RESUMEN
Eukaryotic translation initiation factors (eIFs) are the primary targets for overcoming RNA virus resistance in plants. In a previous study, we mapped a BjeIF2Bß from Brassica juncea representing a new class of plant virus resistance genes associated with resistance to Turnip mosaic virus (TuMV). However, the mechanism underlying eIF2Bß-mediated virus resistance remains unclear. In this study, we discovered that the natural variation of BjeIF2Bß in the allopolyploid B. juncea was inherited from one of its ancestors, B. rapa. By editing of eIF2Bß, we were able to confer resistance to TuMV in B. juncea and in its sister species of B. napus. Additionally, we identified an N6-methyladenosine (m6A) demethylation factor, BjALKBH9B, for interaction with BjeIF2Bß, where BjALKBH9B co-localized with both BjeIF2Bß and TuMV. Furthermore, BjeIF2Bß recruits BjALKBH9B to modify the m6A status of TuMV viral coat protein RNA, which lacks the ALKB homologue in its genomic RNA, thereby affecting viral infection. Our findings have applications for improving virus resistance in the Brassicaceae family through natural variation or genome editing of the eIF2Bß. Moreover, we uncovered a non-canonical translational control of viral mRNA in the host plant.
RESUMEN
Mitochondrial F1F0-ATP synthase (F1F0-ATPase) inhibitor factor 1 (IF1) has been extensively characterized as an endogenous inhibitor that prevents the hydrolysis of adenosine-5'-triphosphate (ATP) by mitochondrial ATPases in mammals and yeasts; however, IF1's functions in plants remain unclear. Here, a comprehensive bioinformatic analysis was performed to identify plant mitochondrial F1F0-ATPase IF1 orthologs. Plant IF1s contain a conserved F1F0-ATPase inhibitory domain, but lack the antiparallel α-helical coiled-coil structure compared with mammalian IF1s. A subcellular localization analysis in Arabidopsis thaliana revealed that AtIF1-green fluorescent protein was present only in mitochondria. Additionally, AtIF1 was widely expressed in diverse organs and intense ß-glucuronidase staining was observed in reproductive tissues and germinating seeds. Compared with the wild-type and p35S:AtIF1-if1 etiolated seedlings, the ATP/ADP ratio was significantly lower in the AtIF1 T-DNA knockout seedlings (if1 mutant) growing under dark conditions, suggesting that AtIF1 can influence the energy state of cells. A significant reduction in seed yield and strong growth retardation under dark conditions were observed in the if1 mutant line. Furthermore, if1 plants exhibited a substantially decreased sensitivity to abscisic acid. Thus, the A. thaliana mitochondrial IF1, which is a conserved F1F0-ATPase inhibitor, is crucial for plant growth and responses to abscisic acid.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas/metabolismo , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas Fluorescentes Verdes , Mitocondrias/enzimología , Mitocondrias/metabolismo , Filogenia , Proteínas/genética , ATPasas de Translocación de Protón/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Alineación de Secuencia , Proteína Inhibidora ATPasaRESUMEN
Eukaryotic translation initiation factors (eIFs) are essential protein complexes involved in the translation of mRNA into proteins. These initiation factors are generally used as targets in the control of plant RNA virus infections. In the present study, we identified a total 190 eIFs, clustered phylogenetically into 40 distinct subfamilies in the allopolyploid Brassica juncea. Extensive evolutionary duplications of the eIFs in B. juncea suggest their increased genetic diversity and wide adaptability. The induction of expressions in some of the eIFs after inoculation against Turnip mosaic virus (TuMV) provided candidate targets to be used in the control of viral infections. In addition, the expression profiles of eIFs under different temperatures suggested that the TuMV epidemic was temperature dependent. The eIFs expressions suggested that the systemic viral infections were more acute in plants grown between 20 °C and 28 °C. In addition, our results revealed that new subgroups of eIFs, eIF2ß, eIF2α, eIF2Bß, EF1A, and PABP could be represented as targets for antiviral strategies in B. juncea. In summary, our findings would be helpful in studying the complex mechanisms of eIF-mediated, temperature-dependent RNA virus control in B. juncea.
RESUMEN
Following publication of this article, the authors have corrected 426 chimeric scaffolds in this genome (total scaffold number 10,684). The genome assembly has now been improved as V1.5, and the updated genome assembly is available to be downloaded from http://brassicadb.org/brad/datasets/pub/Genomes/Brassica_juncea/V1.5/ .
RESUMEN
Recessive resistances to plant viruses in the Potyvirus genus have been found to be based on mutations in the plant eukaryotic translation initiation factors, eIF4E and eIF4G or their isoforms. Here we report that natural, monogenic recessive resistance to the Potyvirus Turnip mosaic virus (TuMV) has been found in a number of mustard (Brassica juncea) accessions. Bulked segregant analysis and sequencing of resistant and susceptible plant lines indicated the resistance is controlled by a single recessive gene, recessive TuMV resistance 03 (retr03), an allele of the eukaryotic translation initiation factor 2B-beta (eIF2Bß). Silencing of eIF2Bß in a TuMV-susceptible mustard plant line and expression of eIF2Bß from a TuMV-susceptible line in a TuMV-resistant mustard plant line confirmed the new resistance mechanism. A functional copy of a specific allele of eIF2Bß is required for efficient TuMV infection. eIF2Bß represents a new class of virus resistance gene conferring resistance to any pathogen. eIF2B acts as a guanine nucleotide exchange factor (GEF) for its GTP-binding protein partner eIF2 via interaction with eIF2·GTP at an early step in translation initiation. Further genotyping indicated that a single non-synonymous substitution (A120G) in the N-terminal region of eIF2Bß was responsible for the TuMV resistance. A reproducible marker has been developed, facilitating marker-assisted selection for TuMV resistance in B. juncea. Our findings provide a new target for seeking natural resistance to potyviruses and new opportunities for the control of potyviruses using genome editing techniques targeted on eIF2Bß.
Asunto(s)
Factores Eucarióticos de Iniciación/metabolismo , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Enfermedades de las Plantas/genética , Proteínas de Plantas/metabolismo , Virus de Plantas/fisiología , Resistencia a la Enfermedad/genética , Factores Eucarióticos de Iniciación/genética , Genotipo , Proteínas de Plantas/genéticaRESUMEN
The Brassica genus encompasses three diploid and three allopolyploid genomes, but a clear understanding of the evolution of agriculturally important traits via polyploidy is lacking. We assembled an allopolyploid Brassica juncea genome by shotgun and single-molecule reads integrated to genomic and genetic maps. We discovered that the A subgenomes of B. juncea and Brassica napus each had independent origins. Results suggested that A subgenomes of B. juncea were of monophyletic origin and evolved into vegetable-use and oil-use subvarieties. Homoeolog expression dominance occurs between subgenomes of allopolyploid B. juncea, in which differentially expressed genes display more selection potential than neutral genes. Homoeolog expression dominance in B. juncea has facilitated selection of glucosinolate and lipid metabolism genes in subvarieties used as vegetables and for oil production. These homoeolog expression dominance relationships among Brassicaceae genomes have contributed to selection response, predicting the directional effects of selection in a polyploid crop genome.