Efficient production of genetically engineered,male-sterile <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> using anther-specific promoters and genes derived from <Emphasis Type="Italic">Brassica oleracea</Emphasis> and <Emphasis Type="Italic">B. rapa</Emphasis>

Prevention of transgene flow from genetically modified crops to food crops and wild relatives is of concern in agricultural biotechnology. We used genes derived from food crops to produce complete male sterility as a strategy for gene confinement as well as to reduce the food purity concerns of consumers. Anther-specific promoters (A3, A6, A9, MS2, and MS5) were isolated from Brassica oleracea and B. rapa and fused to the β-glucuronidase (GUS) reporter gene and candidate genes for male sterility, including the cysteine proteases BoCysP1 and BoCP3, and negative regulatory components of phytohormonal responses involved in male development. These constructs were then introduced into Arabidopsis thaliana. GUS analyses revealed that A3, A6, and A9 had tapetum-specific promoter activity from the anther meiocyte stage. Male sterility was confirmed in tested constructs with protease or gibberellin insensitive (gai) genes. In particular, constructs with BoCysP1 driven by the A3 or A9 promoter most efficiently produced plants with complete male sterility. The tapetum and middle layer cells of anthers expressing BoCysP1 were swollen and excessively vacuolated when observed in transverse section. This suggests that the ectopic expression of cysteine protease in the meiocyte stage may inhibit programmed cell death. The gai gene also induced male sterility, although at a low frequency. This is the first report to show that plant cysteine proteases and gai from food crops are available as a novel tool for the development of genetically engineered male-sterile plants. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Colocalization of NO and VIP in neurons of the submucous plexus in the rat intestine

Since very few previous studies have carried out the quantitative analysis for the colocalization of nitric oxide (NO) and vasoactive intestinal peptide (VIP) in the submucous neurons in the rat digestive tract, we applied in vivo treatment of colchicine to enhance the immunoreactivity and examined the colocalization of NO synthase (nNOS) and VIP in neurons of the submucous plexus throughout the rat digestive tract. The density of nNOS-containing neurons in the submucous plexus in the stomach corpus (103±25 cells/cm², n=3) and that in the antrum (157±9 cells/cm², n=3) were significantly lower than those in small and large intestine. However no difference was detected in the cell density among duodenum (1967±188 cells/cm², n=3), jejunum (2640±140 cells/cm², n=3), ileum (2070±42 cells/cm², n=3), proximal colon (2243±138 cells/cm², n=3) and distal colon (2633±376 cells/cm², n=3). The proportion of nNOS-immunoreactive (IR), nNOS/VIP-IR and VIP-IR neurons to the total number of submucous neurons was examined. nNOS/VIP-IR neurons comprised 45–55% of total number of submucous neurons from the duodenum to the proximal colon, however those comprised 66.4±5.1% in the distal colon. The results showed that the dense distribution of nNOS-containing neurons was found in the submucous plexus throughout the small and large intestine, and large population of submucous neurons co-stored nNOS and VIP.     

Temporal variations in seed dispersal patterns of a bird-dispersed tree, <Emphasis Type="Italic">Swida controversa</Emphasis> (Cornaceae), in a temperate forest

The seed dispersal patterns of bird-dispersed trees often show substantial seasonal and annual variation due to temporal changes in frugivorous bird and bird-dispersed fruit distributions. Elucidating such variation and how it affects plant regeneration is important for understanding the evolution and seed dispersal maintenance strategies of these plants. In this study, we investigated the seed dispersal quantity and distance of a bird-dispersed plant, Swida controversa, for 2 years and detected large seasonal variations in dispersal pattern. Early in the fruiting season, short seed dispersal distance and large amounts of fruit consumption by birds (seed dispersal quantity) were observed. In contrast, late in the fruiting season, a long seed dispersal distance and small seed dispersal quantity were observed. This relationship between seed dispersal distance and quantity may help to maintain constant seed dispersal effectiveness during the long S. controversa fruiting season. Annual variation was also detected for both seed dispersal quantity and distance. More effective seed dispersal was achieved in the masting year, because both seed dispersal quantity and distance were greater than that in the non-masting year. These seed dispersal dynamics may contribute to the evolution and maintenance of S. controversa masting behavior. Thus, we identified substantial temporal variation on both seasonal and annual scales in the seed dispersal pattern of a bird-dispersed plant. The temporal variation in seed dispersal pattern revealed in this study probably plays a substantial role in the life history and population dynamics of S. controversa.     

Retarded decline in poly-ADPR content and poly-ADPR synthetase activity in chicken dystrophic muscle

The activity of poly-ADPR synthetase declines just after hatching in normal chick muscle nuclei. However, in dystrophic chick muscle nuclei it decreases 5 weeks after hatching. A delayed decrease in the amount of poly-ADPR is also observed in dystrophic chick muscle nuclei. These observations suggest that dystrophic muscle follows an abnormal developmental program.     

DNA Lesions Induced by Replication Stress Trigger Mitotic Aberration and Tetraploidy Development

During tumorigenesis, cells acquire immortality in association with the development of genomic instability. However, it is still elusive how genomic instability spontaneously generates during the process of tumorigenesis. Here, we show that precancerous DNA lesions induced by oncogene acceleration, which induce situations identical to the initial stages of cancer development, trigger tetraploidy/aneuploidy generation in association with mitotic aberration. Although oncogene acceleration primarily induces DNA replication stress and the resulting lesions in the S phase, these lesions are carried over into the M phase and cause cytokinesis failure and genomic instability. Unlike directly induced DNA double-strand breaks, DNA replication stress-associated lesions are cryptogenic and pass through cell-cycle checkpoints due to limited and ineffective activation of checkpoint factors. Furthermore, since damaged M-phase cells still progress in mitotic steps, these cells result in chromosomal mis-segregation, cytokinesis failure and the resulting tetraploidy generation. Thus, our results reveal a process of genomic instability generation triggered by precancerous DNA replication stress.     

Meristem-specific gene expression directed by the promoter of the S-phase-specific gene,cyc07, in transgenic Arabidopsis

A genomic clone for the cyc07 gene, which is expressed specifically at the S phase during the cell cycle in synchronous cultures of periwinkle (Catharanthus roseus) cells, was isolated. Determination of the nucleotide sequence of the clone revealed that the cyc07 gene consists of seven exons separated by six introns. Genomic Southern analysis indicated that the cyc07 gene is present as a single copy per haploid genome in periwinkle. Expression of related genes was detected in a wide range of other plants. Transgenic Arabidopsis plants were generated that expressed the gene for -glucuronidase (GUS) under the control of the promoter of the cyc07 gene. The tissue-specific pattern of expression directed by the promoter was investigated by analysis of GUS activity. Histochemical tests demonstrated that 589 bp of the 5-upstream sequence of the cyc07 gene could direct specifical expression of the GUS reporter gene in meristematic tissues in transgenic plants. The spatial pattern of expression directed by the promoter was closely correlated with meristematic activity and cell proliferation, suggesting an association between the function of the cyc07 gene and cell proliferation.     

Ranging behavior of Mahale chimpanzees: a 16 year study

We have analyzed the ranging patterns of the Mimikire group (M group) of chimpanzees in the Mahale Mountains National Park, Tanzania. During 16 years, the chimpanzees moved over a total area of 25.2 or 27.4 km², as estimated by the grid-cell or minimum convex polygon (MCP) methods, respectively. Annually, the M group used an average of 18.4 km², or approximately 70 %, of the total home-range area. The chimpanzees had used 80 % of their total home range after 5 years and 95 % after 11 years. M group chimpanzees were observed more than half of the time in areas that composed only 15 % of their total home range. Thus, they typically moved over limited areas, visiting other parts of their range only occasionally. On average, the chimpanzees used 7.6 km² (in MCP) per month. Mean monthly range size was smallest at the end of the rainy season and largest at the end of the dry season, but there was much variability from year to year. The chimpanzees used many of the same areas every year when Saba comorensis fruits were abundant between August and January. In contrast, the chimpanzees used several different areas of their range in June. Here range overlap between years was relatively small. Over the 16 years of the study we found that the M group reduced their use of the northern part of their range and increased their frequency of visits to the eastern mountainous side of their home range. Changes in home-range size correlated positively with the number of adult females but not with the number of adult males. This finding does not support a prediction of the male-defended territory model proposed for some East African chimpanzee unit-groups.     

Spectroscopic characterization of mononuclear, binuclear, and insoluble polynuclear oxovanadium(IV)-Schiff base complexes and their oxidation catalysis

The objective was to prepare mononuclear, binuclear, and insoluble polynuclear oxovanadium(IV)-Schiff base complexes and to use them for sulfoxidation and epoxidation of organic substrates. [VO(salen)] (complex 1) with tetradentate salen(salicylideneethylenediamine) being coordinated in the equatorial plane of oxovanadium(IV), [VO(salap)] (complex 2), and [(VO)₂(sal₂-dhdabp)] (complex 3) with tridentate salap(salicylideneorthoaminophenol) and sal₂-dhdabp(salicylidene-3,3^′-dihydroxy-4,4^′-diaminobiphenyl) being bound, respectively, in the equatorial plane, of which polynuclear complexes were constituted as monomer units, were prepared and spectroscopically characterized. A sulfide and olefins were oxidized by use of complexes 1 and 2 (mononuclear), complex 3 (binuclear), and the polynuclear complexes (poly-1 and poly-3) synthesized with 1 and 3, respectively. The reaction rates for poly-1 and -3 were a little lower than those of the corresponding 1 and 3. On oxidation of sulfides, poly-3 exhibited lowering of activity by about 15% in three cycles, while poly-1 showed significant lose of activity with each use. Poly-3 was efficient for the oxidation of the olefins only in the first cycle. It was suggested that the loss of activity depends not only on the coordination geometry of the oxovanadium complex, but also on the kind of the substrate.     

Difference of pine ectomycorrhizal biomass in relation to forest conditions

In this study, two plots in a secondary and another two in planted Pinus densiflora stands were exposed to different forest treatments, and the ectomycorrhizal (EM) biomass and its ergosterol content was measured for a year. The unmanaged plot in the secondary stand had greater EM biomass than those in any other plots. Whereas understory cutting had less effect on EM biomass, litter and humus removal decreased pine EM biomass and its ergosterol content, suggesting that such forest treatment alters EM biomass and its structure.