Acylated anthocyanins from red radish (Raphanus sativus L.)

Twelve acylated anthocyanins were isolated from the red radish (Raphanus sativus L.) and their structures were determined by spectroscopic analyses. Six of these were identified as pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-beta-D-glucopyranosyl]-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-caffeoyl-2-O-(6-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-p-coumaroyl-2-O-(6-(E)-caffeoyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-(6-(E)-caffeoyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-p-coumaroyl-2-O-(6-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), and pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-(2-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside).     

The Structure and Organization of the Human NPAT Gene

Ataxia telangiectasia (AT) is an autosomal recessive gene disorder, andATM,a housekeeping gene, has been identified as the gene responsible for AT. Recently we found that another housekeeping gene,NPAT,is located upstream ofATMon human chromosome 11. The two housekeeping genes are transcribed in opposite directions and share a 0.5-kb 5′ flanking sequence. The structure and organization ofNPATwere determined by direct sequencing of cosmid clones carrying the gene and by application of the long and accurate (LA)-PCR method to amplify regions encompassing the exon/intron boundaries and all of the exons. The gene spans at least 44 kb and consists of 18 exons and 17 introns. It has been suggested that AT heterozygotes have an increased risk of developing cancer, especially breast cancer in women. Frequently, loss of heterozygosity at loci on 11q22–q24 has been observed in DNA isolated from tumors of the breast, uterine cervix, and colon, perhaps suggesting the location of a tumor suppressor gene in 11q22–q24. For investigation of the role ofNPATin AT and these tumors with allelic loss of 11q22–q24, appropriate primer sequences and PCR conditions for amplification of all theNPATexons from genomic DNA were determined. We previously reported that no recombinations are found amongAtm, Npat,andAcat1(acetoacetyl-CoA thiolase) loci as determined by fine genetic linkage mapping of the mouse AT region. The results of the LA-PCR analysis usingNPAT- andACAT-specific primers and human genomic DNA allowed us to mapACAT12 kb centromeric toNPAT.     

NSrp70 is a novel nuclear speckle-related protein that modulates alternative pre-mRNA splicing in vivo

Nuclear speckles are known to be the storage sites of mRNA splicing regulators. We report here the identification and characterization of a novel speckle protein, referred to as NSrp70, based on its subcellular localization and apparent molecular weight. This protein was first identified as CCDC55 by the National Institutes of Health Mammalian Gene Collection, although its function has not been assigned. NSrp70 was colocalized and physically interacted with SC35 and ASF/SF2 in speckles. NSrp70 has a putative RNA recognition motif, the RS-like region, and two coiled-coil domains, suggesting a role in RNA processing. Accordingly, using CD44, Tra2β1 and Fas constructs as splicing reporter minigenes, we found that NSrp70 modulated alternative splice site selection in vivo. The C-terminal 10 amino acids (531-540), including (536)RD(537), were identified as a novel nuclear localization signal, and the region spanning 290-471 amino acids was critical for speckle localization and binding to SC35 and ASF/SF2. The N-terminal region (107-161) was essential for the pre-mRNA splicing activity. Finally, we found that knockout of NSrp70 gene in mice led to a lack of progeny, including fetal embryos. Collectively, we demonstrate that NSrp70 is a novel splicing regulator and essentially required early stage of embryonic development.     

Specific detection of wheat residues in processed foods by polymerase chain reaction

A sensitive qualitative detection method for wheat in foods using polymerase chain reaction (PCR) was developed. Trace amounts of wheat in commercial food products could be qualitatively detected by this method. The sensitivity of the proposed PCR method appears to be similar to that of ELISA. The present method should be very useful for detecting wheat residues in processed foods.     

Cultivation of mushrooms of edible ectomycorrhizal fungi associated with Pinus densiflora by in vitro mycorrhizal synthesis

Edible mushroom fungi in the genera Lyophyllum, Tricholoma, Leucopaxillus, Suillus, Rhizopogon, Lactarius, and Morchella were tested for mycorrhization with Pinus densiflora in vitro. Most of the tested fungi in the genera Lyophyllum, Tricholoma, Suillus, Rhizopogon, and Lactarius formed ectomycorrhizas 2–4 months after fungal inoculation. Mycorrhizal seedlings were then acclimatized in open-pot soil under growth-chamber conditions. Almost all mycorrhizal seedlings sustained their symbiont and developed new mycorrhizas for 8–9 months after transplantation. Under these conditions, more than half of the tested species formed primordia and Tricholoma flavovirens, Rhizopogon rubescens, and Lactarius akahatsu developed basidiocarps with young host plants. Accepted: 28 November 2000     

Phosphorylation-Dependent Ubiquitination of Paraxial Protocadherin (PAPC) Controls Gastrulation Cell Movements

Paraxial protocadherin (PAPC) has been shown to be involved in gastrulation cell movements during early embryogenesis. It is first expressed in the dorsal marginal zone at the early gastrula stage and subsequently restricted to the paraxial mesoderm in Xenopus and zebrafish. Using Xenopus embryos, we found that PAPC is also regulated at the protein level and is degraded and excluded from the plasma membrane in the axial mesoderm by the late gastrula stage. Regulation of PAPC requires poly-ubiquitination that is dependent on phosphorylation. PAPC is phosphorylated by GKS3 in the evolutionarily conserved cytoplasmic domain, and this in turn is necessary for poly-ubiquitination by an E3 ubiquitin ligase β-TrCP. We also show that precise control of PAPC by phosphorylation/ubiquitination is essential for normal Xenopus gastrulation cell movements. Taken together, our findings unveil a novel mechanism of regulation of a cell adhesion protein and show that this system plays a crucial role in vertebrate embryogenesis.     

Chromosome mapping of the human cytidine-5′-triphosphate synthetase (CTPS) gene to band 1p34.1–p34.3 by fluorescence in situ hybridization

Summary The human cytidine-5-triphosphate synthetase (CTPS) gene was mapped by a direct mapping system combined with fluorescence in situ hybridization and replicated prometaphase R-bands. By high-resolution banding analysis, the signals were localized to band 34.1–34.3 of the short arm of chromosome 1; 1p34.1–p34.3. Simple procedures for the detection of R-bands are described.