共查询到20条相似文献,搜索用时 15 毫秒
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Anshika Bajaj Nicole E. LaPlante Victoria E. Cotero Kenneth M. Fish Roger M. Bjerke Tiberiu Siclovan Cristina A. Tan Hehir 《The journal of histochemistry and cytochemistry》2013,61(1):19-30
The ability to visualize myelin is important in the diagnosis of demyelinating disordersand the detection of myelin-containing nerves during surgery. The development ofmyelin-selective imaging agents requires that a defined target for these agents beidentified and that a robust assay against the target be developed to allow for assessmentof structure-activity relationships. We describe an immunohistochemical analysis and afluorescence polarization binding assay using purified myelin basic protein (MBP) thatprovides quantitative evidence that MBP is the molecular binding partner of previouslydescribed myelin-selective fluorescent dyes such as BMB, GE3082, and GE3111. 相似文献
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Carlos E. Pedraza Christopher Taylor Albertina Pereira Michelle Seng Chui-Se Tham Michal Izrael Michael Webb 《ASN neuro》2014,6(4)
In inflammatory demyelinating diseases such as multiple sclerosis (MS), myelin
degradation results in loss of axonal function and eventual axonal degeneration.
Differentiation of resident oligodendrocyte precursor cells (OPCs) leading to
remyelination of denuded axons occurs regularly in early stages of MS but halts as
the pathology transitions into progressive MS. Pharmacological potentiation of
endogenous OPC maturation and remyelination is now recognized as a promising
therapeutic approach for MS. In this study, we analyzed the effects of modulating the
Rho-A/Rho-associated kinase (ROCK) signaling pathway, by the use of selective
inhibitors of ROCK, on the transformation of OPCs into mature, myelinating
oligodendrocytes. Here we demonstrate, with the use of cellular cultures from rodent
and human origin, that ROCK inhibition in OPCs results in a significant generation of
branches and cell processes in early differentiation stages, followed by accelerated
production of myelin protein as an indication of advanced maturation. Furthermore,
inhibition of ROCK enhanced myelin formation in cocultures of human OPCs and neurons
and remyelination in rat cerebellar tissue explants previously demyelinated with
lysolecithin. Our findings indicate that by direct inhibition of this signaling
molecule, the OPC differentiation program is activated resulting in morphological and
functional cell maturation, myelin formation, and regeneration. Altogether, we show
evidence of modulation of the Rho-A/ROCK signaling pathway as a viable target for the
induction of remyelination in demyelinating pathologies. 相似文献
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Inhibition of the Gravitropic Response of Snapdragon Spikes by
the Calcium-Channel Blocker Lanthanum Chloride 总被引:6,自引:0,他引:6 下载免费PDF全文
Haya Friedman Shimon Meir Ida Rosenberger Abraham H. Halevy Peter B. Kaufman Sonia Philosoph-Hadas 《Plant physiology》1998,118(2):483-492
The putative Ca2+-channel blocker LaCl3 prevented the gravitropic bending of cut snapdragon (Antirrhinum majus L.) spikes (S. Philosoph-Hadas, S. Meir, I. Rosenberger, A.H. Halevy [1996] Plant Physiol 110: 301–310) and inhibited stem curvature to a greater extent than vertical and horizontal stem elongation at the bending zone. This might indicate that LaCl3, which modulates cytosolic Ca2+, does not influence general stem-growth processes but may specifically affect other gravity-associated processes occurring at the stem-bending zone. Two such specific gravity-dependent events were found to occur in the bending zone of snapdragon spikes: sedimentation of starch-containing chloroplasts at the bottom of stem cortex cells, as seen in cross-sections, and establishment of an ethylene gradient across the stem. Our results show that the lateral sedimentation of chloroplasts associated with gravity sensing was prevented in cross-sections taken from the bending zone of LaCl3-treated and subsequently gravistimulated spikes and that LaCl3 completely prevented the gravity-induced, asymmetric ethylene production established across the stem-bending zone. These data indicate that LaCl3 inhibits stem curvature of snapdragon spikes by preventing several gravity-dependent processes. Therefore, we propose that the gravitropic response of shoots could be mediated through a Ca2+-dependent pathway involving modulation of cytosolic Ca2+ at various stages. 相似文献
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Abstract: [3 H]Strychnine specifically binds to membrane fractions isolated from rat retinae. The binding is saturable, with an apparent dissociation constant, K D , of 14.3 × 10−9 M and 205 fmol bound/mg protein. Specific binding is time-dependent and proportional to protein concentration. Glycine and taurine are equally potent inhibitors of [3 H]strychnine binding ( K i = 4 × 10−5 M); no other amino acids endogenously present in the retina inhibited [3 H]strychnine binding. 相似文献
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Candida albicans, a dimorphic fungus, undergoes hyphal development in response to many different environmental cues, including growth in contact with a semi-solid matrix. C. albicans forms hyphae that invade agar when cells are embedded in or grown on the surface of agar, and the integral membrane protein Dfi1p is required for this activity. In addition, Dfi1p is required for full activation of mitogen activated protein kinase Cek1p during growth on agar. In this study, we identified a putative calmodulin binding motif in the C-terminal tail of Dfi1p. This region of Dfi1p bound to calmodulin in vitro, and mutations that affected this region affected both calmodulin binding in vitro and invasive filamentation when incorporated into the full length Dfi1p protein. Moreover, increasing intracellular calcium levels led to calcium-dependent, Dfi1p-dependent Cek1p activation. We propose that conformational changes in Dfi1p in response to environmental conditions encountered during growth allow the protein to bind calmodulin and initiate a signaling cascade that activates Cek1p. 相似文献
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Insertion Mutagenesis of the Ferric Pyoverdine Receptor FpvA of Pseudomonas aeruginosa: Identification of Permissive Sites and a Region Important for Ligand Binding 下载免费PDF全文
Laurie Kilburn Keith Poole Jean-Marie Meyer Shadi Neshat 《Journal of bacteriology》1998,180(24):6753-6756
Insertion of an 18-amino-acid-encoding sequence within the fpvA gene identified permissive sites at residues Y350, A402, R451, R521, and R558, consistent with these residues occurring in extramembranous loop regions of the protein. Insertions at R451, R521, and R558 did not adversely affect receptor function, although insertions at Y350 and A402 compromised ferric pyoverdine binding and uptake. The latter region likely contributes to or interacts with the ligand-binding site. 相似文献
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血管生成素(angiogenin,ANG)调控细胞增殖、迁移、分化等生物学过程,但其作用的分子机制尚未完全明了. 目前认为,ANG可结合到rDNA区域促进rRNA转录,也可能与mRNA有结合.为全面鉴定细胞内可结合ANG的基因组序列,我们利用染色质免疫共沉淀结合DNA芯片技术(ChIP-chip)对HeLa细胞的基因组DNA进行了筛选,共获得了1 248个结合片段. 我们进一步分析了这些结合片段附近分布的基因,发现有699个可能受ANG结合调控的基因. 基因注释和聚类分析显示,这些可能受ANG调控的基因主要与肿瘤发生发展有关(特别是结直肠癌和前列腺癌),并且与TGF-β和Wnt信号通路相关. 最后,我们验证了ANG不仅与WNT6、CCNE1、APC2、FZD8和EGFR基因的启动子区域有直接结合,而且调控其表达.以上研究结果为深入研究ANG的功能机制提供了线索. 相似文献
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以拟南芥悬浮培养细胞为实验体系,借助外源荧光及同位素标记钙调素,研究结果表明外源钙调素不能被主动内吞入细胞内,而是主要以完整分子形式结合在细胞外表面;外源纯化钙调素可促进正向型质膜囊泡中的鸟苷酸三磷酸水解酶活性升高,也可引起拟南芥悬浮细胞质游离钙离子浓度的特异升高,表明外源钙调素可能通过细胞表面位点跨膜信号转换为细胞内信号,从而调节生物学活性。 相似文献
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Two Genetically Separable Phases of Growth Inhibition Induced by
Blue Light in Arabidopsis Seedlings 总被引:8,自引:3,他引:5
High fluence-rate blue light (BL) rapidly inhibits hypocotyl growth in Arabidopsis, as in other species, after a lag time of 30 s. This growth inhibition is always preceded by the activation of anion channels. The membrane depolarization that results from the activation of anion channels by BL was only 30% of the wild-type magnitude in hy4, a mutant lacking the HY4 BL receptor. High-resolution measurements of growth made with a computer-linked displacement transducer or digitized images revealed that BL caused a rapid inhibition of growth in wild-type and hy4 seedlings. This inhibition persisted in wild-type seedlings during more than 40 h of continuous BL. By contrast, hy4 escaped from the initial inhibition after approximately 1 h of BL and grew faster than wild type for approximately 30 h. Wild-type seedlings treated with 5-nitro-2-(3-phenylpropylamino)-benzoic acid, a potent blocker of the BL-activated anion channel, displayed rapid growth inhibition, but, similar to hy4, these seedlings escaped from inhibition after approximately 1 h of BL and phenocopied the mutant for at least 2.5 h. The effects of 5-nitro-2-(3-phenylpropylamino)-benzoic acid and the HY4 mutation were not additive. Taken together, the results indicate that BL acts through HY4 to activate anion channels at the plasma membrane, causing growth inhibition that begins after approximately 1 h. Neither HY4 nor anion channels appear to participate greatly in the initial phase of inhibition. 相似文献
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Analysis of proteins of the banana and citrus race of Radopholus similis was carried out by several different types of polyacrylamide gel electrophoresis. These included standard slab gel, SDS slab gel, gradient slab gel, and two-ditnensional slab gel electrophoresis. A major band difference was detected between the two races by slab gel electrophoresis. However, several other poorly resolved but consistent hands of high molecular weight proteins near the gel origin also were considered as diagnostic. Resolution of protein bands was greatly improved by SDS and gradient slab gel electrophoresis, but no differences could be detected among the proteins resolved between the two rares with these techniques. Two-dimensional gels revealed a large number of proteins, but background staining obscured them hindering interpretation. When nematode races were reared on three different host plants, no differences in protein patterns were detected between them, indicating host preferences does not play a role in determining the types proteins occurring in these nematodes. 相似文献
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Qinli Wang Bo Chen Peng Liu Maozhong Zheng Yuqing Wang Sujuan Cui Daye Sun Xiaohong Fang Chun-Ming Liu William J. Lucas Jinxing Lin 《The Journal of biological chemistry》2009,284(18):12000-12007
Calmodulin (CaM) is a highly conserved intracellular calcium sensor. In
plants, CaM also appears to be present in the apoplasm, and application of
exogenous CaM has been shown to influence a number of physiological functions
as a polypeptide signal; however, the existence and localization of its
corresponding apoplasmic binding sites remain controversial. To identify the
site(s) of action, a CaM-conjugated quantum dot (QD) system was employed for
single molecule level detection at the surface of plant cells. Using this
approach, we show that QD-CaM binds selectively to sites on the outer surface
of the plasma membrane, which was further confirmed by high resolution
transmission electron microscopy. Measurements of Ca2+ fluxes
across the plasma membrane, using ion-selective microelectrodes, demonstrated
that exogenous CaM induces a net influx into protoplasts. Consistent with
these flux studies, calcium-green-dextran and FRET experiments confirmed that
applied CaM/QD-CaM elicited an increase in cytoplasmic Ca2+ levels.
These results support the hypothesis that apoplasmic CaM can act as a
signaling agent. These findings are discussed in terms of CaM acting as an
apoplasmic peptide ligand to mediate transmembrane signaling in the plant
kingdom.Calmodulin (CaM)2
is a conserved multifunctional calcium sensor that mediates intracellular
Ca2+ signaling and regulates diverse cellular processes by
interacting with calmodulin-binding proteins
(1–3).
Interestingly, in both animals and plants, CaM may also act as an
extracellular agent to regulate physiological events
(4). Consistent with this
notion, extracellular CaM has been detected within the cell walls of a broad
range of plant species (4,
5).Functional studies have established that exogenously applied CaM can
stimulate the proliferation of suspension-cultured plant cells
(6) as well as affect
intracellular activities of heterotrimeric G proteins and phospholipases in
protoplasts (7,
8). Based on these findings, it
has been proposed that, in plants, extracellular CaM may function as a
signaling agent involved in the regulation of cell growth and development
(4). However, as a 17-kDa
hydrophilic protein, exogenously applied CaM could well be retrieved from the
apoplasmic space and then exert its effects on components within the
cytoplasm. Evidence against this hypothesis was provided by studies with
Arabidopsis thaliana suspension-cultured cells in which it was shown
that 24 h of incubation in exogenous CaM did not result in protein uptake or
degradation (4).To exert an effect from the apoplasm, it would seem logical to assume that
a protein(s) within the plant plasma membrane would have a CaM-binding site
exposed to the apoplasm. Although a number of studies have addressed the
molecular mechanism(s) by which extracellular CaM might act as a signal
(6,
9) and attempts have been made
to identify extracellular CaM-binding proteins
(4,
6), currently there is no
direct evidence in support of the hypothesis that specific CaM-binding sites
exist at the surface of plant cells.To address this question, a CaM-conjugated quantum dot (QD) system was
employed for single molecule level detection
(10–13)
at the surface of plant cells. These nanoparticles have several advantages
over conventional fluorophores for light microscopic imaging, including their
higher brightness and photostability
(14,
15). In addition, because of
their electron dense nature, QDs can be used for single labeling studies at
the transmission electron microscope level
(16,
17). Using this QD-CaM system,
we demonstrate that QD-CaM binds selectively to sites on the outer surface of
the plant plasma membrane. We also show by three independent methods that
applied CaM can modulate Ca2+ fluxes across the plasma membrane,
leading to alterations in cytoplasmic Ca2+ status. These findings
support the hypothesis that, in plants, apoplasmic CaM can act as a signaling
agent. 相似文献