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1.
J Photochem Photobiol B ; 197: 111550, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31330424

RESUMO

The plant species of the genus Epimedium L. are well-known traditional Chinese medicinal herbs with special therapeutic effects on human beings and animals in invigorating sexuality and strengthening muscles and bones. In large-scale cultivating Epimedium that is a typical shade plant species, they are arbitrarily covered with black colored shade nets. However, their optimal growth conditions, especially light, are still less understood. During the investigation of different light qualities on the growth of Epimedium pseudowushanense, it was found that, all the values of plant growth characteristics (except shoot number) and photosynthetic characteristics were lower under red, yellow, or blue light treatment than under white light treatment. However, yellow light treatment had beneficial effects on shoot number, dry biomass (per plant) as well as net photosynthesis rate (Pn) and maximal apparent quantum efficiency (AQY) in E. pseudowushanense when compared with red or blue light treatment. More importantly, we found that E. pseudowushanense accumulated higher levels of bioactive flavonoids under yellow light treatment than under white, red, or blue light treatment. Furthermore, both RNAseq and qPCR analyses revealed that yellow light could highly up-regulate the expression levels of flavonoid biosynthetic genes, in particular CHS1, F3H1, PT_5, and raGT_5 that possibly contributed to the enhanced accumulation of bioactive flavonoids in E. pseudowushanense. Taken together, our study revealed that yellow light is the optimal light for the growth of E. pseudowushanense. Our results provided key information on how to improve the cultivation condition and concurrently enhance the accumulation of bioactive flavonoids in E. pseudowushanense.


Assuntos
Epimedium/metabolismo , Flavonoides/metabolismo , Luz , Biomassa , Cromatografia Líquida de Alta Pressão , Medicamentos de Ervas Chinesas , Epimedium/crescimento & desenvolvimento , Epimedium/efeitos da radiação , Flavonoides/análise , Fotossíntese/efeitos da radiação , Folhas de Planta/metabolismo , Folhas de Planta/efeitos da radiação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Brotos de Planta/metabolismo , Brotos de Planta/efeitos da radiação , RNA de Plantas/genética , RNA de Plantas/metabolismo , Análise de Sequência de RNA , Transcriptoma/efeitos da radiação
2.
Zhongguo Zhong Yao Za Zhi ; 44(12): 2480-2485, 2019 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-31359714

RESUMO

Tanshinones are abietane-type norditerpenoid quinones that make up the main bioactive ingredients of traditional Chinese medicine Salvia miltiorrhiza. Cytochrome CYP450 plays an important role in the post-structural modification of tanshinone biosynthesis pathway. Long non-coding RNA( lncRNA) have been defined as transcripts longer than 200 nucleotides,which have been functionally characterized in regulating the growth and development,secondary metabolism and stress of medicinal plants. In this study,we perform a comprehensive identification of lncRNAs in response to tanshinone metabolism induced by yeast extract( YE) and Ag~+ S. miltiorrhiza hairy roots. Deep RNA sequencing was used to identify a set of different 8 942 lncRNAs,of which 6 755 were intergenic lncRNAs. We predicted a total of 1 115 814 lncRNA-coding gene pairs,including 122 lncRNA-coding gene as cis pairs. The correlation analysis between lncRNA and CYP450 related to tanshinone biosynthesis was carried out and a total of 16 249 lncRNA-CYP450 target gene pairs were identified. Further analysis with functional known CYP76 AH1,CYP76 AH3 and CYP76 AK1 involved in tanshinone biosynthesis,we also identified a set of 216 target genes. These candidate genes will be the important target in the downstream regulation mechanism analysis of the tanshinone biosynthesis pathway.


Assuntos
Sistema Enzimático do Citocromo P-450/genética , Diterpenos de Abietano/biossíntese , RNA Longo não Codificante/genética , Salvia miltiorrhiza/genética , Regulação da Expressão Gênica de Plantas , Raízes de Plantas , RNA de Plantas/genética
3.
Nat Commun ; 10(1): 2878, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253789

RESUMO

Brassica napus, an allotetraploid crop, is hypothesized to be a hybrid from unknown varieties of Brassica rapa and Brassica oleracea. Despite the economic importance of B. napus, much is unresolved regarding its phylogenomic relationships, genetic structure, and diversification. Here we conduct a comprehensive study among diverse accessions from 183 B. napus (including rapeseed, rutabaga, and Siberian kale), 112 B. rapa, and 62 B. oleracea and its wild relatives. Using RNA-seq of B. napus accessions, we define the genetic diversity and sub-genome variance of six genetic clusters. Nuclear and organellar phylogenies for B. napus and its progenitors reveal varying patterns of inheritance and post-formation introgression. We discern regions with signatures of selective sweeps and detect 8,187 differentially expressed genes with implications for B. napus diversification. This study highlights the complex origin and evolution of B. napus providing insights that can further facilitate B. napus breeding and germplasm preservation.


Assuntos
Brassica napus/genética , Brassica napus/metabolismo , Ploidias , Regulação da Expressão Gênica de Plantas , Genômica , Organelas , Filogenia , Folhas de Planta/crescimento & desenvolvimento , Tubérculos , Polimorfismo de Nucleotídeo Único , RNA de Plantas/genética , Análise de Sequência de RNA , Transcriptoma
4.
BMC Plant Biol ; 19(1): 274, 2019 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-31234787

RESUMO

BACKGROUND: miRNAs are major regulators of gene expression and have proven their role in understanding the genetic regulation of biosynthetic pathways. Stevioside and rebaudioside-A, the two most abundant and sweetest compounds found in leaf extract of Stevia rebaudiana, have been used for many years in treatment of diabetes. It has been found that the crude extract is more potent than the purified extract. Stevioside, being accumulated in higher concentration, imparts licorice like aftertaste. Thus, in order to make the sweetener more potent and palatable, there is a need to increase the intrinsic concentration of steviol glycosides and to alter the ratio of rebaudioside-A to stevioside. Doing so would significantly increase the quality of the sweeteners, and the potential to be used on a wider scale. To do so, in previous report, miRNAs associated with genes of steviol glycosides biosynthetic pathway were identified in S. rebaudiana. In continuation to that in this study, the two miRNAs (miR319g and miRStv_11) targeting key genes of steviol glycosides biosynthetic pathway were modulated and their impact was evaluated on steviol glycosides contents. RESULTS: The over-expression results showed that miRStv_11 induced, while miR319g had repressive action on its target genes. The knock-down constructs for miR319g and miRStv_11 were then prepared and it was demonstrated that the expression of anti-miR319g produced inhibitory effect on its target miRNA, resulting in enhanced expression of its target genes. On the other hand, anti-miRStv_11 resulted in down-regulation of miRStv_11 and its target gene. Further miRStv_11 and anti-miR319gwere co-expressed which resulted in significant increase in stevioside (24.5%) and rebaudioside-A (51%) contents. CONCLUSION: In conclusion, the role of miR319g and miRStv_11 was successfully validated in steviol gycosides biosynthetic pathway gene regulation and their effect on steviol gycosides contents. In this study, we found the positively correlated miRNA-mRNA interaction network in plants, where miRStv_11 enhanced the expression of KAH gene. miRNAs knock-down was also successfully achieved using antisense precursors. Overall, this study thus reveals more complex nature and fundamental importance of miRNAs in biosynthetic pathway related gene networks and hence, these miRNAs can be successfully employed to enhance the ratio of rebaudioside-A to stevioside, thus enhancing the sweetening indices of this plant and making it more palatable.


Assuntos
Diterpenos de Caurano/biossíntese , Glucosídeos/biossíntese , MicroRNAs/metabolismo , RNA de Plantas/metabolismo , Stevia/metabolismo , Diterpenos de Caurano/química , Diterpenos de Caurano/genética , Regulação da Expressão Gênica de Plantas , Técnicas de Inativação de Genes , Inativação Gênica , Glucosídeos/química , Glucosídeos/genética , MicroRNAs/genética , Folhas de Planta/química , Regiões Promotoras Genéticas , RNA de Plantas/genética , Stevia/genética , Edulcorantes/química
5.
BMC Plant Biol ; 19(1): 247, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31185902

RESUMO

BACKGROUND: MiRNAs (microRNA) are 18-24 nt endogenous noncoding RNAs that regulate gene expression at the post-transcriptional level, including tissue-specific, developmental timing and evolutionary conservation gene expression. RESULTS: This study used high-throughput sequencing technology for the first time in Larix olgensis, predicted 78 miRNAs, including 12,229,003 reads sRNA, screened differentially expressed miRNAs. Predicting target genes was helpful for understanding the miRNA regulation function and obtained 333 corresponding target genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional annotation were analysed, mostly including nucleic acid binding, plant hormone signal transduction, pantothenate and CoA biosynthesis, and cellulose synthase. This study will lay the foundation for clarifying the complex miRNA-mediated regulatory network for growth and development. In view of this, spatio-temporal expression of miR396, miR950, miR164, miR166 and miR160 were analysed in Larix olgensis during the growth stages of not lignified, beginning of lignification, and completely lignified in different tissues (root, stem, and leaf) by quantitative real-time PCR (qRT-PCR). There were differences in the expression of miRNAs in roots, stems and leaves in the same growth period. At 60 days, miR160, miR166 and miR396-2 exhibited the highest expression in leaves. At 120 days, most miRNAs in roots and stems decreased significantly. At 180 days, miRNAs were abundantly expressed in roots and stems. Meanwhile, analysis of the expression of miRNAs in leaves revealed that miR396-2 was reduced as time went on, whereas other miRNAs increased initially and then decreased. On the other hand, in the stems, miR166-1 was increase, whereas other miRNAs, especially miR160, miR164, miR396 and miR950-1, first decreased and then increased. Similarly, in the roots, miR950-2 first decreased and then increased, whereas other miRNAs exhibited a trend of continuous increase. CONCLUSIONS: The present investigation included rapid isolation and identification of miRNAs in Larix olgensis through construction of a sRNA library using Solexa and predicted 78 novel miRNAs, which showed differential expression levels in different tissues and stages. These results provided a theoretical basis for further revealing the genetic regulation mechanism of miRNA in the growth and development of conifers and the verification of function in target genes.


Assuntos
Regulação da Expressão Gênica de Plantas , Larix/genética , MicroRNAs/genética , RNA de Plantas/genética , Perfilação da Expressão Gênica , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Larix/metabolismo , MicroRNAs/metabolismo , RNA de Plantas/metabolismo
6.
BMC Plant Biol ; 19(1): 232, 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31159725

RESUMO

BACKGROUND: Compared with white-fleshed sweetpotato (WFSP), purple-fleshed sweetpotato (PFSP) is a desirable resource for functional food development because of the abundant anthocyanin accumulation in its tuberous roots. Some studies have shown that the expression regulation mediated by miRNA plays an important role in anthocyanin biosynthesis in plants. However, few miRNAs and their corresponding functions related to anthocyanin biosynthesis in tuberous roots of sweetpotato have been known. RESULTS: In this study, small RNA (sRNA) and degradome libraries from the tuberous roots of WFSP (Xushu-18) and PFSP (Xuzishu-3) were constructed, respectively. Totally, 191 known and 33 novel miRNAs were identified by sRNA sequencing, and 180 target genes cleaved by 115 known ib-miRNAs and 5 novel ib-miRNAs were identified by degradome sequencing. Of these, 121 miRNAs were differently expressed between Xushu-18 and Xuzishu-3. Integrated analysis of sRNA, degradome sequencing, GO, KEGG and qRT-PCR revealed that 26 differentially expressed miRNAs and 36 corresponding targets were potentially involved in the anthocyanin biosynthesis. Of which, an inverse correlation between the expression of ib-miR156 and its target ibSPL in WFSP and PFSP was revealed by both qRT-PCR and sRNA sequencing. Subsequently, ib-miR156 was over-expressed in Arabidopsis. Interestingly, the ib-miR156 over-expressing plants showed suppressed abundance of SPL and a purplish phenotype. Concomitantly, upregulated expression of four anthocyanin pathway genes was detected in transgenic Arabidopsis plants. Finally, a putative ib-miRNA-target model involved in anthocyanin biosynthesis in sweetpotato was proposed. CONCLUSIONS: The results represented a comprehensive expression profiling of miRNAs related to anthocyanin accumulation in sweetpotato and provided important clues for understanding the regulatory network of anthocyanin biosynthesis mediated by miRNA in tuberous crops.


Assuntos
Antocianinas/biossíntese , Regulação da Expressão Gênica de Plantas , Ipomoea batatas/genética , MicroRNAs/genética , RNA de Plantas/genética , Antocianinas/genética , Sequenciamento de Nucleotídeos em Larga Escala , Ipomoea batatas/metabolismo , MicroRNAs/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , RNA de Plantas/metabolismo , Análise de Sequência de RNA
7.
World J Microbiol Biotechnol ; 35(7): 97, 2019 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-31222457

RESUMO

Endophytic Streptomyces sp. SSD49 inhibited eight pathogens, including the human opportunistic pathogenic microorganisms, the plant pathogenic fungi and bacteria. The growth of soybeans, tomatoes, peppers and Populus tomentosa seedings inoculated with SSD49 are remarkably promoted. Here, we constructed two P. tomentosa seedling microRNA (miRNA) libraries inoculated with (PS30d) and without SSD49 (PC30d) to explore the molecular regulatory roles in the plant response to the beneficial bacteria. Totals of 314 known and 144 novel miRNAs were identified, among which 27 known and 11 novel miRNA had significantly different expression. The targets of up-regulated miR160, miR156, ptc114 and down-regulated miR319 and other differential expressed miRNAs primarily regulated genes encoding transcription factors (auxin response factor, small auxin-up RNA, and GRAS proteins), disease resistance proteins, phytohormone oxidase, and response regulators, which could promote plant growth, influence disease resistance and miRNA biosynthesis in P. tomentosa. This is the first report on the genome-wide identification of biocontrol endophytic Streptomyces inoculation-responsive miRNAs using small RNA sequencing in P. tomentosa and these findings provide new insight into understanding the biocontrol effects of endophytic Streptomyces.


Assuntos
MicroRNAs/genética , Reguladores de Crescimento de Planta , Populus/genética , RNA de Plantas/isolamento & purificação , Streptomyces/metabolismo , Agentes de Controle Biológico , Regulação da Expressão Gênica de Plantas , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , MicroRNAs/metabolismo , Conformação de Ácido Nucleico , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Populus/metabolismo , Populus/microbiologia , RNA de Plantas/genética , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/isolamento & purificação , Análise de Sequência de RNA
8.
Plant Biol (Stuttg) ; 21(5): 796-804, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31081576

RESUMO

Thellungiella salsuginea is highly tolerant to abiotic stress, while its a close relative Arabidopsis thaliana is sensitive to stress. This characteristic makes T. salsuginea an excellent model for uncovering the mechanisms of abiotic stress tolerance. Abscisic acid (ABA) plays essential roles in plant abiotic and biotic stress tolerance. To test the changes in gene expression of T. salsuginea under ABA treatment, in this study, the transcriptomes of T. salsuginea roots and leaves were compared in response to exogenously application of ABA. The results showed that ABA treatment caused different expression of 2,200 and 3,305 genes in leaves and roots, respectively, compared with the untreated control. In particular, genes encoding transcription factors such as WRKY, MYB, NAC, GATA, ethylene-responsive factors (ERFs), heat stress transcription factors, basic helix-loop-helix, PLATZ and B3 domain-containing family members were enriched. In addition, 49 and 114 differentially expressed genes were identified as ABA-regulated genes, separately in leaves and roots, respectively, which were related to biotic and abiotic stresses. The expression levels of some genes were validated by qRT-PCR. Different responses of genes to ABA treatment were discovered in T. salsuginea and A. thaliana. This transcriptome analysis expands our understanding of the role of ABA in stress tolerance in T. salsuginea. Our study provides a wealth of information for improving stress tolerance in crop plants.


Assuntos
Brassicaceae/fisiologia , Ácido Abscísico/farmacologia , Brassicaceae/genética , Brassicaceae/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genoma de Planta/genética , Sequenciamento de Nucleotídeos em Larga Escala , Reguladores de Crescimento de Planta/farmacologia , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , RNA de Plantas/genética , Reação em Cadeia da Polimerase em Tempo Real , Estresse Fisiológico , Fatores de Transcrição/metabolismo
9.
Nat Plants ; 5(5): 539-550, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31076735

RESUMO

Post-transcriptional gene silencing (PTGS) is a major mechanism regulating gene expression in higher eukaryotes. To identify novel players in PTGS, a forward genetics screen was performed on an Arabidopsis thaliana line overexpressing a strong growth-repressive gene, ETHYLENE RESPONSE FACTOR6 (ERF6). We identified six independent ethyl-methanesulfonate mutants rescuing the dwarfism of ERF6-overexpressing plants as a result of transgene silencing. Among the causative genes, ETHYLENE-INSENSITIVE5, SUPERKILLER2 and HASTY1 have previously been reported to inhibit PTGS. Notably, the three other causative genes have not, to date, been related to PTGS: UTP:RNA-URIDYLYLTRANSFERASE1 (URT1), C-TERMINAL DOMAIN PHOSPHATASE-LIKE3 (CPL3) and RESURRECTION1 (RST1). We show that these genes may participate in protecting the 3' end of transgene transcripts. We present a model in which URT1, CPL3 and RST1 are classified as PTGS suppressors, as compromisation of these genes provokes the accumulation of aberrant transcripts which, in turn, trigger the production of small interfering RNAs, initiating RNA silencing.


Assuntos
Proteínas de Arabidopsis/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Proteínas de Membrana/fisiologia , Fosfoproteínas Fosfatases/fisiologia , Interferência de RNA , RNA Nucleotidiltransferases/fisiologia , RNA de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação/genética , Fosfoproteínas Fosfatases/genética , Fosfoproteínas Fosfatases/metabolismo , RNA Nucleotidiltransferases/genética , RNA Nucleotidiltransferases/metabolismo , RNA de Plantas/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transgenes/genética
10.
BMC Plant Biol ; 19(1): 194, 2019 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-31077147

RESUMO

BACKGROUND: Our study is the first to provide RNA-Seq data analysis related to transcriptomic responses towards drought across different crops. The aim was to identify and map which genes play a key role in drought response on leaves across different crops. Forty-two RNA-seq samples were analyzed from 9 published studies in 7 plant species (Arabidopsis thaliana, Solanum lycopersicum, Zea mays, Vitis vinifera, Malus X domestica, Solanum tuberosum, Triticum aestivum). RESULTS: Twenty-seven (16 up-regulated and 11 down-regulated) drought-regulated genes were commonly present in at least 7 of 9 studies, while 351 (147 up-regulated and 204 down-regulated) were commonly drought-regulated in 6 of 9 studies. Across all kind of leaves, the drought repressed gene-ontologies were related to the cell wall and membrane re-structuring such as wax biosynthesis, cell wall organization, fatty acid biosynthesis. On the other hand, drought-up-regulated biological processes were related to responses to osmotic stress, abscisic acid, water deprivation, abscisic-activated signalling pathway, salt stress, hydrogen peroxide treatment. A common metabolic feature linked to drought response in leaves is the repression of terpenoid pathways. There was an induction of AL1 (alfin-like), UGKYAH (trihelix), WRKY20, homeobox genes and members of the SET domain family in 6 of 9 studies. Several genes involved in detoxifying and antioxidant reactions, signalling pathways and cell protection were commonly modulated by drought across the 7 species. The chromosome (Chr) mapping of these key abiotic stress genes highlighted that Chr 4 in Arabidopsis thaliana, Chr 1 in Zea mays, Chr 2 and Chr 5 in Triticum aestivum contained a higher presence of drought-related genes compared to the other remaining chromosomes. In seedling studies, it is worth notice the up-regulation of ERF4 and ESE3 (ethylene), HVA22 (abscisic acid), TIR1 (auxin) and some transcription factors (MYB3, MYB94, MYB1, WRKY53 and WRKY20). In mature leaves, ERF1 and Alfin-like 1 were induced by drought while other transcription factors (YABBY5, ARR2, TRFL2) and genes involved phospholipid biosynthesis were repressed. CONCLUSIONS: The identified and mapped genes might be potential targets of molecular breeding activities to develop cultivars with enhanced drought resistance and tolerance across different crops.


Assuntos
Cromossomos de Plantas/genética , Produtos Agrícolas/genética , Genes de Plantas/genética , Folhas de Planta/metabolismo , RNA de Plantas/genética , Arabidopsis/genética , Arabidopsis/fisiologia , Mapeamento Cromossômico , Cromossomos de Plantas/fisiologia , Desidratação , Genes de Plantas/fisiologia , Lycopersicon esculentum/genética , Lycopersicon esculentum/fisiologia , Malus/genética , Malus/fisiologia , Folhas de Planta/anatomia & histologia , RNA de Plantas/fisiologia , Solanum tuberosum/genética , Solanum tuberosum/fisiologia , Triticum/genética , Triticum/fisiologia , Vitis/genética , Vitis/fisiologia , Zea mays/genética , Zea mays/fisiologia
11.
BMC Plant Biol ; 19(1): 214, 2019 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-31122194

RESUMO

BACKGROUND: MicroRNA319 (miR319) acts as an essential regulator of gene expression during plant development and under stress conditions. Although the role of miR319a in regulating leaf development has been well studied in tomato (Solanum lycopersicum), the function of the recently discovered wild tomato Solanum habrochaites miRNA319d (sha-miR319d) remains poorly understood. In this study, we overexpressed sha-miR319d in cultivated tomato 'Micro-Tom' to further investigate its role in tomato temperature stress responses. RESULTS: Under chilling or heat stress, sha-miR319d-overexpressing plants showed enhanced stress tolerance, including lower relative electrolyte leakage (REL), malondialdehyde (MDA) concentration, O2- generation and H2O2 concentration and higher chlorophyll contents and Fv/Fm values than wild-type (WT) plants. Overexpression of sha-miR319d enhanced the activities of superoxide dismutase (SOD) and catalase (CAT), with possible correlation with elevated expression levels of the genes FeSOD, CuZnSOD and CAT. Moreover, different expression levels of key genes involved in chilling (MYB83 and CBF1), heat (HsfA1a, HsfA1b and Hsp90), and reactive oxygen species (ROS) (ZAT12 and ZAT10) signaling in transgenic plants and WT were determined, suggesting a role for sha-miR319d in regulating tomato temperature stress via chilling, heat and ROS signaling. Silencing GAMYB-like1 increased tomato chilling tolerance as well as the expression levels of CBF1, CuZnSOD, CAT, APX1, APX2, ZAT12 and ZAT10. Additionally, overexpression of sha-miR319d in tomato caused plant leaf crinkling and reduced height. CONCLUSIONS: Overexpression of sha-miR319d confers chilling and heat stress tolerance in tomato. Sha-miR319d regulates tomato chilling tolerance, possibly by inhibiting expression of GAMYB-like1 and further alters chilling, heat and ROS signal transduction. Our research provides insight for further study of the role of sha-miR319d in tomato growth and stress regulation and lays a foundation for the genetic improvement of tomato.


Assuntos
Resposta ao Choque Frio/genética , Regulação da Expressão Gênica de Plantas , Resposta ao Choque Térmico/genética , Lycopersicon esculentum/fisiologia , MicroRNAs/genética , RNA de Plantas/genética , Solanum/fisiologia , Lycopersicon esculentum/genética , MicroRNAs/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/fisiologia , RNA de Plantas/metabolismo , Solanum/genética , Termotolerância/genética
12.
Gene ; 703: 145-152, 2019 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-30940526

RESUMO

We developed a novel assay system to quantitatively detect amber codon suppression by tRNAs expressed in plant cells. The assay was based on recovery of the expression of the green fluorescent protein (GFP) as a reporter, in which a fourth Lys codon (AAG) was changed to a premature amber codon TAG, designated as GFP/amber. Plasmids carrying GFP/amber, suppressor tRNA, and red fluorescent protein (RFF) as an internal control, respectively, were introduced into onion epidermal cells to monitor cell numbers with GFP and RFP fluorescence. First, an amber suppressor tRNASer from tobacco (NtS2) to suppress a TAG codon in GFP mRNA was examined, leading to the recovery of GFP fluorescence. Second, we used two different tRNAs (i.e., AtY3II-am and AtY3II-amiG7), both of which are intron-containing amber suppressor tRNAsTyr, the former impaired precursor-tRNA splicing but the latter did not, as confirmed previously using two different approaches (Szeykowska-Kulinska and Beier, 1991; Akama and Beier, 2003). As expected, coexpression of GFP/amber with AtY3II-am gave no green fluorescence, but significant fluorescence was observed with AtY3II-amiG7. Then, we applied this system for the analysis of 5'-regulatory sequences of the tRNAGln gene family from Arabidopsis. A 5'-flanking sequence of each of the 17 tRNAGln genes was fused to a coding region of an amber suppressor tRNASer gene (NtS2/amber) and its 3'-flanking sequence. Chimeric tRNASer gene, GFP/amber, and RFP were coexpressed, and the GFP or RFP fluorescence intensity was determined in cells using laser-scanning microscopy. In parallel, 17 kinds of original Arabidopsis tRNAGln genes and their chimeric genes with NtS2/amber were all analyzed in cell-free nuclear extract (Yukawa et al., 1997). Comparison of in vitro and in vivo expression of these chimeric tRNA genes displayed generally similar results, accompanied by a wide range of variance in the expression of each gene. Nevertheless, the expression patterns of several genes were clearly the opposite of each other comparing between the two different system, demonstrating the importance of in vivo systems in the study on tRNA expression in plants.


Assuntos
Proteínas de Fluorescência Verde/genética , Plantas/genética , RNA de Transferência/genética , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Expressão Gênica , Regulação da Expressão Gênica de Plantas , Proteínas de Fluorescência Verde/metabolismo , Cebolas/genética , Cebolas/crescimento & desenvolvimento , RNA de Plantas/genética
13.
BMC Plant Biol ; 19(1): 164, 2019 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-31029105

RESUMO

BACKGROUND: Circular RNAs (circRNAs) are 3'-5' head-to-tail covalently closed non-coding RNA that have been proved to play essential roles in many cellular and developmental processes. However, no information relate to cucumber circRNAs is available currently, especially under salt stress condition. RESULTS: In this study, we sequenced circRNAs in cucumber and a total of 2787 were identified, with 1934 in root and 44 in leaf being differentially regulated under salt stress. Characteristics analysis of these circRNAs revealed following features: most of them are exon circRNAs (79.51%) and they prefer to arise from middle exon(s) of parent genes (2035/2516); moreover, most of circularization events (88.3%) use non-canonical-GT/AG splicing signals; last but not least, pairing-driven circularization is not the major way to generate cucumber circRNAs since very few circRNAs (18) contain sufficient flanking complementary sequences. Annotation and enrichment analysis of both parental genes and target mRNAs were launched to uncover the functions of differentially expressed circRNAs induced by salt stress. The results showed that circRNAs may be paly roles in salt stress response by mediating transcription, signal transcription, cell cycle, metabolism adaptation, and ion homeostasis related pathways. Moreover, circRNAs may function to regulate proline metabolisms through regulating associated biosynthesis and degradation genes. CONCLUSIONS: The present study identified large number of cucumber circRNAs and function annotation revealed their possible biological roles in response to salt stress. Our findings will lay a solid foundation for further structure and function studies of cucumber circRNAs.


Assuntos
Cucumis sativus/genética , Cucumis sativus/fisiologia , RNA de Plantas/genética , RNA/genética , Estresse Salino/genética , Sequência de Bases , Biomassa , Cucumis sativus/crescimento & desenvolvimento , Éxons/genética , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Redes Reguladoras de Genes , Genes de Plantas , Transporte de Íons , MicroRNAs/genética , MicroRNAs/metabolismo , Anotação de Sequência Molecular , Raízes de Plantas/genética , Raízes de Plantas/fisiologia , RNA/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo
14.
Planta ; 249(6): 2015-2020, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30976910

RESUMO

MAIN CONCLUSION: Isolation of high-quality RNA, including miRNA, from microscopic woody apple bud meristem using laser capture microdissection-based method. It is often challenging to study the expression of microRNAs (miRNAs) or genes in less accessible inner tissues of tree species rich in polyphenols or polysaccharides. Here, we report a laser capture microdissection (LCM)-based method for efficient and cost-effective isolation and expression analysis of miRNAs and genes in the meristem tissue of woody apple bud. The tissue fixation, processing, infiltration, and sectioning steps were optimized for LCM-based excision and subsequent RNA isolation. Further, we have confirmed that RNA isolated from LCM-derived apple bud meristem contained miRNAs and was of good quantity and quality, sufficient for downstream expression analysis.


Assuntos
Microdissecção e Captura a Laser , Malus/genética , MicroRNAs/genética , Perfilação da Expressão Gênica , Malus/ultraestrutura , Meristema/genética , Meristema/ultraestrutura , RNA de Plantas/genética , Fixação de Tecidos , Madeira
15.
Food Chem ; 289: 278-284, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-30955613

RESUMO

Fusarium rot of muskmelon is a common and frequently-occurring postharvest disease, which leads to quality deterioration and neosolaniol (NEO) contamination. New strategies to control postharvest decay and reduce NEO contamination are of paramount importance. The effects of acetylsalicylic acid (ASA) treatment on the growth of Fusarium sulphureum in vitro, and Fusarium rot development and NEO accumulation in fruits inoculated with F. sulphureum in vivo were investigated. The results showed that ASA inhibited the growth of F. sulphureum, evident morphological and major cellular changes were observed under the microscope. In vivo testing showed that 3.2 mg/mL ASA significantly suppressed Fusarium rot development and NEO accumulation after 6 and 8 d of pathogen inoculation. Meanwhile, Tri gene expressions involved in NEO biosynthesis were down-regulated after treatment. Taken together, ASA treatment not only reduced Fusarium rot development by inhibiting the growth of F. sulphureum, but decreased NEO accumulation by suppressing NEO biosynthesis pathway.


Assuntos
Aspirina/farmacologia , Cucurbitaceae/química , Tricotecenos/metabolismo , Cucurbitaceae/metabolismo , Cucurbitaceae/microbiologia , Frutas/química , Frutas/metabolismo , Frutas/microbiologia , Fusarium/efeitos dos fármacos , Fusarium/crescimento & desenvolvimento , Fusarium/metabolismo , Fusarium/ultraestrutura , Doenças das Plantas/prevenção & controle , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Tricotecenos/química
16.
Genome ; 62(5): 329-339, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30933665

RESUMO

Fluorescence in situ hybridization (FISH) using oligonucleotides is a simple and convenient method for chromosome research. In this study, 34 of 46 previously developed oligonucleotides produced signals in barley. Together with two plasmid clones and one PCR-amplified cereal centromere repeat (CCS1) probe, 37 repetitive sequences were chromosomally located produced three types of signals covering different positions on the chromosomes. The centromeric and pericentric regions had a more complex genomic organization and sequence composition probably indicative of higher contents of heterochromatin. An efficient multi-plex probe containing eight oligonucleotides and a plasmid clone of 45S rDNA was developed. Thirty-three barley karyotypes were developed and compared. Among them, 11 irradiation-induced mutants of cultivar 08-49 showed no chromosomal variation, whereas 22 cultivar and landrace accessions contained 28 chromosomal polymorphisms. Chromosome 4H was the most variable and 6H was the least variable based on chromosome polymorphic information content (CPIC). Five polymorphic chromosomes (1H-2, 2H-1, 3H-3, 5H-2, and 6H-2) were dominant types, each occurring in more than 50% of accessions. The multi-plex probe should facilitate identification of further chromosomal polymorphisms in barley.


Assuntos
Cromossomos de Plantas/genética , Hordeum/genética , Polimorfismo Genético/genética , Sequências Repetitivas de Ácido Nucleico/genética , Centrômero/genética , Sondas de DNA/genética , Hibridização in Situ Fluorescente , Cariótipo , Cariotipagem , Oligonucleotídeos/genética , RNA de Plantas/genética , RNA Ribossômico/genética
17.
Methods Mol Biol ; 1933: 33-48, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30945177

RESUMO

Noncoding RNAs perform diverse regulatory functions in living cells. In plants, two RNA polymerase II-related enzymes, RNA polymerases IV and V (Pol IV and V), specialize in the synthesis of noncoding RNAs that silence a subset of transposable elements and genes via RNA-directed DNA methylation (RdDM). In this process, Pol IV partners with RNA-dependent RNA polymerase 2 (RDR2) to produce double-stranded RNAs that are then cut by an RNase III enzyme, Dicer-like 3 (DCL3), into 24 nt small interfering RNAs (siRNAs). The siRNAs are loaded into an Argonaute family protein, primarily AGO4, and guide the complex to complementary DNA target sequences where RdDM and repressive chromatin modifications ensue. The dependence of 24 nt siRNA biogenesis on Pol IV and RDR2 has been known for more than a decade, but the elusive pre-siRNA transcripts synthesized by Pol IV and RDR2 have only recently been identified. This chapter describes the approaches that enabled our identification of Pol IV/RDR2-dependent RNAs (P4R2 RNAs) in Arabidopsis thaliana. These included the use of a triple Dicer mutant (dcl2 dcl3 dcl4) to cause P4R2 RNAs to accumulate, genome-wide identification and mapping of P4R2 RNAs using a modified Illumina small RNA-Seq protocol, and multiple bioinformatic pipelines for data analysis and displaying results.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA Polimerase II/genética , Precursores de RNA/genética , RNA de Plantas/genética , RNA Interferente Pequeno/genética , Proteínas de Arabidopsis/antagonistas & inibidores , Biologia Computacional/métodos , Regulação da Expressão Gênica de Plantas , RNA Polimerase II/antagonistas & inibidores
18.
BMC Genomics ; 20(1): 269, 2019 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-30947685

RESUMO

BACKGROUND: Chinese kale (Brassica alboglabra) contains high nutritional elements and functional molecules, especially anticarcinogenic and antioxidant glucosinolates (GS), which was highly affected by environment temperature. To investigate the link of GS biosynthesis with heat stress response in Chinese kale, global transcription profiles of high-GS line (HG), low-GS line (LG), high-GS line under heat stress (HGT) and low-GS line under heat stress (LGT) were analyzed. RESULTS: Based on three biological replicates of each RNA sequencing data, 3901, 4062 and 2396 differentially expressed genes in HG vs HGT, LG vs LGT and HGT vs LGT were obtained, respectively. GO annotation, KEGG pathway analysis and a comprehensive analysis of DEGs showed a strong correlation between the GS biosynthesis and heat stress response. It was noticed that 11 differentially expressed genes tied to the GS biosynthesis were down-regulated, 23 heat shock transcription factors and 61 heat shock proteins were up-regulated upon the heat treatment. Another two Chinese kale varieties Cuibao and Shunbao with high- and low- GS content respectively, were used to validate the relationship of GS content and heat-response, and the results showed that high-GS content variety were more thermotolerant than the low-GS content one although GS significantly decreased in both varieties under heat stress. In addition, HSP100/ClpB, HSP90, HSP70 and sHSPs were differentially expressed in high- and low-GS varieties. Notably, HSP90 and sHSPs showed an obviously early response to heat stress than other related genes. CONCLUSION: The higher heat resistance of high-GS Chinese kale and the sharp decrease of glucosinolate content under heat stress indicated a strong relationship of GS accumulation and heat stress response. Combined with the previous report on the low expression of HSP90 at elevated temperatures in GS-deficient mutant TU8 of Arabidopsis, the differential expression pattern of HSP90 in high- and low- GS varieties and its early heat response implied it might be a key regulator in GS metabolism and heat-resistance in Chinese kale.


Assuntos
Brassica/genética , Regulação da Expressão Gênica de Plantas , Resposta ao Choque Térmico , Plântula/química , Transcriptoma , Antioxidantes/metabolismo , Brassica/fisiologia , Perfilação da Expressão Gênica , Glucosinolatos/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , RNA de Plantas/genética
19.
J Agric Food Chem ; 67(18): 5072-5084, 2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-30986354

RESUMO

Alternative splicing (AS), the process of removing introns from pre-mRNA and the rearrangement of exons to produce several types of mature transcripts, is a remarkable step preceding protein synthesis. In particular, it has now been conclusively shown that up to ∼95% of genes are alternatively spliced to generate a complex and diverse proteome in eukaryotic organisms. Consequently, AS is one of the determinants of the functional repertoire of cells. Many studies have revealed that AS in plants can be regulated by cell type, developmental stage, environmental stress, and the circadian clock. Moreover, increasing amounts of evidence reveal that chemical compounds can affect various steps during splicing to induce major effects on plant physiology. Hence, the chemical modulation of AS can serve as a good strategy for molecular-target identification in attempts to potentially control plant genetics. However, the kind of mechanisms involved in the chemical modulation of AS that can be used in agrochemical research remain largely unknown. This review introduces recent studies describing the specific roles AS plays in plant adaptation to environmental stressors and in the regulation of development. We also discuss recent advances in small molecules that induce alterations of AS and the possibility of using this strategy in agrochemical-target identification, giving a new direction for potential genetic control in agrochemical research.


Assuntos
Agroquímicos/farmacologia , Processamento Alternativo , Plantas/efeitos dos fármacos , Plantas/genética , Processamento Alternativo/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Desenvolvimento Vegetal/efeitos dos fármacos , Plantas/química , Plantas/metabolismo , RNA de Plantas/genética
20.
Methods Mol Biol ; 1933: 49-65, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30945178

RESUMO

Maize endosperm consists of three distinct types of tissues, including the starchy endosperm (SE), the basal endosperm transfer cell layer (BETL), and the aleurone cell layer (AL). Compartmentalization of these tissues during endosperm differentiation makes the endosperm development an excellent model to study changes in gene expression during development. By utilizing cryo-dissection of developing endosperm, morphologically distinct samples can be obtained for transcriptome and epigenome analysis. Here, we describe methods for the isolation of tissues from developing maize endosperm and for the transcriptome analysis to identify novel long noncoding RNAs. The transcriptome data can be further analyzed to illustrate spatiotemporal changes in both coding and noncoding transcripts during the endosperm development.


Assuntos
Endosperma/genética , Perfilação da Expressão Gênica/métodos , Genes de Plantas/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA Longo não Codificante/genética , RNA de Plantas/genética , Zea mays/genética , Biologia Computacional/métodos , Endosperma/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas , RNA de Plantas/isolamento & purificação , Transcriptoma , Zea mays/crescimento & desenvolvimento
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