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31.
Human Microtubule-Associated Protein-2c Localizes to Dendrites and Axons in Fetal Spinal Motor Neurons 总被引:2,自引:0,他引:2
Joanna S. Albala Yvonne Kress Wan-Kyng Liu Karen Weidenheim Shu-Hui C. Yen Bridget Shafit-Zagardo 《Journal of neurochemistry》1995,64(6):2480-2490
Abstract: Microtubule-associated protein-2 (MAP-2) functions to maintain neuronal morphology by promoting the assembly of microtubules. MAP-2c is an alternately spliced form of MAP-2, containing the first 151 amino acids of high-molecular-weight (HMW) MAP-2 joined to the last 321 amino acids, eliminating 1,352 amino acids specific to HMW MAP-2. A polyclonal antibody generated to the splice site of human MAP-2c was used to determine its cellular localization. The MAP-2c antiserum was depleted of any HMW MAP-2 reactivity by absorption with HMW MAP-2 fusion protein. Western blot analysis of human fetal spinal cord homogenates demonstrated that the antibody is specific for human MAP-2c. MAP-2c immunoreactivity was found in the perinuclear cytoplasm and processes of anterior motor neurons and large processes of the posterior column in sections from 22–24-week human fetal spinal cord. Double-label confocal microscopy was performed using the MAP-2c polyclonal antibody and either a HMW MAP-2 or a neurofilament protein (highly phosphorylated 160- and 200-kDa protein) monoclonal antibody to identify these processes as dendrites or axons, respectively. HMW MAP-2 and MAP-2c colocalized in cell bodies and dendrites of anterior motor neurons, demonstrating for the first time the presence of native MAP-2c within dendrites. In addition, immunoelectron microscopy showed MAP-2c associated with microtubules in dendrites of motor neurons. MAP-2c and the neurofilament proteins were found in axons of the dorsal and ventral roots. The presence of MAP-2c within axons and dendrites suggests that MAP-2c contributes to neuronal plasticity during human fetal development. 相似文献
32.
Expression of Biologically Active Basic Fibroblast Growth Factor by Genetically Modified Rat Primary Skin Fibroblasts 总被引:1,自引:0,他引:1
Jasodhara Ray Joanna Hogg reas S. Beutler Hideichi Takayama Andrew Baird Fred H. Gage 《Journal of neurochemistry》1995,64(2):503-513
Abstract: Basic fibroblast growth factor (FGF-2) is normally expressed as a cell-associated protein, and accordingly it is not clear how it exerts its action on target cells in vivo. It has been proposed that cells release, by death or other mechanisms, small amounts of FGF-2 that then acts in an autocrine manner. To address the question of whether it is necessary that FGF-2 remain cell associated or needs to be secreted from cells to have biological activity, we expressed the 18-kDa form of FGF-2 in primary fibroblasts as a cell-associated (FGF-2-B) or as a secreted (FGF-2-S) protein. FGF-2 protein is detected in cell lysates and membrane fractions of both cell types, whereas it is present in significant amounts only in the conditioned medium of FGF-2-S cells. No FGF-2 is detected in control (untransfected) cells. FGF-2-S cells also grow faster than the control or FGF-2-B cells. Yet, when evaluated for their ability to promote the survival of embryonic hippocampal neurons in vitro, both the cell types are active, establishing the activity of the transgene product. We conclude that FGF-2 is active when engineered to be expressed as a cell-associated form or secreted from cells. 相似文献
33.
Joanna K. Bowen Matthew D. Templeton Keith R. Sharrock Ross N. Crowhurst Erik H. A. Rikkerink 《Molecular & general genetics : MGG》1995,246(2):196-205
The feasibility of performing routine transformation-mediated mutagenesis in Glomerella cingulata was analysed by adopting three one-step gene disruption strategies targeted at the pectin lyase gene pnIA. The efficiencies of disruption following transformation with gene replacement- or gene truncation-disruption vectors were compared. To effect replacement-disruption, G. cingulata was transformed with a vector carrying DNA from the pnlA locus in which the majority of the coding sequence had been replaced by the gene for hygromycin B resistance. Two of the five transformants investigated contained an inactivated pnlA gene (pnlA
–
);both also contained ectopically integrated vector sequences. The efficacy of gene disruption by transformation with two gene truncation-disruption vectors was also assessed. Both vectors carried a 5and 3truncated copy of the pnlA coding sequence, adjacent to the gene for hygromycin B resistance. The promoter sequences controlling the selectable marker differed in the two vectors. In one vector the homologous G. cingulata gpdA promoter controlled hygromycin B phosphotransferase expression (homologous truncation vector), whereas in the second vector promoter elements were from the Aspergillus nidulans gpdA gene (heterologous truncation vector). Following transformation with the homologous truncation vector, nine transformants were analysed by Southern hybridisation; no transformants contained a disrupted pnlA gene. Of nineteen heterologous truncation vector transformants, three contained a disrupted pnlA gene; Southern analysis revealed single integrations of vector sequence at pnlA in two of these transformants. pnlA mRNA was not detected by Northern hybridisation in pnlA-transformants. pnlA-transformants failed to produce a PNLA protein with a pI identical to one normally detected in wild-type isolates by silver and activity staining of isoelectric focussing gels. Pathogenesis on Capsicum and apple was unaffected by disruption of the pnlA gene, indicating that the corresponding gene product, PNLA, is not essential for pathogenicity. Gene disruption is a feasible method for selectively mutating defined loci in G. cingulata for functional analysis of the corresponding gene products. 相似文献
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The effects of calystegines isolated from edible fruits and vegetables on mammalian liver glycosidases 总被引:1,自引:0,他引:1
Asano Naoki; Kato Atsushi; Matsui Katsuhiko; Watson Alison A.; Nash Robert J.; Molyneux Russell J.; Hackett Lucy; Topping Joanna; Winchester Bryan 《Glycobiology》1997,7(8):1085-1088
The polyhydroxylated nortropane alkaloids called calyste-ginesoccur in many plants of the Convolvulaceae, Solanaceae, andMoraceae families. Certain of these alkaloids exhibit potentinhibitory activities against glycosidases and the recentlydemonstrated occurrence of calystegines in the leaves, skins,and sprouts of potatoes (Solatium tuberosum), and in the leavesof the eggplant (S.melongena), has raised concerns regardingthe safety of these vegetables in the human diet. We have surveyedthe occurrence of calystegines in edible fruits and vegetablesof the families Convolvulaceae, Solanaceae, and Moraceae byGC-MS. Calystegines A3, B1, B2, and C1 were detected in allthe edible fruits and vegetables tested; sweet and chili peppers,potatoes, eggplants, tomatoes, Physalis fruits, sweet potatoes,and mulberries. Calystegines B1 and C1 were potent competitiveinhibitors of the bovine, human, and rat β-glucosidaseactivities, with K1 values of 150, 10, and 1.9 µM, respectivelyfor B1 and 15,1.5, and 1 µM, respectively, for C1. CalystegineB2 was a strong competitive inhibitor of the -galactosidaseactivity in all the livers. Human β-xylosidase was inhibitedby all four nortropanes, with calystegine C1 having a K1 of0.13 µM. Calystegines A3 and B2 selectively inhibitedthe rat liver β-glucosidase activity. The potent inhibitionof mammalian β-glucosidase and -galactosidase activitiesin vitro raises the possibility of toxicity in humans consuminglarge amounts of plants that contain these compounds. edible plants calystegines glycosidase inhibitors bovine, human, and rat liver 相似文献
36.
Size, age and number of offspring at the first and subsequent tworeproductions were compared in Daphnia treated with cues frominvertebrate predators (adult Cyclopoida fed Daphnia)and from crushed conspecifics. Daphnia in the presence ofinvertebrate predators had earlier maturation, greater size atfirst and subsequent reproductions and a larger number of offspringthan control animals, while the opposite (later maturation andsmaller size at subsequent reproductions) happened when Daphnia were treated with homogenate from crushedconspecifics. 相似文献
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38.
Effects of acute hypoxia on incorporation of [1-14C]arachidonic acid into glycerolipids of rat brain
The effects of 1 min of acute hypoxic treatment (1% O2 in N2) on incorporation of [1-14C]arachidonic acid into brain lipids of 16-day-old rats were investigated at 3, 6, and 12 min after intracerebral injection of the labeled fatty acid. The hypoxic-hypoxia condition associated with convulsive seizures caused a decrease in the conversion of labeled arachidonate to its acyl-CoA as well as incorporation of the label into the brain phospholipids. Among the phospholipids, there was a specific decrease in the labeling of diacylglycerophosphoinositol (GPI), and this change was accompanied by an increase in labeling of the diacylglycerols. These results indicate that metabolism of the long-chain fatty acids and some glycero-lipids in brain are vulnerable to acute hypoxic treatment. 相似文献
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