Proliferation of chick primordial germ cells cultured on stroma cells from the germinal ridge.

Fluorescent reagent-labelled PGCs isolated from the blood of 2-day-old chick embryos were cultured on stroma cells derived from 5-day-old germinal ridge in Medium 199 supplemented with 10% FBS, human IGF-1, bovine FGF-b, and murine LIF. In 7 experiments, the number of MCs increased by an average of 4.8 fold in 4 days. Intrinsic PGCs in the 5-day embryonic germinal ridge were observed loosely attached to the stroma cells, and they also increased 3.8 fold during culture for 4 days. These results indicate the possibility of applying this culture method to the production of transgenic chickens.     

Photoactivation of the latent water-oxidizing complex in photosystem II membranes isolated from dark-grown spruce seedlings

Photosystem II membranes (D-PSII) were isolated from dark-grown spruce seedlings. All major PSII proteins except the 17- and 23-kDa extrinsic proteins were present in D-PSII. O₂ evolution and Mn content in D-PSII were negligible, while PSII-donor activity showed a value comparable to that of NH₂OH-treated PSII membranes (NH₂OH-L-PSII) from light-grown seedlings. Light incubation of D-PSII with 1 m M MnCl₂, 50 m M CaCl₂ and 100 μ M DCIP at pH 5.3 resulted in activation of the latent water-oxidizing complex. Accomplishment of photoactivation of PSII membranes from dark-grown spruce seedlings clearly indicates that only ligation of Mn²⁺ to the apo-water oxidizing complex is required for expression of O₂ evolution, and that protein synthesis is not involved in the photoactivation process. There was no essential difference between 'photoactivation' of naturally Mn-free PSII membranes and 'photoreactivation' of artificially Mn-depleted PSII membranes on kinetics, pH dependence, Mn²⁺-concentration dependence. However, kinetics and pH dependence of photoactivation were appreciably different in spruce PSII membranes and in PSII membranes of angiosperms such as wheat and spinach.     

Computer-aided image analysis applied to immunogold-silver staining: evaluation of proliferating cell nuclear antigen (PCNA)-reactive sites in paraffin sections

Feasibility of the combination of the immunogold-silver staining method (IGSS) and computer-aided image analysis was assessed for the detection of antigen in an immunostained, paraffin-embedded section. Using low-temperature IGSS, we stained a specimen of human oral squamous cell carcinoma with a monoclonal antibody, PC 10, against a proliferating cell nuclear antigen (PCNA/cyclin), and the section was analyzed by ACAS 570 interactive laser cytometry. The PCNA-positive cells, exhibiting a heteromorphic texture, were contrasted by the dark staining of their nuclei, but showed heterogeneity in staining intensity from cell to cell. Using a conventional microscope light source rather than a laser, and by employing the COMPLEMENT DATA program (which permits inversion of the data values) installed in the ACAS 570 software system, we were able to obtain a complemental image which replicated the real immunohisto-morphology. Approximately 30–35 cells from three different areas in the same section were selected by DEFINE CELL and MARK AREA programs, and quantitative image analysis was performed in terms of cell integrated value, area, perimeter, and shape factor indicated in histogram form. The combined utilization of IGSS with computer-aided image analysis was demonstrated to offer a crucial advantage for the quantitative assessment of immunostained sections.     

Seasonal change in the food habits of the Iriomote catFelis iriomotensis

The food habits of the Iriomote catFelis iriomotensis were studied by analyzing 177 feces collected monthly from 1987 to 1988. A total of 26 food items were identified. The frequency of lizards and frogs in the feces was higher than those of mammals and birds. The food habits changed greatly seasonally.Eumeces skinks were fed on most frequently, and found in the feces with a similar high frequency occurrence in March–April and July–September, while their proportion to the total number of food items was larger in March–April than in July–September. The cats fed on larg-sized skinks, adultEumeces kishinouyei, more in March–April than in other seasons. The number of skinks sighted in the course of a road census was greater from March to August, and large-sized skinks were sighted more in March–April than in July–August. The cats fed selectively on large-sized skinks in every season. Therefore, changes in the food habits depended on the food availability. Characteristics of food habits in the Iriomote cat are discussed in comparison with the food habits of other felids in temperate and tropical regions.     

Acidiphilium aminolytica sp. nov.: An acidophilic chemoorganotrophic bacterium isolated from acidic mineral environment

Acidiphilium aminolytica is proposed for a species of the genusAcidiphilium. Acidiphilium aminolytica can be phenotypically differentiated from all other species of the genusAcidiphilium. The seven strains of this species that have been studied are Gram-negative, aerobic, mesophilic, non-sporeforming, motile, and rod-shaped bacteria. They grow between pH 3.0 and 6.0, but not at pH 6.5. They yield positive results in tests for hippuric acid hydrolysis, catalase and urease production. Oxidase, esculin hydrolysis, and -galactosidase tests are negative. They can used-glucose,d-galactose, inositol, sorbitol,l-lysine,l-glutamate,l-arginine, -alanine,dl-4-aminobutyrate,dl-5-aminovalerate, sperimine, or diaminobutane as a sole carbon source, but cannot use elemental sulfur and ferrous iron as an energy source. The DNA base composition is 58.7–59.2 G+C mol%. The major isoprenoid quinone is ubiquinone with ten isoprene unit (Q-10). The major fatty acid is the C_18:1 fatty acid. Two ornithine amide lipids, the C_18:1 fatty acid esters of -N-3-hydroxystearylornithyltaurine and -N-3-hydroxystearylornithine, are detected as the polar aminolipid. DNA relatedness between this species and the other species ofAcidiphilium, the generaAcidomonas, andAcidobacterium was 29 to 2%. These results indicate, that this new species should be placed in the genusAcidiphilium. The type strain (strain 101) ofA. aminolytica is JCM 8796.     

Growth Characteristics and Intraspecies Host Specificity of a Large Virus Infecting the Dinoflagellate Heterocapsa circularisquama

The growth characteristics and intraspecies host specificity of Heterocapsa circularisquama virus (HcV), a large icosahedral virus specifically infecting the bivalve-killing dinoflagellate H. circularisquama, were examined. Exponentially growing host cells were more sensitive to HcV than those in the stationary phase, and host cells were more susceptible to HcV infection in the culture when a higher percent of the culture was replaced with fresh medium each day, suggesting an intimate relationship between virus sensitivity and the physiological condition of the host cells. HcV was infective over a wide range of temperatures, 15 to 30°C, and the latent period and burst size were estimated at 40 to 56 h and 1,800 to 2,440 infective particles, respectively. Transmission electron microscopy revealed that capsid formation began within 16 h postinfection, and mature virus particles appeared within 24 h postinfection at 20°C. Compared to Heterosigma akashiwo virus, HcV was more widely infectious to H. circularisquama strains that had been independently isolated in the western part of Japan, and only 5.3% of the host-virus combinations (53 host and 10 viral strains) showed resistance to viral infection. The present results are helpful in understanding the ecology of algal host-virus systems in nature.     

Isolation of cDNA for a Plasma Membrane H+-ATPase from Guard Cells of Vicia faba L.

cDNA encoding the plasma membrane H⁺-ATPase of guard cells ofVicia faba L. was isolated. The clone encoded a 105-kDa polypeptide(956 amino acids) that was 79–85% identical in terms ofamino acid sequence to other plant H⁺-ATPases. High levels ofmRNA explain the high H⁺-ATPase activity of these plasma membranes. (Received December 24, 1994; Accepted April 12, 1995)     

Lethality of the covalent linkage between mislocalized major outer membrane lipoprotein and the peptidoglycan of Escherichia coli.

The major outer membrane lipoprotein (Lpp) of Escherichia coli possesses serine at position 2, which is thought to function as the outer membrane sorting signal, and lysine at the C terminus, through which Lpp covalently associates with peptidoglycan. Arginine (R) is present before the C-terminal lysine in the wild-type Lpp (LppSK). By replacing serine (S) at position 2 with aspartate (D), the putative inner membrane sorting signal, and by deleting lysine (K) at the C terminus, Lpp mutants with a different residue at either position 2 (LppDK) or the C terminus (LppSR) or both (LppDR) were constructed. Expression of LppSR and LppDR little affected the growth of E. coli. In contrast, the number of viable cells immediately decreased when LppDK was expressed. Prolonged expression of LppDK inhibited separation of the inner and outer membranes by sucrose density gradient centrifugation, whereas short-term expression did not. Pulse-labeled LppDK and LppDR were localized in the inner membrane, indicating that the amino acid residue at position 2 functions as a sorting signal for the membrane localization of Lpp. LppDK accumulated in the inner membrane covalently associated with the peptidoglycan and thus prevented the separation of the two membranes. Globomycin, an inhibitor of lipoprotein-specific signal peptidase II, was lethal for E. coli only when Lpp possessed the C-terminal lysine. Taken together, these results indicate that the inner membrane accumulation of Lpp per se is not lethal for E. coli. Instead, a covalent linkage between the inner membrane Lpp having the C-terminal lysine and the peptidoglycan is lethal for E. coli, presumably due to the disruption of the cell surface integrity.     

Numerical simulation of neuronal population coding: influences of noise and tuning width on the coding error

Performance of neuronal population coding is investigated numerically, in neurons with Gaussian tuning functions of various widths and noise ratios. The present model is applicable to both direction coding and orientation coding. It is shown that the coding error exhibits peculiar dependence on the width of the tuning function and that the dependence under the influence of noise is different from that of the noise-free case. In the absence of noise, the coding error increases monotonically with the width of the tuning function. The increment obeys the power law (the exponent estimated is 0.501) when the width is less than the critical value. In this region of the width a scaling law is obtained, which shows that the root-mean-square error is proportional to the square root of the ratio of the width of the tuning function to the population size. When the width exceeds the critical value, the coding error increases more rapidly than the power law. The reason for this anomalous increase, not seen previously, is argued. Existence of noise changes the dependence of the coding error on the width of the tuning function. Unlike the noise-free case, the error under the influence of noise becomes minimum at an intermediate value of the width. The width that gives the minimum coding error is termed the optimum width in this article. The numerical results suggest that the optimum width is roughly proportional to the square root of the noise ratio but has only a weak dependence on the population size. It is further shown that the coding error for the optimum width increases sharply when the noise ratio exceeds about 0.5 and is inversely proportional to the square root of the population size.     

Distribution of growth regulators in relation to the light-induced geotropic responsiveness in Zea roots

Growth regulators were measured in extracts from the upper and lower halves of 7-mm apical segments of horizontally oriented, red-light-irradiated and non-irradiated roots of Zea mays L. cv. Golden Cross Bantam 70 which exhibit a georesponse only after an exposure to light. Abscisic acid (ABA) was measured by gas-liquid chromatography, auxin (indole-3-acetic acid, IAA) by the Avena straight-growth assay, and an unidentified growth inhibitor by a Zea root-growth assay. The ratio of ABA in the upper and lower halves was 1.6 in the irradiated roots and 1.0 in the non-irradiated ones. The total amount of ABA after irradiation was increased by a factor of ca. 1.8. The ratio of IAA in the upper and lower halves of irradiated and non-irradiated roots was 1:3.4 and 1:2.9, respectively. The content (or activity) of an unidentified growth inhibitor was highest in the lower halves of horizontally oriented roots which had been irradiated with red light. The unidentified growth inhibitor, rather than IAA or ABA, may be the major factor in the light-induced geotropic responsiveness in Zea roots.