Virus-Mediated Targeted DNA Methylation Illuminates the Dynamics of Methylation in an Endogenous Plant Gene.

DNA methylation maintains genome stability and regulates gene expression in plants. RNA-directed DNA methylation (RdDM) is critical for appropriate methylation. However, no efficient tools are available for the investigation of the functions of specific DNA methylation.

Role of salicylic acid glucosyltransferase in balancing growth and defence for optimum plant fitness.

Salicylic acid (SA), an essential secondary messenger for plant defence responses, plays a role in maintaining a balance (trade-off) between plant growth and resistance induction, but the detailed mechanism has not been explored. Because the SA mimic benzothiadiazole (BTH) is a more stable inducer of plant defence than SA after exogenous application, we analysed expression profiles of defence genes after BTH treatment to better understand SA-mediated immune induction. Transcript levels of the salicylic acid glucosyltransferase (SAGT) gene were significantly lower in BTH-treated Nicotiana tabacum (Nt) plants than in SA-treated Nt control plants, suggesting that SAGT may play an important role in SA-related host defence responses.

Rapid transient protein production by the coat protein-deficient cucumber mosaic virus vector: non-packaged CMV system, NoPaCS.

We developed a non-packaged CMV system (NoPaCS) for CMV-agroinfection with a virus-inescapable transgenic plant platform, enabling rapid, high production of a large-sequence target protein. For rapidly producing high levels of a desirable protein, many plant virus vectors have been developed. However, there is always a concern that such recombinant viruses may escape into the environment.

A simple agroinfiltration method for transient gene expression in plant leaf discs.

In the present study, we developed a simple transient gene expression system based on Agrobacterium-mediated transformation. Vacuum infiltration was applied to leaf discs from Nicotiana benthamiana plants with Agrobacterium suspension solution under conventional vacuum conditions in a needleless plastic syringe. Model proteins, green fluorescent protein, β-glucuronidase, mouse granulocyte-macrophage colony-stimulating factor, and human fibroblast growth factor 1 were successfully expressed in leaf discs within 4 days after infiltration.

NMR-based structural validation of therapeutic antibody produced in Nicotiana benthamiana.

We successfully developed a method for metabolic isotope labeling of recombinant proteins produced in transgenic tobacco. This enabled assessment of structural integrity of plant-derived therapeutic antibodies by NMR analysis. A variety of expression vehicles have been developed for the production of promising biologics, including plants, fungi, bacteria, insects, and mammals.

Production of biologically active Atlantic salmon interferon in transgenic potato and rice plants.

Interferons (IFNs) are cytokines that induce an antiviral state in vertebrate cells. The Atlantic salmon (Salmo salar) IFN gene (SasaIFN-alpha1) was introduced in potato and rice plants by Agrobacterium-mediated transformation to produce a biologically active fish IFN in these plants. The transgenes and their transcripts were detected by PCR and Northern blot analysis.

Risk-managed production of bioactive recombinant proteins using a novel plant virus vector with a helper plant to complement viral systemic movement.

A plant viral vector has the potential to efficiently produce recombinant proteins at a low cost in a short period. Although recombinant proteins can be also produced by transgenic plants, a plant viral vector, if available, may be more convenient when urgent scale-up in production is needed. However, it is difficult to use a viral vector in open fields because of the risk of escape to the environment.

HC-Pro, a potyvirus RNA silencing suppressor, cancels cycling of Cucumber mosaic virus in Nicotiana benthamiana plants.

The mixed infection of Cucumber mosaic virus (CMV) and a potyvirus has been known to increase CMV titer in Nicotiana benthamiana plants, resulting in synergistic viral symptoms. We found that among three potyviruses--Potato virus Y (PVY), Turnip mosaic virus (TuMV), and Clover yellow vein virus (C1YVV)--synergistic effects on CMV (or a recombinant CMV vector) titers were most efficiently induced by a co-infection with PVY in N. benthamiana plants.

Comparative contribution of the aryl hydrocarbon receptor gene to perinatal stage development and dioxin-induced toxicity between the urogenital complex and testis in the mouse.

TCDD (2,3,7,8-tetrachlorodebenzo-p-dioxin) requires the presence of the aryl hydrocarbon receptor (Ahr) gene for its toxic effects, such as reproductive disorders in male offspring of maternally exposed rats and mice. To study the involvement of the Ahr gene in producing the toxic phenotype with respect to testicular development, we administered a relatively high dose of TCDD to mice with three different maternally derived Ahr genotypic traits, and then compared several Ahr-dependent alterations among male reproductive systems on Postnatal Day 14. Reduction in anogenital distance and expression of prostatic epithelial genes in the urogenital complex (UGC) were detected in Ahr(+/+) and Ahr(+/-) mice exposed to TCDD, whereas no difference was observed in Ahr(-/-) mice.

Testicular cytochrome P450scc and LHR as possible targets of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the mouse.

Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in adult animals has been reported to perturb the regulation of steroidogenesis in the testis, possibly by arylhydrocarbon receptor (AhR). To clarify how AhR is involved in the testicular steroidogenesis, we carried out comparative experiments using wild-type and AhR-null male mice that were intraperitoneally administered TCDD. The TCDD administration to wild-type mice showed significant reduction of P450scc and LHR in the testis, whereas the levels in the AhR-null mouse testis were unchanged.

Administration of estradiol-3-benzoate down-regulates the expression of testicular steroidogenic enzyme genes for testosterone production in the adult rat.

ABSTRACT. To study the role of estrogen in the testes, testosterone and testicular steroidogenic enzyme mRNA levels were investigated in male Sprague-Dawley rats 24 hr after intramuscular administration of a single dose of estradiol-3-benzoate (EB). EB administration resulted in a greater decrease in intra-testicular and serum testosterone in 10-week-old rats than in 3- or 5-week-old rats.