The in vitro relationship between the human p53 DNA binding domain (p53 DBD) and glycolipids was investigated. We isolated the glycolipid fraction from spinach (Spinacia oleracea L.) and found that the fraction inhibited the double-stranded DNA (dsDNA) binding activity of p53 DBD. Since the fraction contained mainly three glycolipids, monogalactosyl diacylglycerol (MGDG), digalactosyl diacylglycerol (DGDG) and sulfoquinovosyl diacylglycerol (SQDG), and each glycolipid was purified using silica gel column chromatography. Purified SQDG inhibited the activity, however, purified MGDG and DGDG had no influence. In this study, we demonstrated the structure-function relationship between chemically synthetic SQDG and p53 DBD. The major action is probably dependent on the fatty acid effect, although SQDG was a much stronger inhibitor than the fatty acid alone present in SQDG. The inhibitory activity of SQDG was weakened by the R248A mutant of p53 DBD, suggesting that R248 in the dsDNA binding site of p53 must be important for the inhibitory activity of SQDG. SQDG binding to p53 DBD could be reversed with a non-ionic detergent, Nonidet P-40. This is the first study of a glycolipid, SQDG, acting as a dsDNA binding inhibitor of p53, and it could be considered that a SQDG-containing thylakoid membrane in plant chloroplasts might regulate the activity of p53 for cell division, cell cycle checkpoint and tumor suppression.
Applying our photo technology, we succeeded in generating silver particles exceeding 10 μm by using 50 nm gold particles as catalysts, which are commonly used in commercially available influenza diagnostic kits. The amplified silver particles are 1,000 times easier to see than gold nano-particles. We have commercialized the influenza detection kit, named “FUJI DRI-CHEM IMMUNO AG cartridge FluAB”, and the densitometry analysis machine to determine automatically positive or negative for influenza infection, named “FUJI DRI-CHEM IMMUNO AG1”, using this amplification technology.
Hollow spherical particles with protein-silica hybrid shell structures have been synthesized through a combination of the catalytic activity of the protein and sonochemical treatment; the morphologies of the particles were controlled by varying the protein concentration.
The basidiomycete Coprinus cinereus has many advantages as a model organism for studying sexual development and meiosis, but it has been difficult to investigate using reverse-genetics methods, such as gene disruption by homologous recombination. Here, gene repression by dsRNA-mediated gene silencing was tried as an alternative method for reverse-genetics studies. It was shown that transformation of the LIM15/DMC1 dsRNA expression construct ( LIM15 dsRNA) resulted in genomic insertion of LIM15 dsRNA and paucity of the LIM15/DMC1 transcript. First, LIM15 dsRNA was transformed into the homothallic strain AmutBmut to generate a homozygote in which both nuclei had a copy of LIM15 dsRNA. The LIM15/DMC1 -repressed strain showed abnormal homologous chromosome synapsis during meiosis. Basidiospore production was reduced to 16 % by the induction of dsRNA. However, approximately 60 % of basidiospores were viable. Next, a heterozygote was generated in which one nucleus had a copy of LIM15 dsRNA. The phenotype was similar to that of the homozygote. These results are not only the first demonstration of dsRNA-mediated gene silencing in a member of the homobasidiomycete fungi, to which 90 % of mushroom species belong, but also the first successful use of a reverse-genetics approach in C. cinereus research.
