共查询到20条相似文献，搜索用时 170 毫秒
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Heterogeneous nuclear ribonucleoproteins: role in RNA splicing 总被引：107，自引：0，他引：107
Splicing in vitro of a messenger RNA (mRNA) precursor (pre-mRNA) is inhibited by a monoclonal antibody to the C proteins (anti-C) of the heterogeneous nuclear RNA (hnRNA)-ribonucleoprotein (hnRNP) particles. This antibody, 4F4, inhibits an early step of the reaction: cleavage at the 3' end of the upstream exon and the formation of the intron lariat. In contrast, boiled 4F4, or a different monoclonal antibody (designated 2B12) to the C proteins, or antibodies to other hnRNP proteins (120 and 68 kilodaltons) and nonimmune mouse antibodies have no inhibitory effect. The 4F4 antibody does not prevent the adenosine triphosphate-dependent formation of a 60S splicing complex (spliceosome). Furthermore, the 60S splicing complex contains C proteins, and it can be immunoprecipitated with 4F4. Depletion of C proteins from the splicing extract by immunoadsorption with either of the two monoclonal antibodies to the C proteins (4F4 or 2B12) results in the loss of splicing activity, whereas mock-depletion with nonimmune mouse antibodies bodies has no effect. A 60S splicing complex does not form in a C protein-depleted nuclear extract. These results indicate an essential role for proteins of the hnRNP complex in the splicing of mRNA precursors. 相似文献
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Translation of unspliced transcripts after heat shock 总被引：19，自引：0，他引：19
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Regulated splicing of the Drosophila P transposable element third intron in vitro: somatic repression 总被引：35，自引：0，他引：35
In eukaryotic cells alternative splicing of messenger RNA precursors (pre-mRNA's) is a means of regulating gene expression. Although a number of the components that participate in regulating some alternative splicing events have been identified by molecular genetic procedures, the elucidation of the biochemical mechanisms governing alternative splicing requires in vitro reaction systems. The tissue specificity of P element transposition in Drosophila depends on the germline restriction of pre-mRNA splicing of the P element third intron (IVS3). Drosophila P element IVS3 pre-mRNA substrates were spliced accurately in vitro in heterologous human cell extracts but not in Drosophila somatic cell splicing extracts. Components in Drosophila somatic cell extracts that specifically inhibited IVS3 splicing in vitro were detected by a complementation assay. Biochemical assays for Drosophila RNA binding proteins were then used to detect a 97-kilodalton protein that interacts specifically with 5' exon sequences previously implicated in the control of IVS3 splicing in vivo. Inhibition of IVS3 splicing in vitro could be correlated with binding of the 97-kD protein to 5' exon sequences, suggesting that one aspect of IVS3 tissue-specific splicing involves somatic repression by specific RNA-protein interactions. 相似文献
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Human immunodeficiency virus type 1 (HIV-1), in contrast with most other retroviruses, encodes trans-regulatory proteins for virus gene expression. It is shown in this study, by means of an in vitro splicing system, that nuclear extracts obtained from cells infected with HIV-1 contain a factor (or factors) that specifically inhibits splicing of a synthetic SP6/HIV pre-messenger RNA (pre-mRNA)-containing donor and acceptor splice sites in the coding region for the envelope protein. It is also shown that the SP6/HIV pre-mRNA is not capable of assembly in a ribonucleoprotein complex, spliceosome, in extracts from infected cells. These findings raise the possibility that specific inhibition of pre-mRNA splicing in the envelope protein coding region by HIV-1 trans-regulatory factors might be one control mechanism for efficient production of structural viral proteins and virion assembly. 相似文献
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Axonal proteins of presynaptic neurons during synaptogenesis 总被引：5，自引：0，他引：5
P Sonderegger M C Fishman M Bokoum H C Bauer P G Nelson 《Science (New York, N.Y.)》1983,221(4617):1294-1297
Changes occur in the synthesis and axonal transport of neuronal proteins in dorsal-root ganglia axons as a result of contact with cells from the spinal cord during synapse formation. Dorsal-root ganglia cells were cultured in a compartmental cel culture system that allows separate access to neuronal cell bodies and their axons. When cells from the ventral spinal cord were cultured with the dorsal-root ganglia axons, synapses were established within a few days. Metabolic labeling and two-dimensional electrophoresis revealed that four of more than 300 axonal proteins had changed in their expression by the time synapses were established. The highly selective nature of these changes suggests that the proteins involved may be important in the processes of axon growth and synapse formation and their regulation by the regional environment. 相似文献
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A broad range of neurodegenerative disorders is characterized by neuronal damage that may be caused by toxic, aggregation-prone proteins. As genes are identified for these disorders and cell culture and animal models are developed, it has become clear that a major effect of mutations in these genes is the abnormal processing and accumulation of misfolded protein in neuronal inclusions and plaques. Increased understanding of the cellular mechanisms for disposal of abnormal proteins and of the effects of toxic protein accumulation on neuronal survival may allow the development of rational, effective treatment for these disorders. 相似文献
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Affinity chromatography of splicing complexes: U2, U5, and U4 + U6 small nuclear ribonucleoprotein particles in the spliceosome 总被引：69，自引：0，他引：69
The splicing process, which removes intervening sequences from messenger RNA (mRNA) precursors is essential to gene expression in eukaryotic cells. This site-specific process requires precise sequence recognition at the boundaries of an intervening sequence, but the mechanism of this recognition is not understood. The splicing of mRNA precursors occurs in a multicomponent complex termed the spliceosome. Such an assembly of components is likely to play a key role in specifying those sequences to be spliced. In order to analyze spliceosome structure, a stringent approach was developed to obtain splicing complexes free of cellular contaminants. This approach is a form of affinity chromatography based on the high specificity of the biotin-streptavidin interaction. A minimum of three subunits: U2, U5, and U4 + U6 small nuclear ribonucleoprotein particles were identified in the 35S spliceosome structure, which also contains the bipartite RNA intermediate of splicing. A 25S presplicing complex contained only the U2 particle. The multiple subunit structure of the spliceosome has implications for the regulation of a splicing event and for its possible catalysis by ribozyme or ribozymes. 相似文献
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Neuronal generation of the leech swimming movement 总被引：5，自引：0，他引：5
G S Stent W B Kristan W O Friesen C A Ort M Poon R L Calabrese 《Science (New York, N.Y.)》1978,200(4348):1348-1357
The swimming movement of the leech is produced by an ensemble of bilaterally symmetric, rhythmically active pairs of motor neurons present in each segmental ganglion of the ventral nerve cord. These motor neurons innervate the longitudinal muscles in dorsal or ventral sectors of the segmental body wall. Their duty cycles are phase-locked in a manner such that the dorsal and ventral body wall sectors of any given segment undergo an antiphasic contractile rhythm and that the contractile rhythms of different segments form a rostrocaudal phase progression. This activity rhythm is imposed on the motor neurons by a central swim oscillator, of which four bilaterally symmetric pairs of interneurons present in each segmental ganglion appear to constitute the major component. These interneurons are linked intra- and intersegmentally via inhibitory connections to form a segmentally iterated and inter-segmentally concatenated cyclic neuronal network. The network appears to owe its oscillatory activity pattern to the mechanism of recurrent cyclic inhibition. 相似文献
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Two domains of yeast U6 small nuclear RNA required for both steps of nuclear precursor messenger RNA splicing 总被引：31，自引：0，他引：31
U6 is one of the five small nuclear RNA's (snRNA's) that are required for splicing of nuclear precursor messenger RNA (pre-mRNA). The size and sequence of U6 RNA are conserved among organisms as diverse as yeast and man, and so it has been proposed that U6 RNA functions as a catalytic element in splicing. A procedure for in vitro reconstitution of functional yeast U6 small nuclear ribonucleoproteins (snRNP's) with synthetic U6 RNA was applied in an attempt to elucidate the function of yeast U6 RNA. Two domains in U6 RNA were identified, each of which is required for in vitro splicing. Single nucleotide substitutions in these two domains block splicing either at the first or the second step. Invariably, U6 RNA mutants that block the first step of splicing do not enter the spliceosome. On the other hand, those that block the second step of splicing form a spliceosome but block cleavage at the 3' splice site of the intron. In both domains, the positions of base changes that block the second step of splicing correspond exactly to the site of insertion of pre-mRNA-type introns into the U6 gene of two yeast species, providing a possible explanation for the mechanism of how these introns originated and adding further evidence for the proposed catalytic role of U6 RNA. 相似文献
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Popow J Englert M Weitzer S Schleiffer A Mierzwa B Mechtler K Trowitzsch S Will CL Lührmann R Söll D Martinez J 《Science (New York, N.Y.)》2011,331(6018):760-764
Splicing of mammalian precursor transfer RNA (tRNA) molecules involves two enzymatic steps. First, intron removal by the tRNA splicing endonuclease generates separate 5' and 3' exons. In animals, the second step predominantly entails direct exon ligation by an elusive RNA ligase. Using activity-guided purification of tRNA ligase from HeLa cell extracts, we identified HSPC117, a member of the UPF0027 (RtcB) family, as the essential subunit of a tRNA ligase complex. RNA interference-mediated depletion of HSPC117 inhibited maturation of intron-containing pre-tRNA both in vitro and in living cells. The high sequence conservation of HSPC117/RtcB proteins is suggestive of RNA ligase roles of this protein family in various organisms. 相似文献
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A specific amino acid binding site composed of RNA 总被引：20，自引：0，他引：20
M Yarus 《Science (New York, N.Y.)》1988,240(4860):1751-1758
A specific, reversible binding site for a free amino acid is detectable on the intron of the Tetrahymena self-splicing ribosomal precursor RNA. The site selects arginine among the natural amino acids, and prefers the L- to the D-amino acid. The dissociation constant is in the millimolar range, and amino acid binding is at or in the catalytic rG splicing substrate site. Occupation of the G site by L-arginine therefore inhibits splicing by inhibiting the binding of rG, without inhibition of later reactions in the splicing reaction sequence. Arginine binding specificity seems to be directed at the side chain and the guanidino radical, and the alpha-amino and carboxyl groups are dispensable for binding. The arginine site can be placed within the G site by structural homology, with consequent implications for RNA-amino acid interaction, for the origin of the genetic code, for control of RNA activities, and for further catalytic capabilities for RNA. 相似文献
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[目的]为了寻求采用响应面分析法提取仙人掌多糖的最佳工艺,为其开发利用提供可用数据。[方法]在单因素试验的基础上,根据Box-Benhnken的中心组合试验设计原理,选取提取温度、提取时间和水料比对仙人掌多糖提取影响显著的三个因素,采用三因素三水平的响应面分析方法,对仙人掌茎多糖提取条件进行优化。[结果]响应面分析结果表明,提取温度、提取时间以及水料比与仙人掌茎多糖多糖得率存在着显著的相关性,仙人掌茎多糖水提的最佳提取工艺条件为:提取温度86.1℃、时间提取时间3.61 h、水料比3.72∶1,仙人掌茎多糖的得率达到理论值0.694%。[结论]采用响应面分析法优化工艺得到的提取条件参数可信,具有可行性与应用价值。 相似文献
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Technologies to assess the molecular targets of biomolecules in living cells are lacking. We have developed a technology called magnetism-based interaction capture (MAGIC) that identifies molecular targets on the basis of induced movement of superparamagnetic nanoparticles inside living cells. Efficient intracellular uptake of superparamagnetic nanoparticles (coated with a small molecule of interest) was mediated by a transducible fusogenic peptide. These nanoprobes captured the small molecule's labeled target protein and were translocated in a direction specified by the magnetic field. Use of MAGIC in genome-wide expression screening identified multiple protein targets of a drug. MAGIC was also used to monitor signal-dependent modification and multiple interactions of proteins. 相似文献
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Edery P Marcaillou C Sahbatou M Labalme A Chastang J Touraine R Tubacher E Senni F Bober MB Nampoothiri S Jouk PS Steichen E Berland S Toutain A Wise CA Sanlaville D Rousseau F Clerget-Darpoux F Leutenegger AL 《Science (New York, N.Y.)》2011,332(6026):240-243
The spliceosome, a ribonucleoprotein complex that includes proteins and small nuclear RNAs (snRNAs), catalyzes RNA splicing through intron excision and exon ligation to produce mature messenger RNAs, which, in turn serve as templates for protein translation. We identified four point mutations in the U4atac snRNA component of the minor spliceosome in patients with brain and bone malformations and unexplained postnatal death [microcephalic osteodysplastic primordial dwarfism type 1 (MOPD 1) or Taybi-Linder syndrome (TALS); Mendelian Inheritance in Man ID no. 210710]. Expression of a subgroup of genes, possibly linked to the disease phenotype, and minor intron splicing were affected in cell lines derived from TALS patients. Our findings demonstrate a crucial role of the minor spliceosome component U4atac snRNA in early human development and postnatal survival. 相似文献
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Stokin GB Lillo C Falzone TL Brusch RG Rockenstein E Mount SL Raman R Davies P Masliah E Williams DS Goldstein LS 《Science (New York, N.Y.)》2005,307(5713):1282-1288
We identified axonal defects in mouse models of Alzheimer's disease that preceded known disease-related pathology by more than a year; we observed similar axonal defects in the early stages of Alzheimer's disease in humans. Axonal defects consisted of swellings that accumulated abnormal amounts of microtubule-associated and molecular motor proteins, organelles, and vesicles. Impairing axonal transport by reducing the dosage of a kinesin molecular motor protein enhanced the frequency of axonal defects and increased amyloid-beta peptide levels and amyloid deposition. Reductions in microtubule-dependent transport may stimulate proteolytic processing of beta-amyloid precursor protein, resulting in the development of senile plaques and Alzheimer's disease. 相似文献
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基因组水平上拟南芥和水稻mrs2基因家族的比较系统发生分析 总被引：1，自引：0，他引：1
mrs2（mitochondrial RNA splicing2）基因是植物线粒体中Ⅱ类内含子自我剪接缺陷的抑制基因，同时参与了植物中镁离子的运输。本研究利用已经分离的植物的mrs2基因，鉴别出MRS2结构域，同时对拟南芥和水稻中的mrs2基因家族的成员进行了鉴定；利用这些基因编码的蛋白质序列构建了系统发生树，并进行了序列保守性分析，最后查找了相关基因的EST表达信息。结果表明：①系统发生分析表明拟南芥和水稻的mrs2基因的结构在拟南芥和水稻分离之前已经形成，并在分离之后按照物种特异性的方式进行了扩张；②MEME分析表明植物的Mrs2蛋白质具有高度保守的基序，并且在蛋白质中的排列顺序也大致相似；③mrs2基因在拟南芥和水稻中的表达有差异，但在部分表达上仍保持了一致性。 相似文献
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Fairman ME Maroney PA Wang W Bowers HA Gollnick P Nilsen TW Jankowsky E 《Science (New York, N.Y.)》2004,304(5671):730-734
Members of the DExH/D superfamily of nucleic acid-activated nucleotide triphosphatases are essential for virtually all aspects of RNA metabolism, including pre-messenger RNA splicing, RNA interference, translation, and nucleocytoplasmic trafficking. Physiological substrates for these enzymes are thought to be regions of double-stranded RNA, because several DExH/D proteins catalyze strand separation in vitro. These "RNA helicases" can also disrupt RNA-protein interactions, but it is unclear whether this activity is coupled to duplex unwinding. Here we demonstrate that two unrelated DExH/D proteins catalyze protein displacement independently of duplex unwinding. Therefore, the essential functions of DExH/D proteins are not confined to RNA duplexes but can be exerted on a wide range of ribonucleoprotein substrates. 相似文献