Detection of Specific RNAs by in situ Hybridization in Plant Protoplasts

Combined exposure of fixed tomato protoplasts to 0.2 N HCl atroom temperature, heating at 70?C in 2 ? SSC for 30 min, anddigestion with 10 µg/ml proteinase K allowed localizationof viroid RNA by in situ hybridization, without destructionof cellular structures. The technique also allowed localizationof viral RNA in tobacco protoplasts. ⁴Present address: National Institute of Animal Health, Tsukuba,Ibaraki, 305 Japan (Received March 1, 1989; Accepted February 21, 1990)     

A model of neural network extracting binocular parallax

A model of neural network extracting binocular parallax is proposed. It is a multilayered network composed of analog threshold elements. Three types of binocular neurons are included in this model. They are binocular simple neurons, binocular gate neurons and binocular depth neurons. The final layers of this model consist of elements which correspond to the binocular depth neurons. The performance of the model has been simulated on a digital computer. The results of the computer simulation show that every element of this model acts like neurons found in cat's and monkey's visual system and this model extracts binocular parallax caused by simple line components satisfactorily.     

Real-time weather analysis reveals the adaptability of direct sea-crossing by raptors

Many animals seasonally travel between their breeding and wintering grounds. With their advanced mobility, birds often migrate over thousands of kilometres. Recently, satellite-tracking studies have revealed peculiar migration routes for some avian species at a global scale. However, the adaptability of such migration routes has not been clearly demonstrated. Using satellite-tracking data for 33 individuals, we show that the Japanese population of Oriental honey-buzzards (Pernis ptilorhynchus) directly crosses the 650-km-wide East China Sea during their autumn migration, although they fly a longer route around the sea rather than directly crossing it during their spring migration. By applying aerodynamic theory, we show that the buzzards could cross the sea by soaring and gliding flight. Moreover, using a high-resolution meteorological-prediction analysis, we demonstrate that the migratory trajectory of the birds strongly depends on the wind direction at their estimated locations. In the area, northeastern tailwinds blow stably only during autumn. Thermals were abundant ca. 500–1,000 m over the East China Sea in autumn, but that was not the case in spring. We suggest that the autumn-migration route across the East China Sea is likely to have evolved in response to the specific weather conditions over the sea. Animations showing movements of Oriental honey-buzzards and temporal change in weather conditions are available at: , , , , , and .     

Mating behavior and the function of the male genital spine in the ground beetle Carabus clathratus

The morphologies of male genitalia often appear harmful or aggressive, as if they may inflict physical damage upon females during copulation. Such male genitalia are often thought to function in intra- and intersexual interactions during mating. In the carabid genus Carabus, division Spinulati, males possess a spine (spinula) on the intromittent organ, of which function is unknown. To reveal the function of the spinula, we studied the mating behavior and genital coupling of a Spinulati species, Carabus (Limnocarabus) clathratus. The males positioned the spinula along the inner wall of the vaginal opening throughout copulation. This placement created a small dent and subsequently a melanized patch (wound) on the vaginal wall, but the spinula rarely penetrated the vaginal wall. The spinula did not reach the innermost part of the vagina where the spermatophore is deposited. These results suggest that the spinula is not used for inflicting damage on female genitalia or manipulating spermatophores of rival males. During spermatophore formation, the male partially withdrew the aedeagus, and only the aedeagal tip and endophallus remained within the female. By placing the spinula against the vaginal wall, the male could hold the endophallus within the vaginal chamber in the unstable copulatory posture. Thus, our observations suggest that the spinula primarily functions as an "anchor" to maintain the coupling of the male and female genitalia and thereby ensure insemination.     

BMP and Hh signaling affects primordial germ cell division in Drosophila

The germline is segregated from the remainder of the soma during early embryonic development in metazoan species. In Drosophila, female primordial germ cells (PGCs) continue to proliferate during larval development, and become germline stem cells at the early pupal stage. To elucidate the roles of growth factors in larval PGC division, we examined expression patterns of a bone morphogenetic protein (BMP) growth factor, Decapentaplegic (Dpp), and Hedgehog (Hh), along with factors downstream of each, in the ovary during larval development. Dpp signaling appeared in the ovarian soma from early larval development, and was prominent in the terminal filament cells at late larval stage, whereas Hh appeared in the ovarian soma and PGCs from the third instar larval stage. The number of PGCs decreased when components of these signal transduction pathways were abrogated by RNAi in the PGCs, indicating that both Dpp and Hh signals directly regulate PGC proliferation. Experiments on the up- and down-regulation of Dpp and Hh with a tissue-specific Gal4 driver indicated that Dpp and Hh act as extrinsic and autocrine growth factors. Furthermore, heat-pulse experiments with hs-Gal4 showed that Dpp is active in PGC proliferation throughout larval development, whereas Hh has effects only during late larval development. In addition to Dpp, the reduction of Glass bottom boat (Gbb), another BMP molecule, caused a decrease in the number of PGCs and initiation of larval PGCs differentiation into cystocytes, indicating that Gbb functions to promote PGC division and repress differentiation.     

Increased expression of co-chaperone HOP with HSP90 and HSC70 and complex formation in human colonic carcinoma

Co-chaperone HOP (also called stress-inducible protein 1) is a co-chaperone that interacts with the cytosolic 70-kDa heat shock protein (HSP70) and 90-kDa heat shock protein (HSP90) families using different tetratricopeptide repeat domains. HOP plays crucial roles in the productive folding of substrate proteins by controlling the chaperone activities of HSP70 and HSP90. Here, we examined the levels of HOP, HSC70 (cognate of HSP70, also called HSP73), and HSP90 in the tumor tissues from colon cancer patients, in comparison with the non-tumor tissues from the same patients. Expression level of HOP was significantly increased in the tumor tissues (68% of patients, n = 19). Levels of HSC70 and HSP90 were also increased in the tumor tissues (95% and 74% of patients, respectively), and the HOP level was highly correlated with those of HSP90 (r = 0.77, p < 0.001) and HSC70 (r = 0.68, p < 0.01). Immunoprecipitation experiments indicated that HOP complexes with HSC70 or HSP90 in the tumor tissues. These data are consistent with increased formation of co-chaperone complexes in colon tumor specimens compared to adjacent normal tissue and could reflect a role for HOP in this process.     

Olfactory Imprinting of Amino Acids in Lacustrine Sockeye Salmon

Juvenile salmon have an olfactory ability to imprint their natal stream odors, but neither the odor properties of natal stream water nor the imprinting timing and duration have been clarified as yet. Here we show, using electrophysiological and behavioral experiments, that one-year-old lacustrine sockeye salmon (Oncorhynchus nerka) can be imprinted around the stage of parr-smolt transformation (PST) by a single amino acid, 1 µM L-proline (Pro), or L-glutamic acid (Glu). We also show by real-time PCR that changes occur in mRNA levels of the salmon olfactory imprinting-related gene (SOIG) around PST. The electro-olfactogram (EOG) responses of test fish exposed to Pro in March (before PST) and April–June (during PST) for 2 weeks were significantly (1.7-fold) greater than those of non-exposed control fish, but not those of test fish exposed in July (after PST). When Pro and control water were added to the water inlets of a two-choice test tank during the spawning season 2 years after the test water exposure, 80% of maturing and matured test fish exposed before and during PST showed a preference for Pro, whereas those exposed after PST did not. The EOG response of test fish exposed to Pro or Glu for 1 hour, 6 hours, 1 day, 7 days, or 14 days in May revealed that only the response after 14 days of exposure was significantly (1.8-fold) greater than the control. The expression levels of SOIG mRNA increased before and during PST, and decreased after PST. We conclude that one-year-old lacustrine sockeye salmon can be imprinted by a single amino acid before and during PST, and that imprinting requires exposure for at least 14 days.     

Study on immunocapture-chemiluminescence assay of lipase activity in a biological sample.

A new approach for the determination of lipase (triacylglycerol lipase, EC.3.1.1.3) activity in a biological sample was investigated by combining an immunocapture technique with a chemiluminescence (CL) assay method in order to eliminate interference with CL detection. The proposed method consists of an immunocapture step to trap lipase and a subsequent step for CL detection of the activity of the captured lipase. The CL detection is based on the luminol-hydrogen peroxide (H(2)O(2))-horseradish peroxidase (HRP) reaction and utilizes a proenhancer substrate [a lauric acid ester of 2-(4-hydroxyphenyl)-4,5-diphenylimidazole (HDI)] which liberates an active enhancer, HDI, by enzymatic hydrolysis. A polyclonal antibody prepared with porcine pancreas lipase was used for the immunocapture. The proposed immunocapture-CL method effectively eliminated the interference with the CL reaction from biological components and enabled the determination of spiked porcine pancreas lipase activity in serum samples in the range 0.41-1.1 U(HDI) (1 U(HDI) corresponds to the amount which liberates 1 pmol HDI/min at 37 degrees C from the substrate). The method was further applied to the assay of the activity for human pancreas lipase in serum and the results showed good correlation (r = 0.871) with those by the conventional colorimetric method.     

Novel emission properties of melem caused by the heavy metal effect of lanthanides(III) in a LB film.

A novel emissive molecular system is constructed by the intercalation of the fluorophore melem (triamino-tri-s-triazine) within a Langmuir-Blodgett (LB) film of stearic acid with the periodic arrangement of lanthanides (Ln(III)), mainly Pr(III) with supporting of Eu(III). From emission spectra, decay curves, quantum yields and XPS measurements, it is clarified that the external heavy metal effect of Pr(III) on melem is much stronger in the film than in the bulk solid state, resulting in producing an unusual triplet state of melem. The triplet state of melem in the LB film donates the excitation energy to Pr(III) in the LB film, which is completely different from the energy transfer pathway of Pr-melem complex in the solid state through the singlet state of melem.     

Enhancement of growth and cellulose accumulation by overexpression of xyloglucanase in poplar

Because the loosening of xyloglucan in the cell wall promotes plant growth (Takeda et al. (2002) Proc. Natl. Acad. Sci. USA 99, 9055-9060; Park et al. (2003) Plant J. 33, 1099-1106), we expressed Aspergillus xyloglucanase constitutively in Populus alba. The expression increased the length of stem even in the presence of sucrose. Increased stem growth was accompanied by a decrease in Young's elastic modulus in the growth zone but an increased elasticity in mature tissue. The increased internode length corresponded to an increase in cellulose content as well as specific gravity, showing that the removal of xyloglucan might cause an increase in cellulose density in the secondary xylem.