High β-carotene accumulation in transgenic eggplant fruits grown under artificial light.

Eggplant ( L.) fruits are known to contain few carotenoids such as β-carotene, which are abundant in congener tomato fruits. In a previous study, we introduced a fruit-specific gene promoter-driven gene encoding phytoene synthase (PSY) of into eggplant 'Senryo No.

IRE1-mediated cytoplasmic splicing and regulated IRE1-dependent decay of mRNA in the liverwort .

The unfolded protein response (UPR) or the endoplasmic reticulum (ER) stress response is a homeostatic cellular response conserved in eukaryotes to alleviate the accumulation of unfolded proteins in the ER. In the present study, we characterized the UPR in the liverwort to obtain insights into the conservation and divergence of the UPR in the land plants. We demonstrate that the most conserved UPR transducer in eukaryotes, IRE1, is conserved in , which harbors a single gene encoding IRE1.

Unpaired nucleotides on the stem of microRNA precursor are important for precise cleavage by Dicer-like 1 in Arabidopsis.

Dicer-like 1 (DCL1) is a core component of the plant microRNA (miRNA) biogenesis machinery. MiRNA is transcribed as a precursor RNA, termed primary miRNA (pri-miRNA), which is cleaved by DCL1 in two steps to generate miRNA/miRNA* duplex. Pri-miRNA is a single-stranded RNA that forms a hairpin structure with a number of unpaired bases, hereafter called mismatches, on its stem.

Genetic engineering of eggplant accumulating β-carotene in fruit.

Genetic engineering of eggplant using fruit-specific EEF48 promoter-driven bacterial PSY gene, crtB, confers β-carotene accumulation in fruit. Eggplant (Solanum melongena L.) is globally cultivated especially in Asia and is an important source of nutrients in the diets of low-income consumers in developing countries.

Unfolded protein-independent IRE1 activation contributes to multifaceted developmental processes in Arabidopsis.

In Arabidopsis, the and double mutant () is unable to activate cytoplasmic splicing of mRNA and regulated IRE1-dependent decay under ER stress, whereas the mutant does not exhibit severe developmental defects under normal conditions. In this study, we focused on the Arabidopsis gene, whose product lacks a sensor domain. We found that the triple mutant is lethal, and heterozygous (/+) mutation in the mutants resulted in growth defects and reduction of the number of pollen grains.

Deficiency in the double-stranded RNA binding protein HYPONASTIC LEAVES1 increases sensitivity to the endoplasmic reticulum stress inducer tunicamycin in Arabidopsis.

Objective: microRNA (miRNA) is a small non-coding RNA that regulates gene expression by sequence-dependent binding to protein-coding mRNA in eukaryotic cells. In plants, miRNA plays important roles in a plethora of physiological processes, including abiotic and biotic stress responses. The present study was conducted to investigate whether miRNA-mediated regulation is important for the endoplasmic reticulum (ER) stress response in Arabidopsis.

CRISPR/Cas9-mediated homologous recombination in tobacco.

Co-transformation of multiple T-DNA in a binary vector enabled CRISPR/Cas9-mediated HR in tobacco. HR occurred in a limited region around the gRNA target site. In this study, CRISPR/Cas9-mediated homologous recombination (HR) in tobacco (Nicotiana tabacum L.

Constitutive BiP protein accumulation in Arabidopsis mutants defective in a gene encoding chloroplast-resident stearoyl-acyl carrier protein desaturase.

The unfolded protein response (UPR) occurs when protein folding and maturation are disturbed in the endoplasmic reticulum (ER). During the UPR, a number of genes including those encoding ER-resident molecular chaperones are induced. In Arabidopsis, BiP3 has been used as a UPR marker gene whose expression is strongly induced in response to ER stress.

DNA double-strand breaks promote endoreduplication in radish cotyledon.

DSBs differently affect endoreduplication and organ size in radish cotyledons and hypocotyls in different light conditions, suggesting that DSBs-mediated endoreduplication varies based on different developmental and environmental cues. Endoreduplication induced by DNA double strand breaks (DSBs) in Arabidopsis thaliana roots and cultured cells has been reported in recent years. In this study, we investigated whether DSBs-mediated endoreduplication also occurs in other tissues, such as cotyledons and hypocotyls of radish (Raphanus sativus var.

A 64-bp sequence containing the GAAGA motif is essential for CaMV-35S promoter methylation in gentian.

This study investigated sequence specificity and perenniality of DNA methylation in the cauliflower mosaic virus (CaMV) 35S promoter of transgenic gentian (Gentiana triflora×G. scabra) plants. Unlike conventional transgene silencing models, 35S promoter hypermethylation in gentian is species-specific and occurs irrespective of the T-DNA copy number and genomic location.

Activation of the Arabidopsis membrane-bound transcription factor bZIP28 is mediated by site-2 protease, but not site-1 protease.

The unfolded protein response (UPR) is a homeostatic cellular response conserved in eukaryotic cells to alleviate the accumulation of unfolded proteins in the endoplasmic reticulum (ER). Arabidopsis bZIP28 is a membrane-bound transcription factor activated by proteolytic cleavage in response to ER stress, thereby releasing its cytosolic portion containing the bZIP domain from the membrane to translocate into the nucleus where it induces the transcription of genes encoding ER-resident molecular chaperones and folding enzymes. It has been widely recognized that the proteolytic activation of bZIP28 is mediated by the sequential cleavage of site-1 protease (S1P) and site-2 protease (S2P).

Inositol-requiring enzyme 1 affects meristematic division in roots under moderate salt stress in Arabidopsis.

The unfolded protein response (UPR) mitigates stress caused by accumulation of unfolded proteins in the endoplasmic reticulum (ER). Inositol-requiring enzyme 1 (IRE1) is the most conserved sensor of the UPR with ribonuclease activity that mediates cytoplasmic splicing and decay of mRNA encoding secretory and membrane proteins. In the present study, we demonstrate that the Arabidopsis mutant defective in two genes exhibit retarded growth of primary roots under moderate salt stress, although such grow retardation is not observed in wild type plants.

Effect of light and auxin transport inhibitors on endoreduplication in hypocotyl and cotyledon.

Enhancement of endoreduplication in dark-grown hypocotyl is a common feature in dicotyledonous polysomatic plants, and TIBA-mediated inhibition of the endoreduplication is partially due to abnormal actin organization. Many higher plant species use endoreduplication during cell differentiation. However, the mechanisms underlying this process have remained elusive.

Tunicamycin-induced inhibition of protein secretion into culture medium of Arabidopsis T87 suspension cells through mRNA degradation on the endoplasmic reticulum.

The N-glycosylation inhibitor tunicamycin triggers endoplasmic reticulum stress response and inhibits efficient protein secretion in eukaryotes. Using Arabidopsis suspension cells, we showed that the reduced secretion of mannose-binding lectin 1 (MBL1) protein by tunicamycin is accompanied by a significant decrease in MBL1 mRNA, suggesting that mRNA destabilization is the major cause of the inhibition of protein secretion in plants.

Arabidopsis tRNA ligase completes the cytoplasmic splicing of bZIP60 mRNA in the unfolded protein response.

Arabidopsis bZIP60 is a major transcription factor that activates the unfolded protein response and is regulated by cytoplasmic splicing. Two Arabidopsis inositol-requiring 1s (IRE1A and IRE1B) cleave bZIP60 mRNA; however, the ligase that connects the two half-molecules of the split bZIP60 mRNA has not yet been identified. We aimed to determine whether the Arabidopsis tRNA ligase RLG1 catalyzes the ligation of cleaved bZIP60 mRNA.

CaMV-35S promoter sequence-specific DNA methylation in lettuce.

We found 35S promoter sequence-specific DNA methylation in lettuce. Additionally, transgenic lettuce plants having a modified 35S promoter lost methylation, suggesting the modified sequence is subjected to the methylation machinery. We previously reported that cauliflower mosaic virus 35S promoter-specific DNA methylation in transgenic gentian (Gentiana triflora × G.

Exogenous salicylic acid activates two signaling arms of the unfolded protein response in Arabidopsis.

The unfolded protein response (UPR) is a highly conserved cellular response that prevents abnormal maturation of proteins in the endoplasmic reticulum (ER). The expression of genes encoding ER chaperones is induced during the UPR. In the Arabidopsis UPR, two membrane-bound transcription factors, bZIP60 and bZIP28, activate the expression of those genes.

The polar auxin transport inhibitor TIBA inhibits endoreduplication in dark grown spinach hypocotyls.

We addressed the question of whether an additional round of endoreduplication in dark-grown hypocotyls is a common feature in dicotyledonous plants having endopolyploid tissues. Ploidy distributions of hypocotyl tissues derived from in vitro-grown spinach (Spinacia oleracea L. cv.

An attempt to detect siRNA-mediated genomic DNA modification by artificially induced mismatch siRNA in Arabidopsis.

Although tremendous progress has been made in recent years in identifying molecular mechanisms of small interfering RNA (siRNA) functions in higher plants, the possibility of direct interaction between genomic DNA and siRNA remains an enigma. Such an interaction was proposed in the 'RNA cache' hypothesis, in which a mutant allele is restored based on template-directed gene conversion. To test this hypothesis, we generated transgenic Arabidopsis thaliana plants conditionally expressing a hairpin dsRNA construct of a mutated acetolactate synthase (mALS) gene coding sequence, which confers chlorsulfuron resistance, in the presence of dexamethasone (DEX).

Defects in IRE1 enhance cell death and fail to degrade mRNAs encoding secretory pathway proteins in the Arabidopsis unfolded protein response.

The unfolded protein response (UPR) is a cellular response highly conserved in eukaryotes to obviate accumulation of misfolded proteins in the endoplasmic reticulum (ER). Inositol-requiring enzyme 1 (IRE1) catalyzes the cytoplasmic splicing of mRNA encoding bZIP transcription factors to activate the UPR signaling pathway. Arabidopsis IRE1 was recently shown to be involved in the cytoplasmic splicing of bZIP60 mRNA.

Arabidopsis IRE1 catalyses unconventional splicing of bZIP60 mRNA to produce the active transcription factor.

IRE1 plays an essential role in the endoplasmic reticulum (ER) stress response in yeast and mammals. We found that a double mutant of Arabidopsis IRE1A and IRE1B (ire1a/ire1b) is more sensitive to the ER stress inducer tunicamycin than the wild-type. Transcriptome analysis revealed that genes whose induction was reduced in ire1a/ire1b largely overlapped those in the bzip60 mutant.

Epigenetic modifications of the 35S promoter in cultured gentian cells.

Our previous studies found strict gene silencing associated with CaMV-35S promoter-specific de novo methylation in transgenic gentian plants. To dissect the de novo methylation machinery, especially in association with histone modification, 35S-driven sGFP-expressing and -silenced gentian cultured cell lines that originated from a single transformation event were produced and used for epigenetic analyses. A sGFP-expressing primarily induced cell suspension culture (PS) was hypomethylated in the 35S promoter region, although a low level of de novo methylation at the 35S enhancer region (-148 to -85) was detected.

Strict de novo methylation of the 35S enhancer sequence in gentian.

A novel transgene silencing phenomenon was found in the ornamental plant, gentian (Gentiana triflora x G. scabra), in which the introduced Cauliflower mosaic virus (CaMV) 35S promoter region was strictly methylated, irrespective of the transgene copy number and integrated loci. Transgenic tobacco having the same vector did not show the silencing behavior.

Genetic engineering of novel flower colour by suppression of anthocyanin modification genes in gentian.

Ornamental gentian plants have vivid-blue flowers. The main factor contributing to the flower colour is the accumulation of a polyacylated delphinidin 'gentiodelphin' in their petals. Although in vitro studies proposed that acylation plays an important role in the stability and development of gentian blue colour, the in vivo stability of the polyacylated anthocyanin was not clearly demonstrated.

Agrobacterium-mediated transformation of protocorm-like bodies in Cymbidium.

Genetically transformed plants of Cymbidium were regenerated after cocultivating protocorm-like bodies (PLB) with Agrobacterium tumefaciens strain EHA101 (pIG121Hm) that harbored genes for beta-glucuronidase (gus), hygromycin phosphotransferase (hpt) and neomycin phosphotransferase II (nptII). PLB of three genotypes maintained in liquid new Dogashima medium (NDM), were subjected to transformation experiments. The PLB inoculated with Agrobacterium produced secondary PLB, 4 weeks after transfer onto 2.