Immunocytochemical studies of hamster oocyte activation

By indirect immunofluorescence, using rabbit anti-heparin-binding placental protein (HBPP) antiserum, we studied HBPP expression by physiologically and non-physiologically (microsurgically) activated hamster gametes. Whereas mature gametes (sperm, metaphase II oocytes) were negative, in vivo conceived preimplantation embryos, from pronuclear to two- and four-cell stages, were HBPP positive. No HBPP was demonstrated in the zona pellucida, but HBPP-dependent immunofluorescence was localized in the perivitelline space. Oocytes incubated with hyaluronidase demonstrated variable responses from negative to positive. (Diluent or sperm) microinjected oocytes were all activated and HBPP positive within 4 h after stimulation. Thus neither activation by microinjection nor HBPP expression required paternal gametes. These kinetics suggest that HBPP may be a cortical granule secretogogue which can be applied to monitor oocyte responses during in vitro manipulations.     

Immunological Characterization and Transmembrane Topology of 5-Hydroxytryptamine3 Receptors by Functional Epitope Tagging

Abstract: 5-Hydroxytryptamine₃ (5-HT₃) receptors are the only known monoamine receptors mediating fast excitatory responses in mammalian neurons. Their primary structure as well as their electrophysiological and pharmacological properties show a phylogenetic relation to nicotinic acetylcholine, GABA_A, and glycine receptors. As a prototypical member of this gene superfamily, we investigated the membrane topology of functional homomeric 5-HT₃ receptors by using epitope tagging of the channel subunits expressed in heterologous systems. Visualization of 5-HT₃ receptors in transfected COS-7 cells, either in western blot (molecular mass 61.2 ± 0.8 kDa) or in situ, was performed with previously characterized antibodies recognizing artificial epitopes as well as with anti-fusion protein antibodies directed against a wild-type receptor intracellular domain. The extracellular location of the distal C-terminal tagged domain demonstrates the presence of a fourth transmembrane domain in 5-HT₃ serotonin-gated channels. In this region, the significant homology between members of this class of neurotransmitter-gated channels suggests strongly that they have a common transmembrane organization basically different from glutamate-gated and ATP-gated channels.     

Multiple roles of the mitogen-activated protein kinase kinase/mitogen-activated protein kinase cascade in Xenopus laevis

Mitogen-activated protein kinase (MAPK) was originally identified as a serine/threonine protein kinase that is rapidly activated in response to various growth factors and tumor promoters in mammalian cultured cells. The kinase cascade including MAPK and its direct activator, MAPK kinase (MAPKK), is now believed to transmit various extracellular signals into their intracellular targets in eukaryotic cells. It has been reported that activation of MAPKK and MAPK occurs during the meiotic maturation of oocytes in several species, including Xenopus laevis . Studies with neutralizing antibodies against MAPKK, MAPK phosphatases and constitutively active MAPKK or MAPK have revealed a crucial role of the MAPKK/MAPK cascade in a number of developmental processes in Xenopus oocytes and embryos.     

Effect of ammonium sulfate on the phytotoxicity, foliar uptake, and translocation of imazamethabenz in wild oat

Experiments were conducted in greenhouse, growth chamber, and laboratory conditions to determine the effect of ammonium sulfate [(NH₄)₂SO₄] on the phytotoxicity, foliar uptake, and translocation of imazamethabenz on wild oat. Rates of (NH₄)₂SO₄ up to 5% (w/v) applied with a greenhouse sprayer did not affect the phytotoxicity of the herbicide when the mix was applied at the one- to two-leaf stage. However, inclusion of 1 and 2% (NH₄)₂SO₄ increased the phytotoxicity of the herbicide when the mix was sprayed at the two- to three-leaf, or the three- to four-leaf stage. At 10%, (NH₄)₂SO₄ decreased the phytotoxicity of the sublethal dosage of the herbicide. When the herbicide was applied as individual drops to the growth chamber-grown plants, inclusion of (NH₄)₂SO₄ at 1% did not affect phytotoxicity as measured by shoot growth. The presence of (NH₄)₂SO₄ did not affect the amount of imazamethabenz retained by wild oat foliage, but it decreased [¹⁴C]imazamethabenz absorption, slightly antagonized acropetal translocation, and increased the basipetal translocation of [¹⁴C]imazamethabenz. It was concluded that application methods greatly modify the effect of (NH₄)₂SO₄ on imazamethabenz phytotoxicity. Herbicide absorption and translocation as determined by one method do not necessarily represent the absorption and translocation patterns when different application methods are used. Absorption and translocation were not the factors that were responsible for the observed effect of (NH₄)₂SO₄ on the herbicide phytotoxicity.Abbreviations SC suspension concentrate     

Intra- and interspecific predation on four life stage groups by the adult females of Metaseiulus occidentalis, Typhlodromus pyri, Neoseiulus fallacis and Amblyseius andersoni

Do adult females of oligophagous species such as Neoseiulus fallacis (Garman) and Metaseiulus occidentalis (Nesbitt) show less intra- and interspecific predation on phytoseiids when other foods are scarce than polyphagous species such as Amblyseius andersoni Chant and Typhlodromus pyri Scheuten? We caged single adult females of each species without food with ten of their own eggs or larvae, with ten eggs or larvae of the other species or with ten nymphs or adult females of M. occidentalis (T. pyri for M. occidentalis). We assessed the ambulatory activity, survival time, egg levels and prey loss in each test. Polyphages (in particular T. pyri) lived longer than oligophages (in particular N. fallacis) without food. The small T. pyri detected its own stages and benefited most by feeding on small active stages of other species. Amblyseius andersoni, the largest mite, fed and gained the most of any species when held with nymphs and female adults. Metaseiulus occidentalis fed on eggs of all four species to enhance survival. The large hyperactive N. fallacis gained the least from these behaviours. Each mite seemed uniquely adapted to survive conditions of scarce prey and these behaviours may explain their roles in phytoseiid mite complexes. Overall, oligophagous adult females fed less and gained less by feeding on phytoseiids than did polyphagous adult females.     

Immunocytochemical localization of callose in the germinated pollen ofCamellia japonica

Summary A polyclonal antibody against -1,3-glucan, callose, extracted from the pollen tube wall ofCamellia japonica was raised in mice and, using it as a probe, the localization of callose in the germinated pollen was studied. By confocal laser scanning microscopy, callose was found in the tip region of the pollen tube and the tube wall; the immuno-fluorescence in the tube wall was less toward the base of the tube. In contrast, the tip region did not fluoresce although the whole of the tube wall did strongly with aniline blue, the specific dye for callose. Immuno-electron microscopy showed that callose was also found in Golgi vesicles which concentrated in the tip region of the pollen tube, the inner layer of the tube wall, callose plugs, and Golgi vesicles in the pollen grain. Immuno-gold labeling was often detected on the fibrous structures in Golgi vesicles and callose plugs. Based on these results, the participation of Golgi vesicles in the formation of the tube wall and callose plugs was discussed.Abbreviation TBS Tris-buffered saline - Tris Tris(hydroxy-methyl)-aminomethane - PBS phosphate-buffered saline - BSA bovine serum albumin - ELISA enzyme-linked immunosorbent assay - CLSM confocal laser scanning microscopy - DP degree of polymerization     

Comparative ultrastructure of aecial and telial infections of the autoecious rust fungus Puccinia tuyutensis

This study demonstrates morphological differences between aecial and telial stages of the autoecious rust Puccinia tuyutensis. The aeciospores possess verrucose ornamentation while the teliospores have smooth surfaces. The aecial and telial haustoria of this rust produced in the mesophyll of Cressa cretica differ morphologically in the following respects:(1) the haustorial mother cell of telial haustorium is more differentiated than that of aecial haustorium and its wall at the penetration site is composed of 4 layers; (2) the aecial haustorium is filamentous in appearance and slightly constricted at the point of entry into the host cell, while the telial haustorium is clavate and possesses a narrow neck with a densely staining neckband and swollen body; (3) the neck of the telial haustorium is always associated with numerous vesicles while that of the aecial haustorium is not. Vascular tissue of host leaves is heavily invaded by aecial haustoria but not by telial haustoria.     

Prediction of a low dose herbicide effect from studies on binding of metribuzin to the chloroplasts of Chenopodium album L.

Chenopodium album L. seedlings at the 4- and 8-leaf stage were exposed to low concentrations metribuzin [4-amino-6-(l, l-dimethyl)-3-(methylthio)-l,2,4-triazin (4 H )-one] in nutrient solution to study herbicide uptake and the effects of low-dose rates. Chlorophyll fluorescence was measured to relate the inhibition of photosynthesis to herbicide dose. The minimum rate at which metribuzin fully inhibited photosynthesis was less than 1 μM for seedlings at the 4-leaf stage of development, and between 1 and 5 μM for the 8-leaf stage seedlings. With isolated chloroplasts, experiments were conducted to establish the relationship between the amount of herbicide molecules bound to each chloroplast and the inhibition of photosynthesis. From the dose-response curves obtained it was calculated that photosynthesis was fully inhibited when 7.5 10⁵ molecules metribuzin were bound to each chloroplast. This amount of binding was used to estimate minimum-lethal dose rates of metribuzin required for seedlings differing in fresh weight of leaves and amounts of chloroplasts present. It is suggested that prediction of a low dose herbicide effect from studies on binding of photosystem-II inhibitors in combination with chlorophyll fluorescence measurements may lead to the development of a new weed management strategy.     

Sex ratio regulation in the waspEupelmus vuilleti (crwf.), an ectoparasitoid on bean weevil larvae (Hymenoptera: Pteromalidae)

In Africa, the seeds and/or pods ofVigna unguiculata andVigna radiata (Papilionacea) are attacked in fields and storage structures by bruchid beetlesCallosobruchus maculatus andBruchidius atrolineatus, on which parasitoid communities can develop. One of these parasitoids is the solitary ectoparasitoidEupelmus vuilleti (Eupelmidae). The storage conditions ofV. unguiculata andV. radiata favor the overlapping of all host stages during several months. These conditions suggest that female parasitoids would vary the sex ratio of their offspring according to the different sizes or developmental stages of hosts. The sex ratio ofE. vuilleti was strongly related to the developmental stage of the hostC. maculatus. Under our experimental conditions, where superparasitism is rare, the proportion of daughters varied between 5 and 25% on the third larval stage but reached 70 to 90% on the pupae. The increase in the proportion of daughters was also observed in the absence of superparasitism. In this case, there was an absolute coincidence between the sex ratio of eggs laid and that of emerged adults. Manipulation of the sex of the egg by the females seems to take place at the time of the egg's fertilization. The relation between host weight and egg sex showed that the male eggs are preferentially laid on lighter host larvae and the female eggs on heavier ones.     

用重组痘苗病毒表达pGHcDNA的研究

构建了含有ｐＧＨｃＤＮＡ的重组痘苗病毒,用ＥＬＩＳＡ证明该重组病毒在被感染的ｈ１４３细胞中,可表达出猪生长激素并将之分泌到培养基中,表达量约为１．０５μｇ／１０￣６细胞（２４ｈ）。用定位免疫化学法进一步证明该病毒可感染小鼠并在小鼠体内表达ｐＧＨｃＤＮＡ。同时还构建了含双拷贝ｐＧＨｃＤＮＡ的重组痘苗病毒,并证明其ｐＧＨ表达量比单拷贝重组病毒有明显提高,约为１．５０μｇ／１０￣６细胞（２４ｈ）。