Localization of MT2a Gene promoter expression is different from the site(s) of copper accumulation in the roots of transgenic Arabidopsis thaliana

Accumulation of mRNA for MT2a, a metallothionein of Arabidopsis thaliana, is known to be upregulated by excess copper (Cu). We compared the localization of the promoter activity of the MT2a gene and the accumulation site(s) of CU in the roots of transgenic A. thaliana. Open reading frame of the β-glucuronidase (GUS) gene was fused with the MT2a gene promoter and introduced into A. thaliana. Strong GUS activity was observed in the region near or within the stele in the presence of CU at a high concentration (high-CU condition). Electron probe X-ray micro-analysis demonstrated that under high-CU conditions, CU accumulated most abundantly in the cortex, where GUS activity was not significantly induced. These results indicate that the sites showing the MT2a promoter activity and CU accumulation did not coincide with each other, suggesting that CU is not a direct activator of the MT2a promoter.     

Rapid and reversible nitrate inhibition of nodule growth and N2 fixation activity in soybean (Glycine max (L.) Merr.)

Nodulated soybean (Glycine max. (L) Merr. cv. Williams) plants were hydroponically cultured, and various combinations of 1-week culture with 5 or 0 mm nitrate were applied using 13-d-old soybean seedlings during three successive weeks. The treatments were designated as 0-0-0, 5-5-5, 5-5-0, 5-0-0, 5-0-5, 0-5-5, and 0-0-5, where the three sequential numbers denote the nitrate concentration (mm) applied in the first-second-third weeks. The size of the individual nodule was measured periodically using a slide caliper. All the plants were harvested after measurement of the acetylene reduction activity (ARA) at the end of the treatments. In the 0-0-0 treatment, the nodules grew continuously during the treatment period. Individual nodule growth was immediately suppressed after 5 mm nitrate supply. However, the nodule growth rapidly recovered by changing the 5 mm nitrate solution to a 0 mm nitrate solution in the 5-0-0 and 5-5-0 treatments. In the 5-0-5 treatment, nodule growth was completely inhibited in the first and the third weeks with 5 mm nitrate, but the nodule growth was enhanced in the second week with 0 mm nitrate. The nodule growth response to 5 mm nitrate was similar between small and large size nodules. After the 5-5-5, 5-0-5, 0-0-5, and 0-5-5 treatments, where the plants were cultured with 5 mm nitrate in the last third week, the ARA per plant was significantly lower compared with the 0-0-0 treatment. On the other hand, the ARA after the 5-0-0 and 5-5-0 treatments was relatively higher than that after the 0-0-0 treatment, possibly due to the higher photosynthate supply associated with the vigorous vegetative growth of the plants supplemented with nitrate nitrogen. It is concluded that both soybean nodule growth and N₂ fixation activity sensitively responded to the external nitrate level, and that these parameters were reversibly regulated by the current status of nitrate in the culture solution, possibly through sensing of the nitrate concentration in roots and / or nodules.     

Photoassimilate partitioning in hypernodulation mutant of soybean (Glycine max (L.) Merr.) NOD1-3 and its parent williams in relation to nitrate inhibition of nodule growth

Nodule growth of a hypernodulating soybean (Glycine max (L.) Merr.) mutant line NOD1-3 was compared to that of its wild-type parent cv. Williams from 14 to 18 days after planting (DAP) in the absence of nitrate treatment (hereafter referred to as “0 mM treatment”) or with 5 mM nitrate treatment. The growth rate determined by increase in the diameter of the nodules was relatively lower in the mutant NOD1-3 than that of the parent Williams under nitrogen-free conditions (0 mM nitrate). The inhibition of nodule growth by 5 mM nitrate started at 1 d after the onset of the nitrate treatment in Williams, while the inhibition did not occur before the application of the nitrate treatment for 2 d in NOD1-3. The nodule growth was completely inhibited after 2 d in Williams and after 3 d in NOD1-3 during the 5 mM nitrate treatment period. After 4 d of 5 mM nitrate treatment, the nodule dry weight decreased by 22% in NOD1-3 and by 58% in Williams, respectively. The treatment with 5 mM nitrate decreased the acetylene reduction activity (ARA) in NOD1-3 by 60% per plant and by 50% per nodule g DW and these parameters were less sensitive to the treatment than those in Williams in which the inhibition rate was 90% per plant and 80% per nodule g DW. These results indicate that NOD1-3 is partially nitrate-tolerant in terms of individual nodule growth as well as total nodule dry weight and Nz fixation activity. A whole shoot of Williams and NOD1-3 plants was exposed to ¹⁴CO₂ for 120 min followed by 0 or 5 mM nitrate treatment for 2 d, and the partitioning of the photoassimilates among the organs was analyzed. Under 0 mM nitrate treatment, the percentages of the distribution of ¹⁴C radioactivity between the nodules and roots were 63 and 37% in Williams and 89 and 11% in NOD1-3. Under the 5 mM nitrate conditions, the percentages of the distribution of ¹⁴C between the nodules and roots changed to 14 and 86% in Williams and 39 and 61% in NOD1-3, respectively. These results indicated that the hypernodulating mutant NOD1-3 supplied a larger amount of photoassimilates to the nodules than to the roots under nitrogen-free conditions, and that the nitrate depression of photoassimilate transport to the nodules was less sensitive than that of the parent line.     

Effect of placement of coated urea fertilizer on root growth and rubidium uptake activity in soybean plant

Abstract

We reported in the previous paper (Takahashi et al. 1991) that the deep placement of slow release N fertilizer (coated urea) contributed to a stable increase of soybean (Glycine max L. Merr.) yield. In the previous study we observed that the deep placement of coated urea did not depress appreciably the nitrogen fixation by root nodules although fertilizer N was efficiently utilized. We assumed that the N absorbed from the roots in the deep layers did not cause nodule senescence, contributed to the maintenance of the leaf activity during the maturation stage, and that the increase in the availability of carbohydrate and N improved seed production. In the current report the effects of placement of coated urea fertilizer on the root growth and activity were studied by measuring the root dry weight and Rb absorption activity.     

Sensitivity and specificity of a commercial BSE kit for the detection of ovine scrapie

To examine the sensitivity of a commercially available bovine spongiform encephalopathy (BSE) kit (NippIBL) for the detection of ovine scrapie, 50 scrapie‐positive ovine samples from the UK, and 54 scrapie‐negative ovine samples from Japan were obtain and tested using this kit. The sensitivity and specificity of NippIBL for ovine samples were 96% and 100%, respectively. The detection limit of the abnormal isoform of prion protein (PrP^Sc) of NippIBL was examined using diluted scrapie‐positive samples. The sensitivity of NippIBL to ovine scrapie was 3–10 times superior to that of another commercial BSE diagnosis kit. Thus, the NippIBL kit proved more effective for the detection of ovine scrapie.     

Haplotype C of growth hormone (GH) gene in Japanese Black cattle: Structure of GH protein and a novel method for detection of the gene

From a series of studies on single nucleotide polymorphisms (SNPs) in the bovine growth hormone (GH) gene of Japanese Black cattle, the type‐C (127^Val and 172^Met) that is specific for this breed has been intensively focused upon because of the economic importance for carcass traits, such as intramuscular oleic acid contents. In the present study, we intended to analyze the 3‐D structure of GH of haplotype C, and developed a novel method to detect the type C gene. Three‐D analysis of the type C protein showed that the amino acid residues (127^Val and 172^Met), which are present in the third and fourth helixes, respectively, and are important for binding with GH receptors, are shifted to deeper positions in the molecule compared with that for type A (127^Leu and 172^Thr), implying the alteration of binding interaction with receptors. A novel, efficient and cost‐effective method (Dot‐blot‐SNP technique) for type C genotyping was successfully established, of which the basal method was a reported genotyping of SNPs for a large number of plants, reducing the cost to 10% or less of direct sequencing.     

Annual reproductive cycle of the greasyback shrimp Metapenaeus ensis in Ise Bay,Japan

The greasyback shrimp Metapenaeus ensis is widely distributed along the coast of India and the West Pacific where it is an important fisheries species. We have examined seasonal changes in ovarian development, spermatogenesis, and mating of Me. ensis in histological studies and by external observations on specimens collected in Ise Bay, its northernmost habitat. Ovaries were found to be previtellogenic from February to May, with the first signs of development being the accumulation of yolk in oocytes in late May. Ovarian shadow ratios were high during the period late July to mid-September. The formation of cortical rods in the peripheries of oocytes and germinal vesicle breakdown were observed in ovaries from late June to September. Male shrimps had sperm in the testes during the period early June to early October, and female shrimps had spermatophores in spermatheca after early July. In late July, some post-spawn female shrimps had exogenous vitellogenic oocytes in their ovaries, indicating that ovarian development had been repeated in preparation for the next spawning. Ovarian shadow ratios, which were positively correlated with gonadosomatic indices and ovarian development, seem to be a useful marker to determine ovarian development. Our results indicate that mating in Me. ensis started in early July and that the spawning season ranged from July to September with more than two cycles of spawning in Ise Bay.

     

Isolation and identification of flavonoids accumulated in proanthocyanidin-free barley

Flavonoids accumulated in proanthocyanidin-free near-isogenic lines iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi, developed by backcross breeding using a leading cultivar, Nishinohoshi, as a recurrent parent and a proanthocyanidin-free mutant as a nonrecurrent parent in Japan, were examined. A new flavanone, (2RS)-dihydrotricin 7-O-β-D-glucopyranoside (1), known flavanones (2RS)-dihydrotricin (2) and (2RS)-homoeriodictyol (3), and known flavones chrysoeriol 7-O-[α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside] (4), chrysoeriol 7-O-β-D-glucopyranoside (5), tricin (6), and chrysoeriol (7) were isolated from iso ant 17 of Nishinohoshi. The structures and stereochemistries of the isolated flavonoids (1-7) were elucidated on the basis of spectroscopic analyses. The concentrations of the isolated flavonoids (1-7) in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi were similar to each other, whereas the flavonoids 1-5 and 7 were not detected in Nishinohoshi, an old Japanese cultivar, Amaginijo, and North American cultivar Harrington. The concentration of tricin (6) in Nishinohoshi was a half those in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi. Except for iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi, the concentration of tricin (6) was highest in Nishinohoshi, followed by Amaginijo and Harrington. Thus, tricin (6), its precursor dihydrotricin (2), and its glucopyranoside, dihydrotricin 7-O-β-D-glucopyranoside (1), as well as chrysoeriol (7) and homoeriodictyol (3) were accumulated in iso ant 13, iso ant 17, and iso ant 22 of Nishinohoshi probably by blocking at the step of flavanone 3-hydroxylase in the procyanidin biogenetic pathway, resulting in enhancement of the alternative biogenetic pathway.     

Isolation and Caenorhabditis elegans lifespan assay of flavonoids from onion

The main flavonoids were isolated from three selected onion cultivars. Three phenolic compounds were obtained by reverse-phase HPLC, and their structures were elucidated by multiple NMR measurements. There were two known compounds, quercetin and quercetin 3'-O-β-D-glucopyranoside (Q3'G), and one novel compound, quercetin 3-O-β-D-glucopyranoside-(4→1)-β-d-glucopyranoside (Q3M), which was identified in onion for the first time. These flavonoids were found to be more abundant in the onion peel than in the flesh or core. Their antioxidative activities were tested using the DPPH method, and their antiaging activities were evaluated using a Caenorhabditis elegans lifespan assay. No direct correlation was found between antioxidative activity and antiaging activity. Quercetin showed the highest antioxidative activity, whereas Q3M showed the strongest antiaging activity among these flavonoids, which might be related to its high hydrophilicity.     

Ultralong-range polaron-induced quenching of excitons in isolated conjugated polymers

In conjugated polymers, radiative recombination of excitons (electron-hole pairs) competes with nonradiative thermal relaxation pathways. We visualized exciton quenching induced by hole polarons in single-polymer chains in a device geometry. The distance-scale for quenching was measured by means of a new subdiffraction, single-molecule technique--bias-modulated intensity centroid spectroscopy--which allowed the extraction of a mean centroid shift of 14 nanometers for highly ordered, single-polymer nanodomains. This shift requires energy transfer over distances an order of magnitude greater than previously reported for bulk conjugated polymers and far greater than predicted by the standard mechanism for exciton quenching, the unbiased diffusion of free excitons to quenching sites. Instead, multistep "energy funneling" to trapped, localized polarons is the probable mechanism for polaron-induced exciton quenching.