GDP As a Regulator of Phosphorylation of Elongation Factor 1 by Casein Kinase II

Paleń, E; Venema, Richard C.; Chang, Yu Yin; Traugh, Jolinda A.

doi:10.1021/bi00194a016

Cited by 16 publications

(9 citation statements)

References 47 publications

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“…The lack of an effect by GDP is consistent with the availability of ␤ and ␦ in the reconstituted complex for phosphorylation by CKII as compared with EF-1. In the same study (35), polylysine was shown to stimulate phosphorylation of ␤ in native EF-1 but not ␦. With the recombinant subunits, polylysine stimulated phosphorylation of ␤ but not the ␦ subunit (data not shown), which is consistent with the studies on native EF-1.…”

Section: Discussionmentioning

confidence: 78%

“…Previously, Palen et al (35) showed that GDP stimulated the phosphorylation of the ␤ and ␦ subunits of native EF-1 by CKII. Such a stimulation was not observed with the recombinant subunits or reconstituted ␤␥␦.…”

Section: Discussionmentioning

confidence: 99%

“…Effects of Phosphorylation of ␤ and ␦ by Protein Kinase CKII on Elongation-CKII has been shown to phosphorylate the ␤ and ␦ subunits of EF-1 (35). When the purified recombinant subunits were examined separately as substrates for CKII, 0.6 and 0.8 mol/mol of phosphate were incorporated into the ␤ and ␦ subunits, respectively.…”

Section: Stimulation Of the Elongation Activity Of Ef-1␣ By Ef-1␤mentioning

confidence: 99%

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Recombinant Subunits of Mammalian Elongation Factor 1 Expressed in Escherichia coli

Sheu¹,

Traugh

1997

Journal of Biological Chemistry

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The first step in elongation requires two different activities; elongation factor (EF)-1␣ transfers aminoacyltRNA to the ribosome and is released upon hydrolysis of GTP, EF-1␤␥␦ catalyzes exchange of GDP on EF-1␣ with GTP. To analyze the role of the individual subunits of EF-1 in elongation, the cDNAs for the ␤, ␥, and ␦ subunits of EF-1 from rabbit were cloned, and proteins of 225, 437, and 280 amino acids, respectively, were expressed in Escherichia coli. The purified recombinant ␤ subunit migrates as a dimer and the ␥ subunit as a trimer upon gel filtration, whereas the ␦ subunit forms a large aggregate. Complexes of ␤␥, ␥␦ and ␤␥␦ were formed by selfassociation and eluted with a molecular mass of approximately 160, 530, and 670 kDa, respectively; no interaction was observed between ␤ and ␦. The activity of the recombinant subunits was determined with native EF-1␣ by measuring stimulation of the rate of elongation by poly(U)-directed polyphenylalanine synthesis. Recombinant ␤ and ␦ alone stimulated the rate of elongation by 10-fold, with a ratio of 5␣:2␤ or ␦. The ␤␥␦ complex stimulated EF-1␣ activity up to 10-fold with a ratio of 20␣ to 1␤␥␦. Phosphorylation of the ␤ and ␦ subunits alone or in ␤␥␦ by protein kinase CKII had no effect on the rate of elongation. Eukaryotic elongation factor (EF)1 1 consists of four subunits, EF-1␣, ␤, ␥, and ␦. EF-1␣ (50 kDa) forms a ternary complex with GTP and aminoacyl-tRNA and transfers the aminoacyl-tRNA to 80 S ribosomes with the hydrolysis of GTP. EF-1␤␥␦ facilitates the exchange of the GDP bound to EF-1␣ for GTP, initiating another round of elongation (1, 2). EF-1␤ and ␦ contain the GTP exchange activity (3); ␥ is tightly associated to ␤ and is removed only under denaturing conditions (4). The cDNAs for ␤ and ␥ have been cloned and sequenced from a number of different organisms and tissues (5-13). Recently, EF-1␦ cDNA has been sequenced from human (14) and Xenopus (15), and partial amino acid sequences have been obtained from Artemia (14). A leucine zipper motif in the amino terminus of ␦ is found in all three species (14). EF-1␤ and ␦ have a highly homologous carboxyl terminus that contains the GDP/GTP exchange activity, whereas the amino-terminal domains differ and appear to be important for regulation of EF-1 activity. The function of the ␥ subunit is unknown, although there is evidence that ␥ can stimulate the nucleotide exchange activity of ␤ (16). EF-1␥ may anchor the complex to the membrane (16) and has been shown to contain a sequence homologous to glutathione S-transferase in the amino-terminal domain, which has been postulated to be involved in regulation of the assembly of multisubunit complexes (17).Eukaryotic EF-1 has been isolated from a variety of organisms and tissues with molecular weights ranging from 50,000 to about 1 ϫ 10 6 . The low molecular weight form is EF-1␣, the intermediate form is EF-1␣␤␥␦, and the high molecular weight form is a complex of five polypeptides, valyl-tRNA synthetase (ValRS) and .Using purified subunits of EF-1 from Artemia, t...

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Section: Discussionmentioning

confidence: 78%

Section: Discussionmentioning

confidence: 99%

Section: Stimulation Of the Elongation Activity Of Ef-1␣ By Ef-1␤mentioning

confidence: 99%

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Recombinant Subunits of Mammalian Elongation Factor 1 Expressed in Escherichia coli

Sheu¹,

Traugh

1997

Journal of Biological Chemistry

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“…EF-1␦, therefore, plays a key role in protein synthesis by regulating the activity of EF-1␣. EF-1␦ is phosphorylated by several cellular protein kinases, including cdc2, casein kinase II (CKII), protein kinase C, and multipotential S6 kinase (3,29,35,46). Studies on the phosphorylation of EF-1␦ suggest that these modifications of the protein alter translational efficiency (3,26,46,47).…”

mentioning

confidence: 99%

Conserved Protein Kinases Encoded by Herpesviruses and Cellular Protein Kinase cdc2 Target the Same Phosphorylation Site in Eukaryotic Elongation Factor 1δ

Kawaguchi

Kato

Tanaka

et al. 2003

J Virol

132

178

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Earlier studies have shown that translation elongation factor 1␦ (EF-1␦) is hyperphosphorylated in various mammalian cells infected with representative alpha-, beta-, and gammaherpesviruses and that the modification is mediated by conserved viral protein kinases encoded by herpesviruses, including UL13 of herpes simplex virus type 1 (HSV-1), UL97 of human cytomegalovirus, and BGLF4 of Epstein-Barr virus (EBV). In the present study, we attempted to identify the site in EF-1␦ associated with the hyperphosphorylation by the herpesvirus protein kinases The family Herpesviridae can be divided into three subfamilies (the Alphaherpesvirinae, the Betaherpesvirinae, and the Gammaherpesvirinae), and to date, approximately 130 herpesviruses have been identified from various animal species (41). Despite the wide range of biological properties and pathogenic manifestations of the herpesviruses, their genomes contain a significant number of conserved genes (41). This conservation suggests that the products of these genes play essential roles in the life cycle of herpesviruses. Herpesviruses contain viral genes that are predicted to encode protein kinases (4, 44). Among them, a subset exemplified by UL13 of herpes simplex virus type 1 (HSV-1) is conserved in all of the Herpesviridae (4, 44). Conceivably herpesviruses universally utilize their products both to regulate their own replicative processes and to modify cellular machinery through the phosphorylation of target viral and cellular proteins.HSV-1 UL13, a subject of this study, is a serine/threonine protein kinase that is packaged into the virion (6, 34). Studies with UL13 mutants showed that the viral protein kinase affects the accumulation of an ␣ protein, ICP0, and a subset of ␥ 2 proteins, including UL26, UL26.5, UL38, UL41, and Us11 (37), suggesting that UL13 plays a role in viral gene expression in infected cells. Several lines of evidence listed below indicate that the function of UL13 is closely linked to that of the other viral regulatory proteins ICP22 and Us1.5, both of which are encoded by the ␣22 gene. First, ICP22 and Us1.5 are hypophosphorylated in cells infected with UL13 mutant viruses, suggesting that the UL13 protein kinase phosphorylates these viral regulatory proteins (38). Second, the phenotype of UL13 deletion mutants cannot be differentiated from that of ICP22 and Us1.5 deletion mutant viruses with respect to the accumulation of ICP0 and the subset of ␥ 2 proteins (37). Third, both UL13 and ICP22 and Us1.5 are involved in the HSV-1-induced activation and modification of cellular enzymes, including the protein kinase cdc2 (1) and the large subunit of RNA polymerase II (22). Although this series of observations suggests that UL13 expresses its regulatory function by phosphorylating ICP22 and Us1.5, the direct linkage between phosphorylation

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“…The ␤ and ␦ subunits are also phosphorylated by protein kinase CKII (13)(14)(15)(16). GDP has been shown to enhance the phosphorylation of ␤ and ␦ by CKII, while little phosphorylation is observed in the absence of GDP (16).…”

mentioning

confidence: 99%

Phosphorylation of Elongation Factor 1 and Ribosomal Protein S6 by Multipotential S6 Kinase and Insulin Stimulation of Translational Elongation

Chang

Traugh

1997

Journal of Biological Chemistry

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Stimulation of protein synthesis in response to insulin is concomitant with increased phosphorylation of initiation factors 4B and 4G and ribosomal protein S6 (Morley, S. J., and Traugh, J. A. (1993) Biochimie 75, 985-989) and is due at least in part to multipotential S6 kinase. When elongation factor 1 (EF-1) from rabbit reticulocytes was examined as substrate for multipotential S6 kinase, up to 1 mol/mol of phosphate was incorporated into the ␣, ␤, and ␦ subunits. Phosphorylation of EF-1 resulted in a 2-2.6-fold stimulation of EF-1 activity, as measured by poly(U)-directed polyphenylalanine synthesis. The rate of elongation was also stimulated by approximately 2-fold with 80 S ribosomes phosphorylated on S6 by multipotential S6 kinase. When the rates of elongation in extracts from serum-fed 3T3-L1 cells and cells serum-deprived for 1.5 h were compared, a 40% decrease was observed upon serum deprivation. The addition of insulin to serum-deprived cells for 15 min stimulated elongation to a rate equivalent to that of serumfed cells. Similar results were obtained with partially purified EF-1, with both EF-1 and ribosomes contributing to stimulation of elongation. These data are consistent with a ribosomal transit time of 3.2 min for serumdeprived cells and 1.6 min following the addition of insulin for 15 min. Taken together, the data suggest that insulin stimulation involves coordinate regulation of EF-1 and ribosomes through phosphorylation by multipotential S6 kinase. Elongation factor 1 (EF-1)1 is composed of a nucleotide-binding protein, EF-1␣, and a nucleotide exchange protein complex, EF-1␤␥␦ (1, 2). The EF-1␣ subunit (50.1 kDa) binds GTP and mediates the attachment of aminoacyl-tRNA to ribosomes (1, 2), while the ␤ (24.8-kDa), ␥ (50.0-kDa), and ␦ (31.1-kDa) subunits stimulate the exchange of GDP for GTP on the ␣ subunit (3-5). Several forms of EF-1 can be isolated from eukaryotic cells, including ␣, ␣␤␥, and ␣␤␥␦ (6 -8). Approximately 15% of the EF-1␣␤␥␦ is complexed with valyl-tRNA synthetase (ValRS) (7-11).The mechanisms involved in the regulation of EF-1 activity are not well understood, although EF-1 activity has been shown to be modulated by mitogens, aging, heat shock, and fertilization (12). All four subunits of EF-1 have been shown to undergo phosphorylation in vivo and in vitro. When the ␣, ␤, and ␦ subunits are phosphorylated in rabbit reticulocytes in response to phorbol 12-myristate 13-acetate, a 2-3-fold stimulation of EF-1 activity is observed (6). A similar effect is observed when EF-1 is phosphorylated in vitro by protein kinase C (7), and this effect is due to stimulation of the rate-limiting step, GTP/GDP exchange on EF-1␣ (8).The ␤ and ␦ subunits are also phosphorylated by protein kinase CKII (13-16). GDP has been shown to enhance the phosphorylation of ␤ and ␦ by CKII, while little phosphorylation is observed in the absence of GDP (16). In addition, the ␥ and ␦ subunits of EF-1 from Xenopus oocytes are substrates for the cell cycle division control kinase (Cdc2) (17, 18). The physiologic...

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GDP As a Regulator of Phosphorylation of Elongation Factor 1 by Casein Kinase II

Cited by 16 publications

References 47 publications

Recombinant Subunits of Mammalian Elongation Factor 1 Expressed in Escherichia coli

Recombinant Subunits of Mammalian Elongation Factor 1 Expressed in Escherichia coli

Conserved Protein Kinases Encoded by Herpesviruses and Cellular Protein Kinase cdc2 Target the Same Phosphorylation Site in Eukaryotic Elongation Factor 1δ

Phosphorylation of Elongation Factor 1 and Ribosomal Protein S6 by Multipotential S6 Kinase and Insulin Stimulation of Translational Elongation

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