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1.
Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs 总被引:9,自引:6,他引:3 下载免费PDF全文
Cortical granules, which are specialized secretory organelles found in ova of many organisms, have been isolated from the eggs of the sea urchins Arbacia punctulata and Strongylocentrtus pupuratus by a simple, rapid procedure. Electron micropscope examination of cortical granules prepared by this procedure reveals that they are tightly attached to large segments of the plasma membrane and its associated vitelline layer. Further evidence that he cortical granules were associated with these cell surface layers was obtained by (125)I-labeling techniques. The cortical granule preparations were found to be rich in proteoesterase, which was purified 32-fold over that detected in a crude homogenate. Similarly, the specific radioactivity of a (125)I-labeled, surface glycoprotein was increased 40-fold. These facts, coupled with electron microscope observations, indicate the isolation procedure yields a preparation in which both the cortical granules and the plasma membrane-vitelline layer are purified to the same extent. Gel electrophoresis of the membrane-associated cortical granule preparation reveals the presence of at least eight polypeptides. The major polypeptide, which is a glycotprotein of apparent mol wt of 100,000, contains most of the radioactivity introduced by (125)I-labeling of the intact eggs. Lysis of the cortical granules is observed under hypotonic conditions, or under isotonic conditions if Ca(2+) ion is present. When lysis is under isotonic conditions is induced by addition of Ca(2+) ion, the electron-dense contents of the granules remain insoluble. In contrast, hypotonic lysis results in release of the contents of the granule in a soluble form. However, in both cases the (125)I-labeled glycoprotein remains insoluble, presumably because it is a component of either the plasma membrane or the vitelline layer. All these findings indicate that, using this purified preparation, it should be possible to carry out in vitro studies to better define some of the initial, surface-related events observed in vivo upon fertilization. 相似文献
2.
Jennifer C. Utting Adrienne M. Flanagan Andrea Brandao‐Burch Isabel R. Orriss Timothy R. Arnett 《Cell biochemistry and function》2010,28(5):374-380
Active pathological bone destruction in humans often occurs in locations where oxygen tension (pO2) is likely to be low, for example, at the sites of tumours, inflammation, infections and fractures, or the poorly vascularized yellow fatty marrow of the elderly. We examined the effect of pO2 on formation of osteoclasts, the cells responsible for bone resorption, in 14‐day cultures of normal human peripheral blood mononuclear cells (hPBMCs) on ivory discs. Hypoxia (1–2% O2) caused threefold increases in the number of osteoclasts formed, compared with 20% O2. Hypoxia also caused a twofold increase in the number of nuclei per osteoclast, leading to stimulations of resorption pit formation of up to 10‐fold. Exposure to hypoxia led to stabilization of the hypoxia‐inducible factors, HIF1α and HIF2α, and upregulation of vascular endothelial growth factor and interleukin‐6 expression by hPBMCs. These findings help explain why extravasation of mononuclear precursors into relatively O2‐deficient bone microenvironments could result in osteoclast formation and suggest a new mechanism for the bone loss associated with the pathophysiological conditions where hypoxia commonly occurs. Copyright © 2010 John Wiley & Sons, Ltd. 相似文献
3.
Phylogenetic utility of the nuclear gene arginine decarboxylase: an example from Brassicaceae 总被引:10,自引:2,他引:8
Arginine decarboxylase (ADC) is an important enzyme in the production of
putrescine and polyamines in plants. It is encoded by a single or low-copy
nuclear gene that lacks introns in sequences studied to date. The rate of
Adc amino acid sequence evolution is similar to that of ndhF for the
angiosperm family studied. Highly conserved regions provide several target
sites for PCR priming and sequencing and aid in nucleotide and amino acid
sequence alignment across a range of taxonomic levels, while a variable
region provides an increased number of potentially informative characters
relative to ndhF for the taxa surveyed. The utility of the Adc gene in
plant molecular systematic studies is demonstrated by analysis of its
partial nucleotide sequences obtained from 13 representatives of
Brassicaceae and 3 outgroup taxa, 2 from the mustard oil clade (order
Capparales) and 1 from the related order Malvales. Two copies of the Adc
gene, Adc1 and Adc2, are found in all members of the Brassicaceae studied
to data except the basal genus Aethionema. The resulting Adc gene tree
provides robust phylogenetic data regarding relationships within the
complex mustard family, as well as independent support for proposed tribal
realignments based on other molecular data sets such as those from
chloroplast DNA.
相似文献
4.
Annerieke C van Groenestijn Ingrid GL van de Port Carin D Schröder Marcel WM Post Hepke F Grupstra Esther T Kruitwagen Harmen van der Linde Reinout O van Vliet Margreet GH van de Weerd Leonard H van den Berg Eline Lindeman 《BMC neurology》2011,11(1):1-11
Background
Neuropathic pain must be correctly diagnosed for optimal treatment. The questionnaire named Neuropathic Pain Symptom Inventory (NPSI) was developed in its original French version to evaluate the different symptoms of neuropathic pain. We hypothesized that the NPSI might also be used to differentiate neuropathic from non-neuropathic pain.Methods
We translated the NPSI into German using a standard forward-backward translation and administered it in a case-control design to patients with neuropathic (n = 68) and non-neuropathic pain (headache and osteoarthritis, n = 169) to validate it and to analyze its discriminant properties, its sensitivity to change, and to detect neuropathic pain subgroups with distinct profiles.Results
Using a sum score (the NPSI-G score), we found sensitivity to change (r between 0.37 and 0.5 for pain items of the graded chronic pain scale) and could distinguish between neuropathic and other pain on a group basis, but not for individual patients. Post hoc development of a discriminant score with optimized diagnostic properties to distinguish neuropathic pain from non-neuropathic pain resulted in an instrument with high sensitivity (91%) and acceptable specificity (70%). We detected six different pain profiles in the patient group with neuropathic pain; three profiles were found to be distinct.Conclusions
The NPSI-G potentially combines the properties of a diagnostic tool and an instrument to identify subtypes of neuropathic pain. 相似文献5.
6.
Cesca F Yabe A Spencer-Dene B Scholz-Starke J Medrihan L Maden CH Gerhardt H Orriss IR Baldelli P Al-Qatari M Koltzenburg M Adams RH Benfenati F Schiavo G 《Cell death and differentiation》2012,19(2):194-208
Signaling downstream of receptor tyrosine kinases controls cell differentiation and survival. How signals from different receptors are integrated is, however, still poorly understood. In this work, we have identified Kidins220 (Kinase D interacting substrate of 220 kDa)/ARMS (Ankyrin repeat-rich membrane spanning) as a main player in the modulation of neurotrophin and vascular endothelial growth factor (VEGF) signaling in vivo, and a primary determinant for neuronal and cardiovascular development. Kidins220(-/-) embryos die at late stages of gestation, and show extensive cell death in the central and peripheral nervous systems. Primary neurons from Kidins220(-/-) mice exhibit reduced responsiveness to brain-derived neurotrophic factor, in terms of activation of mitogen-activated protein kinase signaling, neurite outgrowth and potentiation of excitatory postsynaptic currents. In addition, mice lacking Kidins220 display striking cardiovascular abnormalities, possibly due to impaired VEGF signaling. In support of this hypothesis, we demonstrate that Kidins220 constitutively interacts with VEGFR2. These findings, together with the data presented in the accompanying paper, indicate that Kidins220 mediates the integration of several growth factor receptor pathways during development, and mediates the activation of distinct downstream cascades according to the location and timing of stimulation. 相似文献
7.
Ricci GC De Souza-Kaneshima AM Felismino MF Mendes-Bonato AB Pagliarini MS Do Valle CB 《Journal of genetics》2011,90(2):289-294
A total of 44 accessions of Brachiaria decumbens were analysed for chromosome count and meiotic behaviour in order to identify potential progenitors for crosses. Among them,
15 accessions presented 2n = 18; 27 accessions, 2n = 36; and 2 accessions, 2n = 45 chromosomes. Among the diploid accessions, the rate of meiotic abnormalities was low, ranging from 0.82% to 7.93%. In
the 27 tetraploid accessions, the rate of meiotic abnormalities ranged from 18.41% to 65.83%. The most common meiotic abnormalities
were related to irregular chromosome segregation, but chromosome stickiness and abnormal cytokinesis were observed in low
frequency. All abnormalities can compromise pollen viability by generating unbalanced gametes. Based on the chromosome number
and meiotic stability, the present study indicates the apomictic tetraploid accessions that can act as male genitor to produce
interspecific hybrids with B. ruziziensis or intraspecific hybrids with recently artificially tetraploidized accessions. 相似文献
8.
本文通过建立图象分析方法对免疫组织化学反应结果进行定量,检测观察H-ras在口腔颊粘膜上皮在正常(N)、慢性炎症(IF)、癌旁上皮(EAC)和鳞癌(SCC)的变化过程中的表达并进行分析。结果显示H-ras在SCC组中,以中等分化的SCC无论是H-ras表达的量还是细胞阳性率都较高。此外,组织学观察显示,H-ras在处于分化末期但尚未角化的正常上皮细胞中有较高的表达。本文结果显示了H-ras的过表达与上皮细胞的会化程度密切相关。本研究还显示,所采用的阳性区域透光值、平均总透光值及阳性反应区域与阴性反应区域比值可靠并有相关性。这进一步说明了用免疫组化定量方法检测H-ras癌基因表达的精确和可靠性。 相似文献
9.
Lectin-binding sites on the plasma membranes of rabbit spermatozoa: Changes in surface receptors during epididymal maturation and after ejaculation 总被引:2,自引:5,他引:2 下载免费PDF全文
GL Nicolson N Usui R Yanagimachi H Yanagimachi Smith JR 《The Journal of cell biology》1977,74(3):950-962
Modifications in rabbit sperm plasma membranes during epididymal passage and after ejaculation were investigated by used of three lectins: concanavalin A (Con A); Ricinus communis I (RCA(I)); and wheat germ agglutinin (WGA). During sperm passage from caput to cauda epididymis, agglutination by WGA drastically decreased, and agglutination by RCA(I) slightly decreased, although agglutination by Con A remained approximately unchanged. After ejaculation, spermatozoa were agglutinated to a similar degree or slightly less by Con A, WGA, and RCA(I), compared to cauda epididymal spermatozoa. Ultrastructural examination of sperm lectin-binding sites with ferritin- lectin conjugates revealed differences in the densities of lectin receptors in various sperm regions, and changes in the same regions during epididymal passage and after ejaculation. Ferritin-RCA(I) showed abrupt changes in lectin site densities between acrosomal and postacrosomal regions of sperm heads. The relative amounts of ferritin-RCA(I) bound to heads of caput epididymal or ejaculated spermatozoa. Tail regions were labeled by ferritin RCA(I) almost equally on caput and cauda epididymal spermatozoa, but the middle-piece region of ejaculated spermatozoa was slightly more densely labeled than the principal-piece region, and these two regions on ejaculated spermatozoa were labeled less than on caput and cuada epididymal spermatozoa. Ferritin-WGA densely labeled the acrosomal region of caput epididymal spermatozoa, although labeling of cauda epidiymal spermatozoa was relatively sparse except in the apical area of the acrosomal region. Ejaculated spermatozoa bound only a few molecules of ferritin-WGA, even at the highest conjugate concentrations used. Caput epididymal, but not cauda epididymal or ejaculated spermatozoa, bound ferritin-WGA in the tail regions. Dramatic differences in labeling densities during epididymal passage and after ejaculation were not found with ferritin-Con A. 相似文献
10.
Lee PA Orriss GL Buchanan G Greene NP Bond PJ Punginelli C Jack RL Sansom MS Berks BC Palmer T 《The Journal of biological chemistry》2006,281(45):34072-34085
The cytoplasmic membrane protein TatB is an essential component of the Escherichia coli twin-arginine (Tat) protein translocation pathway. Together with the TatC component it forms a complex that functions as a membrane receptor for substrate proteins. Structural predictions suggest that TatB is anchored to the membrane via an N-terminal transmembrane alpha-helix that precedes an amphipathic alpha-helical section of the protein. From truncation analysis it is known that both these regions of the protein are essential for function. Here we construct 31 unique cysteine substitutions in the first 42 residues of TatB. Each of the substitutions results in a TatB protein that is competent to support Tat-dependent protein translocation. Oxidant-induced disulfide cross-linking shows that both the N-terminal and amphipathic helices form contacts with at least one other TatB protomer. For the transmembrane helix these contacts are localized to one face of the helix. Molecular modeling and molecular dynamics simulations provide insight into the possible structural basis of the transmembrane helix interactions. Using variants with double cysteine substitutions in the transmembrane helix, we were able to detect cross-links between up to five TatB molecules. Protein purification showed that species containing at least four cross-linked TatB molecules are found in correctly assembled TatBC complexes. Our results suggest that the transmembrane helices of TatB protomers are in the center rather than the periphery of the TatBC complex. 相似文献