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The synthesis of ribosomes in HeLa cells was studied during recovery from a 20-hour deprivation for valine. The rates of incorporation of labeled precursors into ribosomal pre-RNA, processed rRNA, total cellular proteins, and proteins of the 60S ribosomal subunit returned to normal or nearly normal levels immediately after restoration of valine to the medium. Specific proteins of the 60S ribosomal subunit, whose apparent net synthesis is reduced more than that of the other proteins of the 60S ribosomal subunit during valine deprivation, were no longer undersynthesized after valine was restored. This rapid recovery suggests that the apparent decrease in the net rate of synthesis of these ribosomal proteins during valine deprivation is effected at the translational or post-translational level. No evidence of significant synchrony in any particular stage of the cell cycle was observed after a 20-hr valine deprivation. Key words: 60S ribosomal subunit; HeLa, cells; valine deprivation. 相似文献
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Mary McMahon Verónica Ayllón Kostya I. Panov Rosemary O'Connor 《The Journal of biological chemistry》2010,285(24):18309-18318
Insulin-like growth factor-I (IGF-I) signaling is strongly associated with cell growth and regulates the rate of synthesis of the rRNA precursor, the first and the key stage of ribosome biogenesis. In a screen for mediators of IGF-I signaling in cancer, we recently identified several ribosome-related proteins, including NEP1 (nucleolar essential protein 1) and WDR3 (WD repeat 3), whose homologues in yeast function in ribosome processing. The WDR3 gene and its locus on chromosome 1p12-13 have previously been linked with malignancy. Here we show that IGF-I induces expression of WDR3 in transformed cells. WDR3 depletion causes defects in ribosome biogenesis by affecting 18 S rRNA processing and also causes a transient down-regulation of precursor rRNA levels with moderate repression of RNA polymerase I activity. Suppression of WDR3 in cells expressing functional p53 reduced proliferation and arrested cells in the G1 phase of the cell cycle. This was associated with activation of p53 and sequestration of MDM2 by ribosomal protein L11. Cells lacking functional p53 did not undergo cell cycle arrest upon suppression of WDR3. Overall, the data indicate that WDR3 has an essential function in 40 S ribosomal subunit synthesis and in ribosomal stress signaling to p53-mediated regulation of cell cycle progression in cancer cells. 相似文献
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Physarum polycephalum has been used as a model system to study the phosphorylation of ribosomal proteins during the cell cycle. The results showed that the phosphate content of S3, the major ribosomal phosphoprotein in this organism, was constant during all phases of the cell cycle. No additional ribosomal phosphoproteins were observed. These results differ significantly from those reported earlier by Rupp, R.G., Humphrey, R.M. and Shaeffer, J.R. (Biochim. Biophys. Acta (1976) 418, 81-92) and suggest that the use of thymidine or hydroxyurea to synchronize cell population may affect the phosphorylation of ribosomal proteins. The results are discussed in relation to protein synthesis and cAMP level during the cell cycle. 相似文献
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Nucleophosmin serves as a rate-limiting nuclear export chaperone for the Mammalian ribosome 总被引:1,自引:0,他引:1
Maggi LB Kuchenruether M Dadey DY Schwope RM Grisendi S Townsend RR Pandolfi PP Weber JD 《Molecular and cellular biology》2008,28(23):7050-7065
Nucleophosmin (NPM) (B23) is an essential protein in mouse development and cell growth; however, it has been assigned numerous roles in very diverse cellular processes. Here, we present a unified mechanism for NPM's role in cell growth; NPM directs the nuclear export of both 40S and 60S ribosomal subunits. NPM interacts with rRNA and large and small ribosomal subunit proteins and also colocalizes with large and small ribosomal subunit proteins in the nucleolus, nucleus, and cytoplasm. The transduction of NPM shuttling-defective mutants or the loss of Npm1 inhibited the nuclear export of both the 40S and 60S ribosomal subunits, reduced the available pool of cytoplasmic polysomes, and diminished overall protein synthesis without affecting rRNA processing or ribosome assembly. While the inhibition of NPM shuttling can block cellular proliferation, the dramatic effects on ribosome export occur prior to cell cycle inhibition. Modest increases in NPM expression amplified the export of newly synthesized rRNAs, resulting in increased rates of protein synthesis and indicating that NPM is rate limiting in this pathway. These results support the idea that NPM-regulated ribosome export is a fundamental process in cell growth. 相似文献
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Ribosome synthesis involves the concomitance of pre-rRNA processing and ribosomal protein assembly. In eukaryotes, this is a complex process that requires the participation of specific sequences and structures within the pre-rRNAs, at least 200 trans-acting factors and the ribosomal proteins. There is little information on the function of individual 60S ribosomal proteins in ribosome synthesis. Herein, we have analysed the contribution of ribosomal protein L35 in ribosome biogenesis. In vivo depletion of L35 results in a deficit in 60S ribosomal subunits and the appearance of half-mer polysomes. Pulse-chase, northern hybridization and primer extension analyses show that processing of the 27SB to 7S pre-rRNAs is strongly delayed upon L35 depletion. Most likely as a consequence of this, release of pre-60S ribosomal particles from the nucleolus to the nucleoplasm is also blocked. Deletion of RPL35A leads to similar although less pronounced phenotypes. Moreover, we show that L35 assembles in the nucleolus and binds to early pre-60S ribosomal particles. Finally, flow cytometry analysis indicated that L35-depleted cells mildly delay the G1 phase of the cell cycle. We conclude that L35 assembly is a prerequisite for the efficient cleavage of the internal transcribed spacer 2 at site C2. 相似文献
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Synthesis of ribosomal proteins during the cell cycle of the yeast Saccharomyces cerevisiae.
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Centrifugal elutriation was used to separate yeast cells by their cell cycle position. The rate of synthesis of ribosomal proteins showed a constant exponential increase through the cell cycle. 相似文献
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The absolute rates of synthesis of specific ribosomal proteins have been determined during growth and meiotic maturation of mouse oocytes, as well as during early embryogenesis in the mouse. These measurements were made possible by the development of a high-resolution twodimensional gel electrophoresis procedure capable of resolving basic proteins with isoelectric points between 9.1 and 10.2. Mouse ribosomal proteins were separated on such gels and observed rates of incorporation of [35S]methionine into each of 12 representative ribosomal proteins were converted into absolute rates of synthesis (femtograms or moles synthesized/hour/oocyte or embryo) by using previously determined values for the absolute rates of total protein synthesis in mouse oocytes and embryos (R. M. Schultz, M. J. LaMarca, and P. M. Wassarman, 1978,Proc. Nat. Acad. Sci. USA,75, 4160;R. M. Schultz, G. E. Letourneau, and P. M. Wassarman, 1979,Develop. Biol.,68, 341–359). Ribosomal proteins were synthesized at all stages of oogenesis and early embryogenesis examined and, while equimolar amounts of ribosomal proteins were found in ribosomes, they were always synthesized in nonequimolar amounts during development. Rates of synthesis of individual ribosomal proteins differed from each other by more than an order of magnitude in some cases. Synthesis of ribosomal proteins accounted for 1.5, 1.5, and 1.1% of total protein synthesis during growth of the oocyte, in the fully grown oocyte, and in the unfertilized egg, respectively. During meiotic maturation of mouse oocytes the absolute rate of synthesis of ribosomal proteins decreased about 40%, from 620 to 370 fg/hr/cell, as compared to a 23% decrease in the rate of total protein synthesis during the same period. On the other hand, during early embryogenesis the absolute rates of synthesis of each of the 12 ribosomal proteins examined increased substantially as compared with those of the unfertilized egg, such that at the eight-cell stage of embryogenesis synthesis of ribosomal proteins (4.17 pg/hr/embryo) accounted for about 8.1% of the total protein synthesis in the embryo. Consequently, while the absolute rate of total protein synthesis increased about 1.5-fold during development from an unfertilized mouse egg to an eight-cell compacted embryo, the absolute rate of ribosomal protein synthesis increased more than 11-fold during the same period. These results seem to reflect the differences reported for the patterns of ribosomal RNA synthesis during early development of mammalian, as compared to nonmammalian, animal species. The results are compared with those obtained using oocytes and embryos fromXenopus laevis. 相似文献
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Synthesis of soluble protein during the cell cycle of the fission yeast Schizosaccharomyces pombe 总被引:2,自引:0,他引:2
W H Wain 《Experimental cell research》1971,69(1):49-56
Many enzymes show a pattern of increase in activity through the cell cycle which is different from the continuous exponential pattern of total protein synthesis. A group of proteins at an intermediate level between single enzymes and total protein, the soluble proteins, was examined to resolve this anomaly. The synthesis of the pH 8.1 soluble proteins of Schizosaccharomyces pombe through the cell cycle was followed by pulse labelling with 3H-leucine in synchronous cultures. The soluble proteins were analysed by electrophoresis on acrylamide gels. Soluble proteins represent 30% of the total proteins of S. pombe and the rates of synthesis showed a continuous increase through the cell cycle. Individual groups of proteins, represented by a single band after electrophoresis, showed a similar continuous increase in synthesis through the cell cycle. Any proteins which may be synthesised discontinuously, such as some enzymes, represent such a small proportion of any one protein group in the electrophoretic separation that their effect was not detectable. These results are different from those described for mammalian cells. 相似文献
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Fernando Gómez-Herreros Olga Rodríguez-Galán Macarena Morillo-Huesca Douglas Maya María Arista-Romero Jesús de la Cruz Sebastián Chávez Mari Cruz Mu?oz-Centeno 《The Journal of biological chemistry》2013,288(44):31689-31700
Cell cycle regulation is a very accurate process that ensures cell viability and the genomic integrity of daughter cells. A fundamental part of this regulation consists in the arrest of the cycle at particular points to ensure the completion of a previous event, to repair cellular damage, or to avoid progression in potentially risky situations. In this work, we demonstrate that a reduction in nucleotide levels or the depletion of RNA polymerase I or III subunits generates a cell cycle delay at the G1/S transition in Saccharomyces cerevisiae. This delay is concomitant with an imbalance between ribosomal RNAs and proteins which, among others, provokes an accumulation of free ribosomal protein L5. Consistently with a direct impact of free L5 on the G1/S transition, rrs1 mutants, which weaken the assembly of L5 and L11 on pre-60S ribosomal particles, enhance both the G1/S delay and the accumulation of free ribosomal protein L5. We propose the existence of a surveillance mechanism that couples the balanced production of yeast ribosomal components and cell cycle progression through the accumulation of free ribosomal proteins. This regulatory pathway resembles the p53-dependent nucleolar-stress checkpoint response described in human cells, which indicates that this is a general control strategy extended throughout eukaryotes. 相似文献
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Stimulation of RNA synthesis at initiation by ribosomal proteins 总被引:2,自引:0,他引:2
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《Insect Biochemistry》1985,15(3):391-401
Stimulation of the accessory gland of Drosophila melanogaster males by copulation was studied at the molecular level by in vivo radiolabelling experiments. In addition, experiments were carried out to clarify the molecular basis of regulation of the gland's synthetic activityThe stimulatory effect of copulation on secretion protein synthesis is observed within minutes after copulation is terminated. Repeated copulations elicit a stronger response. The data indicate a coordinate synthesis of the secretory proteins.In a reticulocyte-lysate system with added total RNA extract from male accessory glands, secretion proteins appear as translation products. It is shown that synthesis of secretory protein messages is not enhanced by copulation and that the messages are stable.Synthesis of ribosomal proteins and ribosomal RNA increases upon copulation. The increase is observed in the time period from 1 to 5 hr after copulation. Again, the synthesis of the various ribosomal proteins is well coordinated. There is no elevation in the accumulation of ribosomal protein mRNAs after copulation. 相似文献
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THE INTRACELLULAR SITE OF SYNTHESIS OF MITOCHONDRIAL RIBOSOMAL PROTEINS IN NEUROSPORA CRASSA 总被引:8,自引:3,他引:5
The intracellular site of synthesis of mitochondrial ribosomal proteins (MRP) in Neurospora crassa has been investigated using three complementary approaches. (a) Mitochondrial protein synthesis in vitro: Tritium-labeled proteins made by isolated mitochondria were compared to 14C-labeled marker MRP by cofractionation in a two-step procedure involving isoelectric focusing and polyacrylamide gel electrophoresis. Examination of the electrophoretic profiles showed that essentially none of the peaks of in vitro product corresponded exactly to any of the MRP marker peaks. (b) Sensitivity of in vivo MRP synthesis to chloramphenicol: Cells were labeled with leucine-3H in the presence of chloramphenicol, mitochondrial ribosomal subunits were subsequently isolated, and their proteins fractionated by isoelectric focusing followed by gel electrophoresis. The labeling of every single MRP was found to be insensitive to chloramphenicol, a selective inhibitor of mitochondrial protein synthesis. (c) Sensitivity of in vivo MRP synthesis to anisomycin: We have found this antibiotic to be a good selective inhibitor of cytoplasmic protein synthesis in Neurospora. In the presence of anisomycin the labeling of virtually all MRP is inhibited to the same extent as the labeling of cytoplasmic ribosomal proteins. On the basis of these three types of studies we conclude that most if not all 53 structural proteins of mitochondrial ribosomal subunits in Neurospora are synthesized by cytoplasmic ribosomes. 相似文献
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Monoiodoamphenicol inhibits protein synthesis in exponentially growing Escherichia coli without measurable delay. The inhibition follows first-order kinetics suggesting that one molecule of monoiodoamphenicol is sufficient to inhibit one ribosome. However, only 60% of the ribosomes were susceptible to irreversible inhibition by monoiodoamphenicol. The residual 40% of the rate of protein synthesis after monoiodoamphenicol treatment was not significantly different, when the time of treatment was increased up to six hours or the drug concentration was raised to 1000 μg/ml.Analysis of the monoiodoamphenicol-labeled cell proteins by various uni- and two-dimensional gel electrophoreses and by sucrose gradient centrifugation shows that the cell target of the covalent labeling reaction of monoiodoamphenicol is almost exclusively the ribosome. The amount of label incorporated into ribosomal protein is proportional to the degree of inhibition of protein synthesis by monoiodoamphenicol. However, more than one ribosomal protein reacted. The predominantly labeled proteins are S6, L16 and L24. It is proposed that all three proteins are associated with the receptor site of monoiodoamphenicol. 相似文献
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Angelo Gismondi Sara Caldarola Gaia Lisi Giada Juli Lidia Chellini Valentina Iadevaia Christopher G. Proud Fabrizio Loreni 《Nucleic acids research》2014,42(20):12668-12680
The synthesis of adequate amounts of ribosomes is an essential task for the cell. It is therefore not surprising that regulatory circuits exist to organize the synthesis of ribosomal components. It has been shown that defect in ribosome biogenesis (ribosomal stress) induces apoptosis or cell cycle arrest through activation of the tumor suppressor p53. This mechanism is thought to be implicated in the pathophysiology of a group of genetic diseases such as Diamond Blackfan Anemia which are called ribosomopathies. We have identified an additional response to ribosomal stress that includes the activation of eukaryotic translation elongation factor 2 kinase with a consequent inhibition of translation elongation. This leads to a translational reprogramming in the cell that involves the structurally defined group of messengers called terminal oligopyrimidine (TOP) mRNAs which encode ribosomal proteins and translation factors. In fact, while general protein synthesis is decreased by the impairment of elongation, TOP mRNAs are recruited on polysomes causing a relative increase in the synthesis of TOP mRNA-encoded proteins compared to other proteins. Therefore, in response to ribosomal stress, there is a change in the translation pattern of the cell which may help restore a sufficient level of ribosomes. 相似文献
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Anna Aspesi Elisa Pavesi Elisa Robotti Rossella Crescitelli Ilenia Boria Federica Avondo Hélène Moniz Lydie Da Costa Narla Mohandas Paola Roncaglia Ugo Ramenghi Antonella Ronchi Stefano Gustincich Simone Merlin Emilio Marengo Steven R. Ellis Antonia Follenzi Claudio Santoro Irma Dianzani 《Gene》2014
Defects in genes encoding ribosomal proteins cause Diamond Blackfan Anemia (DBA), a red cell aplasia often associated with physical abnormalities. Other bone marrow failure syndromes have been attributed to defects in ribosomal components but the link between erythropoiesis and the ribosome remains to be fully defined. Several lines of evidence suggest that defects in ribosome synthesis lead to “ribosomal stress” with p53 activation and either cell cycle arrest or induction of apoptosis. Pathways independent of p53 have also been proposed to play a role in DBA pathogenesis. 相似文献