies that CDAG1 may function in the ethylene-mediated signal pathway. These findings provide new insight into the function of OSR genes, and suggest potential applications of CDAG1 in breeding high-yielding crops.

Ectopic Expression of DREB Transcription Factor, AtDREB1A, Confers Tolerance to Drought in Transgenic Salvia miltiorrhiza.

Drought decreases crop productivity more than any other type of environmental stress. Transcription factors (TFs) play crucial roles in regulating plant abiotic stress responses. The Arabidopsis thaliana gene DREB1A/CBF3, encoding a stress-inducible TF, was introduced into Salvia miltiorrhiza Ectopic expression of AtDREB1A resulted in increased drought tolerance, and transgenic lines had higher relative water content and Chl content, and exhibited an increased photosynthetic rate when subjected to drought stress. AtDREB1A transgenic plants generally displayed lower malondialdehyde (MDA), but higher superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) activities under drought stress. In particular, plants with ectopic AtDREB1A expression under the control of the stress-induced RD29A promoter exhibited more tolerance to drought compared with p35S::AtDREB1A transgenic plants, without growth inhibition or phenotypic aberrations. Differential gene expression profiling of wild-type and pRD29A::AtDREB1A transgenic plants following drought stress revealed that the expression levels of various genes associated with the stress response, photosynthesis, signaling, carbohydrate metabolism and protein protection were substantially higher in transgenic plants. In addition, the amount of salvianolic acids and tanshinones was significantly elevated in AtDREB1A transgenic S. miltiorrhiza roots, and most of the genes in the related biosynthetic pathways were up-regulated. Together, these results demonstrated that inducing the expression of a TF can effectively regulate multiple genes in the stress response pathways and significantly improve the resistance of plants to abiotic stresses. Our results also suggest that genetic manipulation of a TF can improve production of valuable secondary metabolites by regulating genes in associated pathways.