Abstract:These authors contributed equally to this work.
SUMMARYIn Arabidopsis, anthocyanin biosynthesis is controlled by a MYB-bHLH-WD40 (MBW) transcriptional activator complex. The MBW complex activates the transcription of late biosynthesis genes in the flavonoid pathway, leading to the production of anthocyanins. A similar MBW complex regulates epidermal cell fate by activating the transcription of GLABRA2 (GL2), a homeodomain transcription factor required for trichome formation in shoots and non-hair cell formatio… Show more
“…Protoplast isolation, transfection, and GUS activity assay were performed as described previously (Tiwari et al, 2003;Wang et al, 2005Wang et al, , 2007aWang et al, , 2007bWang et al, , 2015Tian et al, 2015;Dai et al,2016;Zheng et al,2016). In brief, protoplasts were isolated from rosette leaves of 3-to 4-week-old soilgrown Col wild-type plants, cotransfected with effector and reporter plasmids, incubated under darkness at room temperature for 20 to 22 h before GUS activities were measured by using a Synergy HT microplate reader (BioTEK).…”
Section: Plant Transformation and Transgenic Plant Selectionmentioning
The NAM, ATAF1/2, and CUC (NAC) are plant-specific transcription factors that regulate multiple aspects of plant growth and development and plant response to environmental stimuli. We report here the identification of NTM1-LIKE8 (NTL8), a membrane-associated NAC transcription factor, as a novel regulator of trichome formation in Arabidopsis (Arabidopsis thaliana). From an activation-tagged Arabidopsis population, we identified a dominant, gain-of-function mutant with glabrous inflorescence stem. By using plasmid rescue and RT-PCR analyses, we found that NTL8 was tagged; thus, the mutant was named ntl8-1 Dominant (ntl8-1D). Recapitulation experiment further confirmed that the phenotype observed in the ntl8-1D mutant was caused by elevated expression of NTL8. Quantitative RT-PCR results showed that the expression level of the single-repeat R3 MYB genes TRIPTYCHON (TRY) and TRICHOMELESS1 (TCL1) was elevated in the ntl8-1D mutant. Genetic analyses demonstrated that NTL8 acts upstream of TRY and TCL1 in the regulation of trichome formation. When recruited to the promoter region of the reporter gene Gal4:GUS by a fused GAL4 DNA-binding domain, NTL8 activated the expression of the reporter gene. Chromatin immunoprecipitation results indicated that TRY and TCL1 are direct targets of NTL8. However, NTL8 did not interact with SQUAMOSA PROMOTER BINDING PROTEIN LIKE9, another transcription factor that regulates the expression of TRY and TCL1, in yeast and plant cells. Taken together, our results suggest that NTL8 negatively regulates trichome formation in Arabidopsis by directly activating the expression of TRY and TCL1.
“…Protoplast isolation, transfection, and GUS activity assay were performed as described previously (Tiwari et al, 2003;Wang et al, 2005Wang et al, , 2007aWang et al, , 2007bWang et al, , 2015Tian et al, 2015;Dai et al,2016;Zheng et al,2016). In brief, protoplasts were isolated from rosette leaves of 3-to 4-week-old soilgrown Col wild-type plants, cotransfected with effector and reporter plasmids, incubated under darkness at room temperature for 20 to 22 h before GUS activities were measured by using a Synergy HT microplate reader (BioTEK).…”
Section: Plant Transformation and Transgenic Plant Selectionmentioning
The NAM, ATAF1/2, and CUC (NAC) are plant-specific transcription factors that regulate multiple aspects of plant growth and development and plant response to environmental stimuli. We report here the identification of NTM1-LIKE8 (NTL8), a membrane-associated NAC transcription factor, as a novel regulator of trichome formation in Arabidopsis (Arabidopsis thaliana). From an activation-tagged Arabidopsis population, we identified a dominant, gain-of-function mutant with glabrous inflorescence stem. By using plasmid rescue and RT-PCR analyses, we found that NTL8 was tagged; thus, the mutant was named ntl8-1 Dominant (ntl8-1D). Recapitulation experiment further confirmed that the phenotype observed in the ntl8-1D mutant was caused by elevated expression of NTL8. Quantitative RT-PCR results showed that the expression level of the single-repeat R3 MYB genes TRIPTYCHON (TRY) and TRICHOMELESS1 (TCL1) was elevated in the ntl8-1D mutant. Genetic analyses demonstrated that NTL8 acts upstream of TRY and TCL1 in the regulation of trichome formation. When recruited to the promoter region of the reporter gene Gal4:GUS by a fused GAL4 DNA-binding domain, NTL8 activated the expression of the reporter gene. Chromatin immunoprecipitation results indicated that TRY and TCL1 are direct targets of NTL8. However, NTL8 did not interact with SQUAMOSA PROMOTER BINDING PROTEIN LIKE9, another transcription factor that regulates the expression of TRY and TCL1, in yeast and plant cells. Taken together, our results suggest that NTL8 negatively regulates trichome formation in Arabidopsis by directly activating the expression of TRY and TCL1.
“…8b2, 4). Because of the negative relationship between mucilage abundance on the seed coat surface and TAGs in the seed [24–26], we believe that the role of T-6b in reducing the mucilage abundance is beneficial to increase seed oil content.Fig. 8Effects of T-6b expressed in the endosperm on seed mucilage.…”
Section: Resultsmentioning
confidence: 99%
“…In Arabidopsis seeds, TAGs are mainly stored in embryo, but the endosperm also accumulates 10–15% of seed oil [22, 23]. The proanthocyanidin and mucilage contents, which are positively controlled by GLABRA2, are negatively correlated with the amount of TAGs in the seed [24–26]. Plant TAGs are important nutrient sources for humans.…”
BackgroundAs an Agrobacterium tumefaciens T-DNA oncogene, T-6b induces the development of tumors and the enation syndrome in vegetative tissues of transgenic plants. Most of these effects are related to increases in soluble sugar contents. To verify the potential roles of T-6b in the distribution of carbon in developing seeds, not in vegetative tissues, we fused an endosperm-specific promoter to the T-6b gene for expression in transgenic Arabidopsis thaliana plants.ResultsThe expression of T-6b in reproductive organs did not induce the development of the enation syndrome, and moreover, promoted endosperm expansion, which increased the total seed biomass by more than 10%. Additionally, T-6b also increased oil content in mature seeds by more than 10% accompanied with the decrease of starch and mucilage content at the same time.ConclusionsT-6b enhances seed biomass and helps oil biosynthesis but not polysaccharides in reproductive organs without disturbing vegetative growth and development. Our findings suggest T-6b may be very useful for increasing oil production in biodiesel plants.
“…The reporter constructs LexA-Gal4:GUS and Gal4:GUS, and the effector constructs LD-VP, GD, GD-GL3 and GD-OFP1-N used for protoplast transient transfection assays have been described previously (Tiwari et al 2004;Wang et al 2007Wang et al , 2014Wang X et al 2015).…”
Section: Plasmid Constructionmentioning
confidence: 99%
“…The procedures for Arabidopsis leaf mesophyll protoplast isolation, transfection and GUS activity assays have been described previously (Tiwari et al 2003;Wang et al 2005;Tian et al 2015;Wang X et al 2015;Dai et al 2016). Briefly, plasmid DNA of the reporters and effectors were isolated using the Gold Hi Endo Free Plasmid Maxi Kit (CWBIO) by following the manufacturer's procedure, and co-transfected into protoplasts isolated from rosette leaves of ∼4-week-old Col wild-type Arabidopsis plants.…”
Section: Protoplast Isolation Transfection and Gus Activity Assaymentioning
Fungi interact with plants in different ways in nature; however, it remains largely unclear if fungus gene may affect plant growth and development. Basic helix-loop-helix (bHLH) transcription factors are found in almost all organisms including plants and fungi. By using bioinformatics analysis, we found that there are 12 genes in Pleurotus ostreatus which encode bHLH transcription factors, namely P. ostreatus basic helix-loop-helix1 through 12 (PobHLH1-PobHLH12). By taking PobHLH1, 2, 5 and 8 as examples, we examined whether PobHLHs may regulate gene expression and plant growth and development when expressed in Arabidopsis. We found that all the four PobHLHs examined were localized in the nucleus; however, PobHLH2, 5 and 8 activated whereas PobHLH1 repressed reporter gene expression in transfected protoplasts. On the other hand, ectopic expression of PobHLH5 or 8 inhibited plant growth and development, whereas transgenic plants expressing PobHLH1 and PobHLH2 were largely morphologically similar to the wild-type plants.ARTICLE HISTORY
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