RESUMO
Brassica rapa L., which includes Chinese cabbage, turnip, and pak choi, has more complex flowering time regulation than does Arabidopsis thaliana due to the presence of multiple paralogous flowering time genes. FLOWERING LOCUS C (FLC) is one of the key genes regulating the flowering time, and B. rapa has four FLC paralogs. BrFLC5 on the reference genome is deemed a pseudogene because of a mutation (from G to A) in the splice site of the third intron, but there are some accessions with a G nucleotide in the splice site. In this study, we genotyped 310 B. rapa accessions and found that 19 had homozygous and 81 had heterozygous putative functional BrFLC5 alleles. Accessions of turnip showed the highest proportion with a functional BrFLC5 allele. BrFLC5 acts as a floral repressor when overexpressed in A. thaliana. The BrFLC5 expression level varied in pre-vernalized plants, and this transcriptional variation was not associated with the G/A polymorphism in the third intron. Three accessions having a higher BrFLC5 expression in pre-vernalized plants had a 584-bp insertion in the promoter region. Many regions homologous to this 584-bp sequence are present in the B. rapa genome, and this 584-bp inserted region has tandem duplications of an AT-rich sequence in its central region. The possibility that a high expression of a functional BrFLC5 could contribute to producing premature bolting-resistant lines in B. rapa vegetables is discussed. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01405-0.
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Glucosinolates (GSLs), precursors of isothiocyanates (ITCs), are present in Brassicaceae plants have been found to have health benefits. Sulforaphane (4-(methylsulfinyl)butyl ITC) is an ITC stored in the form of 4-(methylsulfinyl)butyl GSL (glucoraphanin, 4MSOB) in Brassica vegetables, such as broccoli and kale. Sulforaphane activates Nrf2 expression, a transcription factor responsible for inducing physiological activities such as detoxification in the human body, and it represents a functional component unique to cruciferous vegetables. Raphanobrassica is an inter-generic hybrid between radish and kale, and it contains a high amount of 4MSOB. However, Raphanobrassica contains as much 4-methylsulfinyl-3-butenyl GSL (glucoraphenin, 4MSO3B) as it does 4MSOB. GLUCORAPHASATIN SYNTHASE 1 (GRS1) is an enzyme present in radish that synthesizes 4-methylthio-3-butenyl GSL (glucoraphasatin, 4MT3B), a precursor of 4MSO3B, using 4-(methylthio)butyl GSL (glucoerucin, 4MTB) as a substrate. Since the precursor of 4MSOB is also 4MTB, it was considered that both 4MSOB and 4MSO3B accumulate owing to competition in Raphanobrassica. We hypothesized that owing to the impaired function of GRS1 in Raphanobrassica, it may be possible to breed Raphanobrassica cultivars containing a high 4MSOB content. In this study, we generated Raphanobrassica populations with functional and defective GRS1 and compared the GSL composition in the two populations using high-performance liquid chromatography. The mean 4MSOB content in leaves of the defective-type populations was higher than that in the functional-type population, and the defective/functional ratio ranged from 2.02 to 2.51-fold, supporting this hypothesis. Furthermore, leaves, flower buds, stems, and roots contained higher amounts of 4MSOB in the defective population than in the functional population. The leaf 4MSOB content of defective Raphanobrassica grown in this study was comparable to that of previously studied vegetables (such as broccoli sprouts) with high 4MSOB content. Raphanobrassica with defective GRS1 represents a new leafy vegetable with high 4MSOB content which exhibits anti-cancerous and anti-inflammatory potentials.
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The majority of genes encoding photosynthesis-associated proteins in the nucleus are induced by light during photomorphogenesis, allowing plants to establish photoautotrophic growth. Therefore, optimizing the protein import apparatus of plastids, designated as the translocon at the outer and inner envelope membranes of chloroplast (TOC-TIC) complex, upon light exposure is a prerequisite to the import of abundant nuclear-encoded photosynthesis-associated proteins. However, the mechanism that coordinates the optimization of the TOC-TIC complex with the expression of nuclear-encoded photosynthesis-associated genes remains to be characterized in detail. To address this question, we investigated the mechanism by which plastid protein import is regulated by light during photomorphogenesis in Arabidopsis. We found that the albino plastid protein import2 (ppi2) mutant lacking Toc159 protein import receptors have active photoreceptors, even though the mutant fails to induce the expression of photosynthesis-associated nuclear genes upon light illumination. In contrast, many TOC and TIC genes are rapidly induced by blue light in both WT and the ppi2 mutant. We uncovered that this regulation is mediated primarily by cryptochrome 1 (CRY1). Furthermore, deficiency of CRY1 resulted in the decrease of some TOC proteins in vivo. Our results suggest that CRY1 plays key roles in optimizing the content of the TOC-TIC apparatus to accommodate the import of abundant photosynthesis-associated proteins during photomorphogenesis.
Assuntos
Arabidopsis/fisiologia , Criptocromos/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Genes de Plantas , Luz , Morfogênese , Fotossíntese/genética , Arabidopsis/genéticaRESUMO
The genus Brassica includes oil crops, vegetables, condiments, fodder crops, and ornamental plants. Brassica species underwent a whole genome triplication event after speciation between ancestral species of Brassica and closely related genera including Arabidopsis thaliana. Diploid species such as Brassica rapa and Brassica oleracea have three copies of genes orthologous to each A. thaliana gene, although deletion in one or two of the three homologs has occurred in some genes. The floral transition is one of the crucial events in a plant's life history, and time of flowering is an important agricultural trait. There is a variation in flowering time within species of the genus Brassica, and this variation is largely dependent on a difference in vernalization requirements. In Brassica, like in A. thaliana, the key gene of vernalization is FLOWERING LOCUS C (FLC). In Brassica species, the vernalization response including the repression of FLC expression by cold treatment and the enrichment of the repressive histone modification tri-methylated histone H3 lysine 27 (H3K27me3) at the FLC locus is similar to A. thaliana. B. rapa and B. oleracea each have four paralogs of FLC, and the allotetraploid species, Brassica napus, has nine paralogs. The increased number of paralogs makes the role of FLC in vernalization more complicated; in a single plant, paralogs vary in the expression level of FLC before and after vernalization. There is also variation in FLC expression levels between accessions. In this review, we focus on the regulatory circuits of the vernalization response of FLC expression in the genus Brassica.
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There is a wide variation of flowering time among lines of Brassica rapa L. Most B. rapa leafy (Chinese cabbage etc.) or root (turnip) vegetables require prolonged cold exposure for flowering, known as vernalization. Premature bolting caused by low temperature leads to a reduction in the yield/quality of these B. rapa vegetables. Therefore, high bolting resistance is an important breeding trait, and understanding the molecular mechanism of vernalization is necessary to achieve this goal. In this study, we demonstrated that BrFRIb functions as an activator of BrFLC in B. rapa. We showed a positive correlation between the steady state expression levels of the sum of the BrFLC paralogs and the days to flowering after four weeks of cold treatment, suggesting that this is an indicator of the vernalization requirement. We indicate that BrFLCs are repressed by the accumulation of H3K27me3 and that the spreading of H3K27me3 promotes stable FLC repression. However, there was no clear relationship between the level of H3K27me3 in the BrFLC and the vernalization requirement. We also showed that if there was a high vernalization requirement, the rate of repression of BrFLC1 expression following prolonged cold treatments was lower.
Assuntos
Brassica rapa/fisiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Brassica rapa/classificação , Brassica rapa/genética , Resposta ao Choque Frio , Flores/classificação , Flores/genética , Flores/fisiologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Histonas/metabolismo , Proteínas de Domínio MADS/genética , Proteínas de Domínio MADS/metabolismo , Análise de Sequência de DNA , Verduras/classificação , Verduras/genética , Verduras/fisiologiaRESUMO
Brassica rapa L. is an important agricultural crop that requires a period of prolonged cold for flowering. This process is known as vernalization. Studies have shown that long noncoding RNAs (lncRNAs) play important roles in abiotic stress responses and several cold-responsive noncoding RNAs have been suggested to be involved in vernalization. We examined the transcriptome of the Chinese cabbage inbred line (B. rapa L. var. pekinensis) RJKB-T24, and identified 1,444 long intergenic noncoding RNAs (lincRNAs), 551 natural antisense transcripts (NATs), and 93 intronic noncoding RNAs (incRNAs); 549 of the 2,088 lncRNAs significantly altered their expression in response to four weeks of cold treatment. Most differentially expressed lncRNAs did not lead to a change of expression levels in mRNAs covering or near lncRNAs, suggesting that the transcriptional responses to four weeks of cold treatment in lncRNA and mRNA are independent. However, some differentially expressed mRNAs had NATs with expression altered in the same direction. These genes were categorized as having an abiotic stress response, suggesting that the paired-expression may play a role in the transcriptional response to vernalization or cold treatment. We also identified short-term cold treatment induced NATs in BrFLC and BrMAF genes, which are involved in vernalization. The lncRNAs we identified differed from those reported in Arabidopsis thaliana, suggesting the role of lncRNAs in vernalization differ between these two species.
Assuntos
Brassica rapa/genética , Temperatura Baixa , RNA Longo não Codificante/genética , RNA de Plantas/genética , Arabidopsis/genética , Produtos Agrícolas/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Íntrons , Oligonucleotídeos Antissenso , Plantas Geneticamente Modificadas/genética , RNA Antissenso/genética , RNA-Seq , Estresse Fisiológico , TranscriptomaRESUMO
Flowering time is an important agronomic trait for Brassica rapa crops, and previous breeding work in Brassica has successfully transmitted other important agronomic traits from donor species. However, there has been no previous attempts to produce hybrids replacing the original Brassica FLC alleles with alien FLC alleles. In this paper, we introduce the creation of a chromosome substitution line (CSSL) containing a homozygous introgression of Flowering Locus C from Brassica oleracea (BoFLC2) into a B. rapa genomic background, and characterize the CSSL line with respect to the parental cultivars. The preferential transmission of alien chromosome inheritance and the pattern of transmission observed during the production of the CSSLs are also discussed.
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Epigenetic regulation, covalent modification of DNA and changes in histone proteins are closely linked to plant development and stress response through flexibly altering the chromatin structure to regulate gene expression. In this review, we will illustrate the importance of epigenetic influences by discussing three agriculturally important traits of Brassicaceae. (1) Vernalization, an acceleration of flowering by prolonged cold exposure regulated through epigenetic silencing of a central floral repressor, FLOWERING LOCUS C. This is associated with cold-dependent repressive histone mark accumulation, which confers competency of consequence vegetative-to-reproductive phase transition. (2) Hybrid vigor, in which an F1 hybrid shows superior performance to the parental lines. Combination of distinct epigenomes with different DNA methylation states between parental lines is important for increase in growth rate in a hybrid progeny. This is independent of siRNA-directed DNA methylation but dependent on the chromatin remodeler DDM1. (3) Self-incompatibility, a reproductive mating system to prevent self-fertilization. This is controlled by the S-locus consisting of SP11 and SRK which are responsible for self/non-self recognition. Because self-incompatibility in Brassicaceae is sporophytically controlled, there are dominance relationships between S haplotypes in the stigma and pollen. The dominance relationships in the pollen rely on de novo DNA methylation at the promoter region of a recessive allele, which is triggered by siRNA production from a flanking region of a dominant allele.
Assuntos
Brassicaceae/genética , Produtos Agrícolas/genética , Epigênese Genética , Vigor Híbrido/genética , Autoincompatibilidade em Angiospermas/genética , Proteínas de Arabidopsis/genética , Brassicaceae/fisiologia , Montagem e Desmontagem da Cromatina/fisiologia , Produtos Agrícolas/fisiologia , Metilação de DNA , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica de Plantas , Haplótipos , Proteínas de Domínio MADS/genética , Pólen/genética , RNA Interferente Pequeno , Fatores de Transcrição/genéticaRESUMO
Glucosinolates (GSLs) are secondary metabolites whose degradation products confer intrinsic flavors and aromas to Brassicaceae vegetables. Several structures of GSLs are known in the Brassicaceae, and the biosynthetic pathway and regulatory networks have been elucidated in Arabidopsis (Arabidopsis thaliana). GSLs are precursors of chemical defense substances against herbivorous pests. Specific GSLs can act as feeding blockers or stimulants, depending on the pest species. Natural selection has led to diversity in the GSL composition even within individual species. However, in radish (Raphanus sativus), glucoraphasatin (4-methylthio-3-butenyl glucosinolate) accounts for more than 90% of the total GSLs, and little compositional variation is observed. Because glucoraphasatin is not contained in other members of the Brassicaceae, like Arabidopsis and cabbage (Brassica oleracea), the biosynthetic pathways for glucoraphasatin remain unclear. In this report, we identified and characterized a gene encoding GLUCORAPHASATIN SYNTHASE 1 (GRS1) by genetic mapping using a mutant that genetically lacks glucoraphasatin. Transgenic Arabidopsis, which overexpressed GRS1 cDNA, accumulated glucoraphasatin in the leaves. GRS1 encodes a 2-oxoglutarate-dependent dioxygenase, and it is abundantly expressed in the leaf. To further investigate the biosynthesis and transportation of GSLs in radish, we grafted a grs1 plant onto a wild-type plant. The grafting experiment revealed a leaf-to-root long-distance glucoraphasatin transport system in radish and showed that the composition of GSLs differed among the organs. Based on these observations, we propose a characteristic biosynthesis pathway for glucoraphasatin in radish. Our results should be useful in metabolite engineering for breeding of high-value vegetables.
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Dioxigenases/metabolismo , Glucosinolatos/biossíntese , Ácidos Cetoglutáricos/metabolismo , Raphanus/metabolismo , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/metabolismo , Transporte Biológico , Vias Biossintéticas/genética , Cromatografia Líquida de Alta Pressão , Dioxigenases/classificação , Dioxigenases/genética , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Glucosinolatos/análise , Engenharia Metabólica/métodos , Mutação , Filogenia , Folhas de Planta/genética , Folhas de Planta/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Raphanus/enzimologia , Raphanus/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Plântula/genética , Plântula/metabolismo , Plântula/fisiologia , Homologia de Sequência de AminoácidosRESUMO
Arabidopsis (Arabidopsis thaliana) GOLDEN2-LIKE (GLK) transcription factors promote chloroplast biogenesis by regulating the expression of photosynthesis-related genes. Arabidopsis GLK1 is also known to participate in retrograde signaling from chloroplasts to the nucleus. To elucidate the mechanism by which GLK1 is regulated in response to plastid signals, we biochemically characterized Arabidopsis GLK1 protein. Expression analysis of GLK1 protein indicated that GLK1 accumulates in aerial tissues. Both tissue-specific and Suc-dependent accumulation of GLK1 were regulated primarily at the transcriptional level. In contrast, norflurazon- or lincomycin-treated gun1-101 mutant expressing normal levels of GLK1 mRNA failed to accumulate GLK1 protein, suggesting that plastid signals directly regulate the accumulation of GLK1 protein in a GUN1-independent manner. Treatment of the glk1glk2 mutant expressing functional GFP-GLK1 with a proteasome inhibitor, MG-132, induced the accumulation of polyubiquitinated GFP-GLK1. Furthermore, the level of endogenous GLK1 in plants with damaged plastids was partially restored when those plants were treated with MG-132. Collectively, these data indicate that the ubiquitin-proteasome system participates in the degradation of Arabidopsis GLK1 in response to plastid signals.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Plastídeos/metabolismo , Fatores de Transcrição/metabolismo , Ubiquitina/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Núcleo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Leupeptinas/farmacologia , Plantas Geneticamente Modificadas , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteassoma/farmacologia , Fatores de Transcrição/genéticaRESUMO
To facilitate prevention of clubroot disease, a major threat to the successful cultivation of Chinese cabbage (Brassica rapa L.), we bred clubroot-resistant (CR) cultivars by introducing resistance genes from CR turnips via conventional breeding. Among 11 CR loci found in B. rapa, we identified CRb in Chinese cabbage cultivar 'CR Shinki' as a single dominant gene for resistance against Plasmodiophora brassicae pathotype group 3, against which the stacking of Crr1 and Crr2 loci was not effective. However, the precise location and pathotype specificity of CRb have been controversial, because CRa and Rcr1 also map near this locus. Previously, our fine-mapping study revealed that CRb is located in a 140-kb genomic region on chromosome A03. Here, we determined the nucleotide sequence of an approximately 64-kb candidate region in the resistant line; this region contains six open reading frames (ORFs) similar to NB-LRR encoding genes that are predicted to occur in tandem with the same orientation. Among the six ORFs present, only four on the genome of the resistant line showed a strong DNA sequence identity with each other, and only one of those four could confer resistance to P. brassicae isolate No. 14 of the pathotype group 3. These results suggest that these genes evolved through recent gene duplication and uneven crossover events that could lead to the acquisition of clubroot resistance. The DNA sequence of the functional ORF was identical to that of the previously cloned CRa gene; thus, we showed that the independently identified CRb and CRa are one and the same clubroot-resistance gene.
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Brassica rapa/genética , Genes de Plantas , Doenças das Plantas/genética , Sequências de Repetição em Tandem , Sequência de Bases , Brassica rapa/parasitologia , Mapeamento Cromossômico , Biblioteca Gênica , Genes Dominantes , Vetores Genéticos , Fases de Leitura Aberta , Fenótipo , Mapeamento Físico do Cromossomo , Plasmodioforídeos , Análise de Sequência de DNARESUMO
In diploid organisms, phenotypic traits are often biased by effects known as Mendelian dominant-recessive interactions between inherited alleles. Phenotypic expression of SP11 alleles, which encodes the male determinants of self-incompatibility in Brassica rapa, is governed by a complex dominance hierarchy1-3. Here, we show that a single polymorphic 24 nucleotide small RNA, named SP11 methylation inducer 2 (Smi2), controls the linear dominance hierarchy of the four SP11 alleles (S44 > S60 > S40 > S29). In all dominant-recessive interactions, small RNA variants derived from the linked region of dominant SP11 alleles exhibited high sequence similarity to the promoter regions of recessive SP11 alleles and acted in trans to epigenetically silence their expression. Together with our previous study4, we propose a new model: sequence similarity between polymorphic small RNAs and their target regulates mono-allelic gene expression, which explains the entire five-phased linear dominance hierarchy of the SP11 phenotypic expression in Brassica.
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KEY MESSAGE: Genetic analysis and gene mapping of the 4-methylthio-3-butenyl glucosinolate-less trait of white radish were performed and a white radish cultivar with new glucosinolate composition was developed. A spontaneous mutant having significantly low 4-methylthio-3-butenyl glucosinolate (4MTB-GSL) content was identified from a landrace of Japanese white radish (Raphanus sativus L.) through intensive evaluation of glucosinolate profiles of 632 lines including genetic resources and commercial cultivars using high-performance liquid chromatography (HPLC) analysis. A line lacking 4MTB-GSL was developed using the selected mutant as a gene source. Genetic analyses of F1, F2, and BC1F1 populations of this line suggested that the 4MTB-GSL-less trait is controlled by a single recessive allele. Using SNP and SCAR markers, 96 F2 plants were genotyped, and a linkage map having nine linkage groups with a total map distance of 808.3 cM was constructed. A gene responsible for the 4MTB-GSL-less trait was mapped between CL1753 and CL5895 at the end of linkage group 1. The genetic distance between these markers was 4.2 cM. By selfing and selection of plants lacking 4MTB-GSL, a new cultivar, 'Daikon parental line No. 5', was successfully developed. This cultivar was characterized by glucoerucin, which accounted for more than 90% of the total glucosinolates (GSLs). The total GSL content in roots was ca. 12 µmol/g DW, significantly lower than those in common white radish cultivars. Significance of this line in radish breeding is discussed.
Assuntos
Glucosinolatos/química , Raphanus/química , Raphanus/genética , Alelos , Cromatografia Líquida de Alta Pressão , Mapeamento Cromossômico , Genes de Plantas , Genes Recessivos , Ligação Genética , Genótipo , Melhoramento Vegetal , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Unique secondary metabolites, glucosinolates (S-glucopyranosyl thiohydroximates), are naturally occurring S-linked glucosides found mainly in Brassicaceae plants. They are enzymatically hydrolyzed to produce sulfate ions, D-glucose, and characteristic degradation products such as isothiocyanates. The functions of glucosinolates in the plants remain unclear, but isothiocyanates possessing a pungent or irritating taste and odor might be associated with plant defense from microbes. Isothiocyanates have been studied extensively in experimental in vitro and in vivo carcinogenesis models for their cancer chemopreventive properties. The beneficial isothiocyanates, glucosinolates that are functional for supporting human health, have received attention from many scientists studying plant breeding, plant physiology, plant genetics, and food functionality. This review presents a summary of recent topics related with glucosinolates in the Brassica family, along with a summary of the chemicals, metabolism, and genes of glucosinolates in Brassicaceae. The bioavailabilities of isothiocyanates from certain functional glucosinolates and the importance of breeding will be described with emphasis on glucosinolates.
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Biogenesis of chloroplasts is essential for plant growth and development. A number of homozygous mutants lacking a chloroplast protein exhibit an albino phenotype. In general, it is challenging to grow albino Arabidopsis plants on soil until they set seeds. Homozygous albino mutants are usually obtained as progenies of heterozygous parents. Here, we describe a method of recovering seeds from the seedling lethal Arabidopsis mutant ppi2-2, which lacks the atToc159 protein import receptor at the outer envelope membrane of chloroplast. Using plastic containers, we were able to grow homozygous ppi2-2 plants until these set seed. Although the germination rate of the harvested seeds was relatively low, it was still sufficient to allow us to further analyze the ppi2-2 progeny. Using ppi2-2 homozygous seeds, we were able to analyze the role of plastid protein import in the light-regulated induction of nuclear genes. We propose that this method be applied to other seedling lethal Arabidopsis mutants to obtain homozygous seeds, helping us further investigate the roles of plastid proteins in plant growth and development.
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Radish (Raphanus sativus L., n = 9) is one of the major vegetables in Asia. Since the genomes of Brassica and related species including radish underwent genome rearrangement, it is quite difficult to perform functional analysis based on the reported genomic sequence of Brassica rapa. Therefore, we performed genome sequencing of radish. Short reads of genomic sequences of 191.1 Gb were obtained by next-generation sequencing (NGS) for a radish inbred line, and 76,592 scaffolds of ≥ 300 bp were constructed along with the bacterial artificial chromosome-end sequences. Finally, the whole draft genomic sequence of 402 Mb spanning 75.9% of the estimated genomic size and containing 61,572 predicted genes was obtained. Subsequently, 221 single nucleotide polymorphism markers and 768 PCR-RFLP markers were used together with the 746 markers produced in our previous study for the construction of a linkage map. The map was combined further with another radish linkage map constructed mainly with expressed sequence tag-simple sequence repeat markers into a high-density integrated map of 1,166 cM with 2,553 DNA markers. A total of 1,345 scaffolds were assigned to the linkage map, spanning 116.0 Mb. Bulked PCR products amplified by 2,880 primer pairs were sequenced by NGS, and SNPs in eight inbred lines were identified.
Assuntos
Genoma de Planta , Raphanus/genética , Análise de Sequência de DNA , Brassica rapa/genética , Marcadores Genéticos , Polimorfismo de Nucleotídeo ÚnicoRESUMO
SNP markers for QTL analysis of 4-MTB-GSL contents in radish roots were developed by determining nucleotide sequences of bulked PCR products using a next-generation sequencer. DNA fragments were amplified from two radish lines by multiplex PCR with six primer pairs, and those amplified by 2,880 primer pairs were mixed and sequenced. By assembling sequence data, 1,953 SNPs in 750 DNA fragments, 437 of which have been previously mapped in a linkage map, were identified. A linkage map of nine linkage groups was constructed with 188 markers, and five QTLs were detected in two F(2) populations, three of them accounting for more than 50% of the total phenotypic variance being repeatedly detected. In the identified QTL regions, nine SNP markers were newly produced. By synteny analysis of the QTLs regions with Arabidopsis thaliana and Brassica rapa genome sequences, three candidate genes were selected, i.e., RsMAM3 for production of aliphatic glucosinolates linked to GSL-QTL-4, RsIPMDH1 for leucine biosynthesis showing strong co-expression with glucosinolate biosynthesis genes linked to GSL-QTL-2, and RsBCAT4 for branched-chain amino acid aminotransferase linked to GSL-QTL-1. Nucleotide sequences and expression of these genes suggested their possible function in 4MTB-GSL biosynthesis in radish roots.
Assuntos
DNA de Plantas/genética , Glucosinolatos/biossíntese , Proteínas de Plantas/genética , Raízes de Plantas/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Raphanus/genética , 2-Isopropilmalato Sintase/genética , 2-Isopropilmalato Sintase/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Sequência de Bases , Brassica rapa/genética , Brassica rapa/metabolismo , Mapeamento Cromossômico , Cromossomos de Plantas , Etiquetas de Sequências Expressas , Sequenciamento de Nucleotídeos em Larga Escala , Malato Desidrogenase/genética , Malato Desidrogenase/metabolismo , Dados de Sequência Molecular , Reação em Cadeia da Polimerase Multiplex , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Raphanus/metabolismo , Sintenia , Transaminases/genética , Transaminases/metabolismoRESUMO
To reveal varietally differing glucosinolate (GSL) contents in radish (Raphanus sativus L.) cultivated in Japan, the total and individual GSLs of 28 cultivars were analyzed using high-performance liquid chromatography. In these cultivars, GSL types including three aliphatic GSLs (glucoraphenin, glucoerucin, and 4-methylthio-3-butenyl GSL (4MTB-GSL)) and three indolyl GSLs (4-hydroxyglucobrassicin, glucobrassicin, and 4-methoxy-glucobrassicin) were detected. No cultivar-specific type of GSL was identified. The dominant GSL was 4MTB-GSL, but its contents differed remarkably: 8.6 µmol/g in 'Koushin' to 135.7 µmol/g in 'Karami 199'. Over about 90% of all GSLs in Japanese radish type are 4MTB-GSL, a higher percentage than in Chinese or European garden radish cultivars. A simple, rapid method for estimating total GSL contents in crude extracts was established because of the small variation of glucosinolate composition in Japanese cultivars. The total GSL content can be estimated using an equation for prediction with absorbance at 425 nm in a mixture of GSL crude extract and palladium (II) chloride solution: Total GSL (µmol/g) = 305.47 × A(425) - 29.66. Its coefficient of determination (R(2)) and standard error of prediction (SEP) are 0.968 and 8.052. This method enables total GSL content estimation from more than 200 samples per person per day.
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Retrograde signalling from the plastid to the nucleus, also known as plastid signalling, plays a key role in coordinating nuclear gene expression with the functional state of plastids. Inhibitors that cause plastid dysfunction have been suggested to generate specific plastid signals related to their modes of action. However, the molecules involved in plastid signalling remain to be identified. Genetic studies indicate that the plastid-localized pentatricopeptide repeat protein GUN1 mediates signalling under several plastid signalling-related conditions. To elucidate further the nature of plastid signals, investigations were carried out to determine whether different plastid signal-inducing treatments had similar effects on plastids and on nuclear gene expression. It is demonstrated that norflurazon and lincomycin treatments and the plastid protein import2-2 (ppi2-2) mutation, which causes a defect in plastid protein import, all resulted in similar changes at the gene expression level. Furthermore, it was observed that these three treatments resulted in defective RNA editing in plastids. This defect in RNA editing was not a secondary effect of down-regulation of pentatricopeptide repeat protein gene expression in the nucleus. The results indicate that these three treatments, which are known to induce plastid signals, affect RNA editing in plastids, suggesting an unprecedented link between plastid signalling and RNA editing.
Assuntos
Plastídeos , Transdução de Sinais , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Genes de Plantas , Mutação , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
Plastids are a diverse group of organelles found in plants and some parasites. Because genes encoding plastid proteins are divided between the nuclear and plastid genomes, coordinated expression of genes in two separate genomes is indispensable for plastid function. To coordinate nuclear gene expression with the functional or metabolic state of plastids, plant cells have acquired a retrograde signaling pathway from plastid to nucleus, also known as the plastid signaling pathway. To date, several metabolic processes within plastids have been shown to affect the expression of nuclear genes. Recent progress in this field has also revealed that the plastid signaling pathway interacts and shares common components with other intracellular signaling pathways. This review summarizes our current knowledge on retrograde signaling from plastid to nucleus in plant cells and its role in plant growth and development.