Development of series of gateway binary vectors, pGWBs, for realizing efficient construction of fusion genes for plant transformation

Nakagawa, Tsuyoshi; Kurose, Takayuki; Hino, Takeshi; Tanaka, Katsunori; Kawamukai, Makoto; Niwa, Yasuo; Toyooka, Kiminori; Matsuoka, Ken; Jinbo, Tetsuro; Kimura, Takeshi

doi:10.1263/jbb.104.34

Cited by 1,528 publications

(1,427 citation statements)

References 26 publications

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“…To construct the 35S:uORFs TBF1 -LUC reporter, the 35S promoter and the TBF1 exon1 (including the R-motif, uORF1-uORF2, and the coding sequence of the first 73 amino acids of TBF1) were amplified from p35S:uORF1-uORF2-GUS 1 using Reporter-F/R primers, and ligated into pGWB235 22 via Gateway recombination. The 35S:ccdB cassette-LUC-NOS construct was generated by fusing PCR fragments of the 35S promoter from pMDC140 23 , the ccdB cassette and the NOS terminator from pRNAi-LIC 24 and LUC from pGWB235 22 .…”

Section: Methodsmentioning

confidence: 99%

“…The 35S:ccdB cassette-LUC-NOS construct was generated by fusing PCR fragments of the 35S promoter from pMDC140 23 , the ccdB cassette and the NOS terminator from pRNAi-LIC 24 and LUC from pGWB235 22 . The 35S:ccdB cassette-LUC-NOS was then inserted into pCAMBIA1300 via Pst I and EcoR I and designated as pGX301 for cloning 5′ leader sequences through replacement of the Apa I-flanked ccdB cassette 24 .…”

Section: Methodsmentioning

confidence: 99%

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Global translational reprogramming is a fundamental layer of immune regulation in plants

Greene

Yoo

et al. 2017

Nature

205

212

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In the absence of specialized immune cells, the need for plants to reprogram transcription to transition from growth-related activities to defence is well understood1, 2. However, little is known about translational changes that occur during immune induction. Using ribosome footprinting (RF), we performed global translatome profiling on Arabidopsis exposed to the microbe-associated molecular pattern (MAMP) elf18. We found that during this pattern-triggered immunity (PTI), translation was tightly regulated and poorly correlated with transcription. Identification of genes with altered translational efficiency (TE) led to the discovery of novel regulators of this immune response. Further investigation of these genes showed that mRNA sequence features are major determinants of the observed TE changes. In the 5′ leader sequences of transcripts with increased TE, we found a highly enriched mRNA consensus sequence, R-motif, consisting of mostly purines. We showed that R-motif regulates translation in response to PTI induction through interaction with poly(A)-binding proteins. Therefore, this study provides not only strong evidence, but also a molecular mechanism for global translational reprogramming during PTI in plants.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Global translational reprogramming is a fundamental layer of immune regulation in plants

Greene

Yoo

et al. 2017

Nature

205

212

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“…The TBF1 promoter was amplified from the Arabidopsis genomic DNA and flanked with Hind III/ Spe I using primers P14/P15 and ligated into pGX179, which was cut with Hind III/ Xba I, to generate pGX181 ( TBF1p:uorfs TBF1 -Gateway-NOS ). LUC , GFP ER and snc1 were amplified from pGWB235 26 , GFP-HDEL 27 and the snc1 mutant genomic DNA, respectively. TBF1-YFP and NPR1-EGFP were fused together through PCR, cloned via ligation independent cloning 25 .…”

Section: Methodsmentioning

confidence: 99%

“…For fitness test, rice was grown during the normal rice growing season (From Nov. 2015 to May 2016) under field conditions in Lingshui, Hainan (18° N latitude). Agrobacterium -mediated transformation into the Oryza sativa cultivar ZH11 was used to obtain transgenic rice plants 26 . For Xoo infection in the greenhouse (performed in year 2016), rice was grown for 3 weeks from Feb. 2 and inoculated on Feb. 23 with data collection on Mar.…”

Section: Methodsmentioning

confidence: 99%

uORF-mediated translation allows engineered plant disease resistance without fitness costs

Yuan

et al. 2017

Nature

283

189

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Controlling plant disease has been a struggle for mankind since the advent of agriculture. Studies of plant immune mechanisms have led to strategies of engineering resistant crops through ectopic transcription of plants’ own defence genes, such as the master immune regulatory gene NPR11. However, enhanced resistance obtained through such strategies is often associated with significant penalties to fitness2, making the resulting products undesirable for agricultural applications. To remedy this problem, we sought more stringent mechanisms of expressing defence proteins. Based on our latest finding that translation of key immune regulators, such as TBF13, is rapidly and transiently induced upon pathogen challenge (accompanying manuscript), we developed “TBF1-cassette” consisting of not only the immune-inducible promoter but also two pathogen-responsive upstream open reading frames (uORFsTBF1) of the TBF1 gene. We demonstrate that inclusion of the uORFsTBF1-mediated translational control over the production of snc1 (an autoactivated immune receptor) in Arabidopsis (At) and AtNPR1 in rice enables us to engineer broad-spectrum disease resistance without compromising plant fitness in the laboratory or in the field. This broadly applicable new strategy may lead to reduced use of pesticides and lightening of selective pressure for resistant pathogens.

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“…Supplementary Table 1. The constructs were injected into 3-to 4-week-old N. benthamiana plants as described 39 . At 2-4 days after injection, fluorescence was observed with a Zeiss LSM510 confocal microscope (Jena, Germany).…”

Section: Methodsmentioning

confidence: 99%

ABA signalling is fine-tuned by antagonistic HAB1 variants

et al. 2015

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Group A protein type 2C phosphatases (PP2Cs) are negative regulators of abscisic acid (ABA) signalling and plant adaptation to stress. However, our knowledge of the regulation of PP2C activity is limited. Here we report that the PP2C HAB1 undergoes alternative splicing to produce two splice variants, which encode HAB1.1 and HAB1.2, that play opposing roles in ABA-mediated seed germination and ABA-mediated post-germination developmental arrest. HAB1.2 is predominately formed in the presence of ABA and prevents seed germination and post-germinative growth. HAB1.2 interacts with OST1, but cannot inhibit OST1 kinase activity; thus, it functions as a positive regulator of ABA signalling. We also identified an RNA-recognition motif-containing protein, RBM25, as a potential regulator of HAB1 alternative splicing and molecular diversity. Our results reveal a mechanism for turning ABA signalling on and off and for plant adaptation to abiotic stress.

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Development of series of gateway binary vectors, pGWBs, for realizing efficient construction of fusion genes for plant transformation

Cited by 1,528 publications

References 26 publications

Global translational reprogramming is a fundamental layer of immune regulation in plants

Global translational reprogramming is a fundamental layer of immune regulation in plants

uORF-mediated translation allows engineered plant disease resistance without fitness costs

ABA signalling is fine-tuned by antagonistic HAB1 variants

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