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At least 325 records · Page 18Linked to original sources

PubMed1979

Translational and post-translational cleavage of M13 procoat protein: extracts of both the cytoplasmic and outer membranes of Escherichia coli contain leader peptidase activity.

The coat protein of coliphage M13 is an integral protein of the host cytoplasmic membrane at all stages of the infectious cycle. Both in in vivo and DNA-directed in vitro synthesis, it is initially made with an NH2-terminal "leader peptide" of 23 amino acids and is termed procoat. We now report that leader peptidase, and activity which removes the leader peptide and converts procoat to coat, is found in both the inner (cytoplasmic) and outer membrane of Escherichia coli. However, only cytoplasmic membranes will catalyze cleavage of procoat in the absence of detergent. Leader peptidase will cleave procoat either during translation or after protein synthesis is complete.

Cell Membrane↗

PubMed1999

Universal conservation in translation initiation revealed by human and archaeal homologs of bacterial translation initiation factor IF2.

Binding of initiator methionyl-tRNA to ribosomes is catalyzed in prokaryotes by initiation factor (IF) IF2 and in eukaryotes by eIF2. The discovery of both IF2 and eIF2 homologs in yeast and archaea suggested that these microbes possess an evolutionarily intermediate protein synthesis apparatus. We describe the identification of a human IF2 homolog, and we demonstrate by using in vivo and in vitro assays that human IF2 functions as a translation factor. In addition, we show that archaea IF2 can substitute for its yeast homolog both in vivo and in vitro. We propose a universally conserved function for IF2 in facilitating the proper binding of initiator methionyl-tRNA to the ribosomal P site.

Amino Acid Sequence↗

PubMed2004

Ribosome stalling during translation elongation induces cleavage of mRNA being translated in Escherichia coli.

Recently, it has been found that ribosome pausing at stop codons caused by certain nascent peptides induces cleavage of mRNA in Escherichia coli cells (1, 2). The question we addressed in the present study is whether mRNA cleavage occurs when translation elongation is prevented. We focused on a specific peptide sequence (AS17), derived from SecM, that is known to cause elongation arrest. When the crp-crr fusion gene encoding CRP-AS17-IIA(Glc) was expressed, cAMP receptor protein (CRP) proteins truncated around the arrest sequence were efficiently produced, and they were tagged by the transfer-messenger RNA (tmRNA) system. Northern blot analysis revealed that both truncated upstream crp and downstream crr mRNAs were generated along with reduced amounts of the full-length crp-crr mRNA. The truncated crp mRNA dramatically decreased in the presence of tmRNA due to rapid degradation. The 3' ends of truncated crp mRNA correspond well to the C termini of the truncated CRP proteins. We conclude that ribosome stalling by the arrest sequence induces mRNA cleavage near the arrest point, resulting in nonstop mRNAs that are recognized by tmRNA. We propose that the mRNA cleavage induced by ribosome stalling acts in concert with the tmRNA system as a way to ensure quality control of protein synthesis and possibly to regulate the expression of certain genes.

Adenosine Triphosphatases↗

PubMed2005

The role of ribosomal protein L11 in class I release factor-mediated translation termination and translational accuracy.

It has been suggested from in vivo and cryoelectron micrographic studies that the large ribosomal subunit protein L11 and its N-terminal domain play an important role in peptide release by, in particular, the class I release factor RF1. In this work, we have studied in vitro the role of L11 in translation termination with ribosomes from a wild type strain (WT-L11), an L11 knocked-out strain (DeltaL11), and an L11 N terminus truncated strain (Cter-L11). Our data show 4-6-fold reductions in termination efficiency (k(cat)/K(m)) of RF1, but not of RF2, on DeltaL11 and Cter-L11 ribosomes compared with wild type. There is, at the same time, no effect of these L11 alterations on the maximal rate of ester bond cleavage by either RF1 or RF2. The rates of dissociation of RF2 but not of RF1 from the ribosome after peptide release are somewhat reduced by the L11 changes irrespective of the presence of RF3, and they cause a 2-fold decrease in the missense error. Our results suggest that the L11 modifications increase nonsense suppression at UAG codons because of the reduced termination efficiency of RF1 and that they decrease nonsense suppression at UGA codons because of a decreased missense error level.

Codon, Terminator↗

PubMed1996

Translational control of programmed cell death: eukaryotic translation initiation factor 4E blocks apoptosis in growth-factor-restricted fibroblasts with physiologically expressed or deregulated Myc.

There is increasing evidence that cell cycle transit is potentially lethal, with survival depending on the activation of metabolic pathways which block apoptosis. However, the identities of those pathways coupling cell cycle transit to survival remain undefined. Here we show that the eukaryotic translation initiation factor 4E (eIF4E) can mediate both proliferative and survival signaling. Overexpression of eIF4E completely substituted for serum or individual growth factors in preserving the viability of established NIH 3T3 fibroblasts. An eIF4E mutant (Ser-53 changed to Ala) defective in mediating its growth-factor-regulated functions was also defective in its survival signaling. Survival signaling by enforced expression of eIF4E did not result from autocrine release of survival factors, nor did it lead to increased expression of the apoptosis antagonists Bcl-2 and Bcl-XL. In addition, the execution apparatus of the apoptotic response in eIF4E-overexpressing cells was found to be intact. Increased expression of eIF4E was sufficient to inhibit apoptosis in serum-restricted primary fibroblasts with enforced expression of Myc. In contrast, activation of Ha-Ras, which is required for eIF4E proliferative signaling, did not suppress Myc-induced apoptosis. These data suggest that the eIF4E-activated pathways leading to survival and cell cycle progression are distinct. This dual signaling of proliferation and survival might be the basis for the potency of eIF4E as an inducer of neoplastic transformation.

3T3 Cells↗

PubMed1985

The legacy of Ewald Hecker: a new translation of "Die Hebephrenie". Translated by Marie-Louise Schoelly.

Early descriptions of schizophrenia may be found in the writings of Haslam and Morel, but the turning point in the development of the modern concept was Ewald Hecker's classic paper on hebephrenia in 1871. The syndrome he described--a psychosis of early onset with a deteriorating course characterized by a "silly" affect, behavioral peculiarities, and formal thought disorder--not only adumbrated Kraepelin's generic category of dementia praecox but quite specifically defined the later subtype of hebephrenic, or disorganized, schizophrenia as well. The present translation into English of Hecker's "Die Hebephrenie" makes accessible a crucial milestone in the history of modern psychiatry.

Germany↗

PubMed2004

Correlation between semaphorin3A-induced facilitation of axonal transport and local activation of a translation initiation factor eukaryotic translation initiation factor 4E.

An impressive body of evidence has been accumulated indicating that local protein synthesis is implicated in navigation of neurite extension induced by guidance cues, such as semaphorin3A (Sema3A). We found previously that a Src type tyrosine kinase Fyn and cyclin-dependent kinase 5 (Cdk5) mediate Sema3A-signaling. We also showed that Sema3A elicits axonal transport through neuropilin-1, a receptor for Sema3A, located at the growth cones. Here, we investigate the relationship between Sema3A-induced local signaling, protein synthesis, and axonal transport. Lavendustin A, a tyrosine kinase inhibitor, and olomoucine, a cyclin-dependent kinase inhibitor, suppressed Sema3A-induced facilitation of anterograde and retrograde axonal transport in dorsal root ganglion (DRG) neuron with and without the cell body. Sema3A-induced facilitation of axonal transport was attenuated in DRG neurons of fyn- (fyn-/-) and a Cdk5 activator, p35 (p35-/-)-deficient mice when compared with those of wild-type or heterozygous mice. Inhibition of protein synthesis suppressed Sema3A-induced facilitation of axonal transport in the DRG neuron with and without the cell body. Sema3A enhanced the level of immunoreactivity of phosphorylated eukaryotic translation initiation factor 4E (eIF-4E) within 5 min in growth cones in a time course similar to that of the facilitated axonal transport. This enhanced signal for phospho-eIF4E was blocked by lavendustin A or olomoucine and was not detected in the fyn-/- and p35-/- neurons. These results provide evidence for a mutual regulatory mechanism between local protein synthesis and axonal transport.

Animals↗

PubMed1992

Dissociation of transcription from translation of human IL-1 beta: induction of steady state mRNA by adherence or recombinant C5a in the absence of translation.

Interleukin (IL) 1 is an important mediator of local and systemic disease. Blocking IL-1 using the IL-1 receptor antagonist has reduced the severity of disease in animal models of septic shock, diabetes, graft-vs-host disease, inflammatory bowel disease, and the spontaneous proliferation of leukemia cells. Blocking IL-1 and reduction in the synthesis of IL-1 are important strategies for reducing the progression of inflammatory disease and autoimmune diseases. Nature, however, maintains control over the synthesis of IL-1 by dissociating transcription for translation. In this paper, the basis for the dissociation of IL-1 beta synthesis of mRNA from synthesis of the IL-1 beta protein is reviewed.

Cell Adhesion↗

PubMed1991

Differences between the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase and low density lipoprotein receptor in human hepatoma cells and fibroblasts reside primarily at the translational and post-translational levels.

We have carried out parallel analyses of the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGr) and low density lipoprotein receptor (LDLr) in two highly differentiated human hepatoma cell lines, HepG2 and Hep3B, and primary cultures of human fibroblasts. Analyses of the levels of HMGr and LDLr mRNAs under a variety of culture conditions that perturb intracellular sterol metabolism, or which differ in the levels of extracellular sterols, indicated that the hepatoma cells and fibroblasts responded similarly in terms of the repression or induction ratios of both mRNAs. However, the absolute levels of the mRNAs were severalfold higher in the hepatoma cells. The major difference between the responses of the hepatoma cells and fibroblasts involved the increase in expression of LDLr which occurred upon shifting the cells from complete to lipoprotein-depleted serum. Under these conditions, the 3-fold increase in rate of synthesis of LDLr in the hepatomas was closely matched by increases in the level of its mRNA. In the case of fibroblasts, a 10-fold increase in translational efficiency was required to explain the 30-fold change in rate of synthesis of LDLr. Polysome profiles from both hepatoma cells and fibroblasts suggest that the rate of elongation or termination on LDLr mRNA is relatively low in the presence of reconstituted complete serum, and that it increases in fibroblasts upon lipoprotein depletion, but not in the hepatoma cells. These data indicate that hepatic expression of LDLr may be relatively refractory to induction by decreased circulating levels of lipoprotein when compared with peripheral tissues.

Carcinoma, Hepatocellular↗

PubMed1977

Initiation and translation in vitro of mRNA for MOPC 315 immunoglobulin heavy chain and characterization of translation product.

An initiation study of mineral oil-induced plasmacytoma (MOPC) 315 heavy chain immunoglobulin (H315) in vitro has been conducted using formyl-[35S]methionyl-tRNAfMet and a highly purified 18 S message from MOPC 315 solid tumor in a crude rabbit reticulocyte lysate system. The product was specifically precipitated by antibodies directed against MOPC 315 immunoglobulin and H315. The in vitro H315 products terminally labeled with formyl-[35S]methionine or internally labeled with [3H]leucine were electrophoretically identical with in vivo H315 on sodium dodecyl sulfate-polyacrylamide gels. All of the [35S]-methionine was incorporated at the NH2 terminus, not internally, since there is a near complete recovery of [35S]methionine following one cycle of Edman degradation. The NH2-terminal cyanogen bromide peptide, CN2, of in vivo and in vitro H315 co-migrated exactly on gel electrophoresis under conditions which completely resolved two proteins differing in size by only 14 amino acids. These data strongly suggest that there is no NH2-terminal precursor of H315 in this system. Cyanogen bromide peptide profiles of in vivo and in vitro H315 were chromatographically indistinguishable. Three peptides, CN1, CN2, and CN4, which represent approximately 85% of the total amino acids of H315 were isolated and further characterized by electrophoresis and paper chromatography. All were very similar to the corresponding peptides of authentic H315. We conclude that the fidelity of H315 translation is preserved in vitro.

Cell Line↗

PubMed1983

[Functioning of amino acid operons in Escherichia coli strains with an altered transcription and translation apparatus. II. The effect of mutations in genes coding ribosomal protein S5 and translation elongation factor G on the functioning of the ilv operon].

It has been shown in our previous study that mutations in genes relA, relC and rpsL result in the delay in Escherichia coli ilv operon derepression; the complete inhibition of derepression of the ilv operon is observed in the double mutants having alterations in rpsL and relA or relC genes. At present, some mutations occurring in the fus gene and altering the structure of the translational elongation G factor have been also found to delay derepression of E. coli ilv operon and complete inhibition in fusr and rel double mutants. Phenotypical ile and val auxotrophy is also detected in the double E. coli mutants with spectinomycin resistance mutation in rpsE gene coding for the structure of ribosomal S5 protein and mutations in relA or relC genes. The suggestion of participation of the ilv operon in regulation of other E. coli amino acid operons expression is discussed.

Amino Acid Sequence↗

PubMed1983

[Expression of the amino acid operons in Escherichia coli strains with an altered transcription and translation apparatus. IV. The effect of mutations disturbing the coupling of the transcription and translation processes on ilv-operon expression in cells carrying the mutation in the spoT gene].

The rate of adaptation of Escherichia coli K-12 NF930 spoT1 cells with elevated intracellular level of ppGpp to various minimal media was studied. It has been found that the rate of adaptation of spoT cells, like that of parent and rel strains, depends mainly on the rate of derepression of the ilv operon. The maximal rate of the ilv operon derepression was observed when an optimal concentration of ppGpp was maintained in cells. Derepression of the ilv operon is sharply delayed when the level of ppGpp is elevated or reduced. Mutations altering the translation system do not change the rate of adaptation of spoT cells. Rifampicin resistance mutations which altered the structure of RNA polymerase change the rate of adaptation of spoT cells to minimal media, especially to those containing serine at high concentrations. The possible role of serine in the regulation of ppGpp degradation system is discussed.

Adaptation, Physiological↗

PubMedDate not supplied

Mechanism of interferon action. Translational control and the RNA-dependent protein kinase (PKR): antagonists of PKR enhance the translational activity of mRNAs that include a 161 nucleotide region from reovirus S1 mRNA.

The interferon-inducible, RNA-dependent protein kinase (PKR) is an important regulator of viral protein synthesis. Activated PKR inhibits protein synthesis by phosphorylating initiation factor eIF-2 alpha. The reovirus S4 gene, whose 1196 nucleotide mRNA transcript does not activate the PKR kinase, is efficiently expressed in vector-transfected monkey COS cells. By contrast, the 1463 nucleotide S1 gene of reovirus, which is a potent activator of PKR, is poorly expressed in COS cells. Virus genetic engineering was therefore used to examine the effect of the PKR activator sequence from the reovirus S1 gene on the expression of chimeric genes of reovirus in transfected COS cells. Chimeric S1/S4 and S4/S1/S4 reovirus constructions that included the PKR activator sequence from S1 in the sigma 3 ORF of S4 were expressed much less efficiently than wild-type S4. However, expression of sigma 3 from S4 (3'UTR/S1), which included the PKR activator sequence from S1 within the 3'-UTR of S4, was comparable to that from wild-type S4. Treatment of COS cells with 2-aminopurine, an inhibitor of PKR, increased the expression of the reovirus S1, S1/S4, and S4/S1/S4 chimeric genes but not the S4 gene or S4 (3'UTR/S1) chimera in transfected COS cells. Likewise, coexpression of the phosphotransfer-negative mutant PKR (K296R) increased the expression of reovirus S1, S1/S4 and S4/S1/S4 chimeric genes but not the S4 gene or S4 (3'UTR/S1) chimera in cotransfected COS cells. Truncated PKR(1-243) which includes the dsRNA binding domain but not the kinase catalytic subdomains was able to enhance the expression of reovirus S1, but did not affect S4 expression. The dsRNA binding protein E3L encoded by vaccinia virus also increased S1 expression similar to PKR (1-243) and PKR(K296R). These results suggest that the translational repression in vivo mediated by PKR is selective for mRNAs that possess the kinase activator region, and that the dominant negative effect of PKR on gene expression is likely mediated by the RNA binding activity of the PKR protein.

2-Aminopurine↗

PubMed1996

Expression of a translationally regulated, dominant-negative CCAAT/enhancer-binding protein beta isoform and up-regulation of the eukaryotic translation initiation factor 2alpha are correlated with neoplastic transformation of mammary epithelial cells.

A translationally regulated, dominant-negative isoform of CCAAT/enhancer-binding protein beta is expressed in transplantable and primary mouse mammary tumors of different etiologies but is not expressed in preneoplastic mammary hyperplasias or in primary prostate, lung, lens, ovary or lymphoid tumors. The eukaryotic initiation factor 2alpha protein is also expressed at significantly higher levels (69.8 +/- 7.2%) in these mammary tumors compared with normal and hyperplastic tissues. Thus, misregulation of eukaryotic initiation factor 2alpha may promote the expression of a dominant-negative CCAAT/enhancer-binding protein beta isoform, which may inhibit terminal differentiation and facilitate uncontrolled proliferation of mammary epithelial cells.

Animals↗

PubMedDate not supplied