Nucleotide sequence of human elongation factor-1β cDNA

Sanders, J.P.; Maassen, J. Antonie; Amons, Reinout; Möller, W.

doi:10.1093/nar/19.16.4551

Cited by 38 publications

(13 citation statements)

References 6 publications

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“…It is notable that rapidly growing cultured cells exhibit a large increase in eEF1A levels compared with that seen in tissues (26,37). Indeed, in an attempt to deduce a physiological role for eEF1A, we ectopically expressed eEF1A1 in HEK-293T and CHO cells to determine its effect on cellular SK activity.…”

Section: Figure 5 Phosphorylation Of Sk1 Does Not Alter Binding To Ementioning

confidence: 99%

Eukaryotic Elongation Factor 1A Interacts with Sphingosine Kinase and Directly Enhances Its Catalytic Activity

Leclercq

Moretti

Vadas

et al. 2008

Journal of Biological Chemistry

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Sphingosine 1-phosphate (S1P) has many important roles in mammalian cells, including contributing to the control of cell survival and proliferation. S1P is generated by sphingosine kinases (SKs), of which two mammalian isoforms have been identified (SK1 and SK2). To gain a better understanding of SK regulation, we have used a yeast two-hybrid screen to identify SK1-interacting proteins and established elongation factor 1A (eEF1A) as one such protein that associates with both SK1 and SK2. We show the direct interaction of eEF1A with the SKs in vitro, whereas the physiological relevance of this association was demonstrated by co-immunoprecipitation of the endogenous proteins from cell lysates. Although the canonical role of eEF1A resides in protein synthesis, it has also been implicated in other roles, including regulating the activity of some signaling enzymes. Thus, we examined the potential role of eEF1A in regulation of the SKs and show that eEF1A is able to directly increase the activity of SK1 and SK2 ϳ3-fold in vitro. Substrate kinetics demonstrated that eEF1A increased the catalytic rate of both SKs, while having no observable effect on substrate affinities of these enzymes for either ATP or sphingosine. Overexpression of eEF1A in quiescent Chinese hamster ovary cells increased cellular SK activity, whereas a small interfering RNAmediated decrease in eEF1A levels in MCF7 cells substantially reduced cellular SK activity and S1P levels, supporting the in vivo physiological relevance of this interaction. Thus, this study has established a novel mechanism of regulation of both SK1 and SK2 that is mediated by their interaction with eEF1A. Sphingosine kinases (SKs)2 catalyze the phosphorylation of sphingosine to generate the bio-active phospholipid sphingosine 1-phosphate (S1P). S1P affects many biological processes including calcium mobilization, mitogenesis, apoptosis, atherosclerosis, asthma, hypertension, inflammatory responses, and cytoskeleton rearrangement (1). In humans two sphingosine kinases, SK1 and SK2, have been identified (2, 3). These enzymes appear to be the major control elements regulating cellular S1P levels (4) and thus play a crucial role in modulating the effects attributed to S1P.There is now convincing evidence that SK1 and S1P play roles in tumorigenesis. We have shown that overexpression of SK1 in NIH-3T3 fibroblasts results in neoplastic transformation of these cells (5). Other studies have demonstrated the involvement of SK1 in estrogen-dependent regulation of breast tumor cell growth and survival (6, 7). Furthermore, French et al. (8) have shown that SK1 mRNA levels are elevated in a variety of human solid tumors and, importantly, that tumor growth can be reduced in vivo by SK inhibitors. Similarly, we have shown that the use of SK inhibitors or a dominant negative SK can block neoplastic cell transformation induced by oncogenic H-Ras (5).We have established that activation of SK1 can be mediated by phosphorylation at Ser 225 by ERK1/2 (9). This phosphorylation not only results in ...

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Section: Figure 5 Phosphorylation Of Sk1 Does Not Alter Binding To Ementioning

confidence: 99%

Eukaryotic Elongation Factor 1A Interacts with Sphingosine Kinase and Directly Enhances Its Catalytic Activity

Leclercq

Moretti

Vadas

et al. 2008

Journal of Biological Chemistry

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“…The carboxy-terminal part of EF-1 P possessing the GDP-exchange-stimulating activity is almost as active in stimulating GDP exchange on EF-la as intact EF-1P. It is this same part which shows a strong sequence similarity to not only EF-lP from pig (R. Amons, A. Schipper, H. T. F. van Damme, J. Kriek and W. Moller, unpublished results) and human [46], but also to EF-1P-like protein EF-16 [3]. Remarkably, the exchange of guanine nucleotides, bound to EF-la from Artemia, can be stimulated with EF-1P from pig (H. T. F. van Damme, A. Schipper, R. Amons, W. Moller, unpublished results) as well as with EF-1P from Xenopus [12].…”

Section: Discussionmentioning

confidence: 89%

Identification of the sites in the eukaryotic elongation factor 1α involved in the binding of elongation factor 1β and aminoacyl‐tRNA

Damme¹,

Amons²,

Möller³

1992

European Journal of Biochemistry

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In this article we report the identification of the sites which are involved in the binding of the GDP-exchange factor EF-1P and aminoacyl tRNA to the a-subunit of the eukaryotic elongation factor 1 (EF-1) from Artemiu. For this purpose the polypeptide chain of EF-la, having 461 amino acid residues, was proteolytically cleaved into large fragments by distinct proteases. Under well defined conditions, a mixture of two large fragments, free from intact EF-la and with molecular masses of 37 kDa and 43 kDa, was obtained. The 37-kDa and 43-kDa fragments comprise the residues 129 -461 and 69 -461, respectively. However, in aqueous solution and under non-denaturing conditions, the mixture still contained a short amino-terminal peptide, encompassing the residues 1 -36, that remained tightly bound. The ability of the mixture of the 37 + 43-kDa fragments, including this amino-terminal peptide 1 -36, to bind GDP or to facilitate aminoacyl tRNA binding to saltwashed ribosomes was severely reduced, compared to intact EF-la. However, both of these complexes were able to bind to the GDP-exchange-stimulating subunit EF-1P . A 30-kDa fragment, comprising the residues 1 -287, was generated after treatment of the protein with endoproteinase Glu-C. This fragment contained the complete guanine nucleotide binding pocket. Although it was able to bind GDP and to transport aminoacyl tRNA to the ribosome, no affinity towards EF-1P was observed. We propose that the guanine-nucleotide-exchange stimulation by EF-1P is induced through binding of this factor to the carboxy-terminal part of EF-la. As a result, a decreased susceptibility towards trypsin of the guanine-nucleotide-binding pocket of EF-la, especially in the region of its presumed effector loop is induced.The most abundant eukaryotic translational factor is elongation factor 1 (EF-I), which catalyzes, together with the translocation factor EF-2, the elongation cycle in the eukaryotic protein synthesis [I, 21. EF-1 contains four subunits (a, /J, y and 6). The elongation starts with the formation of a ternary complex between EF-la, aminoacyl tRNA and GTP. Subsequently, aminoacyl tRNA is attached to the mRNAribosome complex. This latter step requires hydrolysis of GTP [2]. The elongation rate is increased by two other subunits, EF-1P and EF-16, which both accelerate the exchange of GDP, bound to EF-la, for GTP [2, 31. EF-1P and EF-16 have been isolated in complex with EF-1y as heterodimers or heterotrimers (EF-Ifiy or EF-lPyG, respectively). Together with EF-1 a, these EF-IPy(6) complexes form high-moleclarmass aggregates, which have been described in many eukaryotic species [4 -81. Furthermore, these aggregates have been found associated with valyl tRNA synthetase [9, 101.

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“…The cDNA probes for human EF 1-␤ (entire coding region) and EF 1-␥ (complete coding sequence except the nucleotides coding for the four NH2-terminal amino acids) were kindly provided into the plasmid PUC120 by William Möller, Sylvius Laboratory, Leiden, The Netherlands (11,12).…”

Section: Methodsmentioning

confidence: 99%

Unbalanced expression of the different subunits of elongation factor 1 in diabetic skeletal muscle

Reynet

Kahn

2001

Proc. Natl. Acad. Sci. U.S.A.

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In studies using subtraction cloning to screen for alterations in mRNA expression in skeletal muscle from humans with Type 2 diabetes mellitus and control subjects, one of the most prominent differences was in the mRNA for elongation factor (EF)-1␣. With Northern blot analysis, EF-1␣ expression was enhanced by 2-to 6-fold in both Types 1 and 2 human diabetics. In contrast, no changes in expression of EF-1␤ or -␥ were noted. We observed similar results in animal models of Type 1 diabetes. EF-1␣ expression, but not EF-1␤ or -␥ expression, was also enhanced in streptozotocin-induced diabetic rats, and this effect was reversed by insulin treatment. An increased level of EF-1␣ mRNA was also observed in nonobese diabetic mice. This unbalanced regulation of the expression of the different subunits of EF-1 may contribute to alterations not only in protein synthesis but also in other cellular events observed in the diabetic state.gene expression ͉ insulin ͉ diabetes mellitus U ncontrolled diabetes mellitus results in complex metabolic and structural alterations, leading to abnormal carbohydrate, lipid, and protein metabolism, as well as long-term complications involving vascular tissue, kidney, and nerve. The molecular mechanisms for many of these changes are not completely understood but involve changes in both the level and posttranslational modification of a number of proteins. The decreased levels of insulin and the insulin resistance present in Types 1 and 2 diabetes lead to a reduction in total protein synthesis, as well as changes in expression of a number of insulin and͞or metabolically regulated genes (1-3).Although there is considerable information about the effects of diabetes and insulin on regulation of genes in liver and adipose tissue, much less is known about their effects on gene expression in skeletal muscle. In an attempt to identify changes in gene expression at the level of skeletal muscle in humans with diabetes mellitus, we have used the technique of subtraction cloning (4). Using this approach, we demonstrated increased expression of several mitochondrially encoded genes (5) and glycogen phosphorylase (6) in muscle from diabetic patients. We also identified Rad, a member of the Ras͞GTPase proteins family, whose expression was selectively increased in muscle of some patients with Type 2 diabetes as compared with muscle from normal individuals or patients with Type 1 diabetes (7).This report details another GTP-binding protein whose expression is increased in diabetes that was uncovered by using this technique, namely eukaryotic elongation factor (EF)-1␣ (also named EF-1A) (8). EF-1, the primary factor involved in the elongation reaction during protein synthesis, is composed of two distinct parts: a nucleotide-binding subunit, EF-1␣, and a nucleotide exchange complex composed of the EF-1␤ and -␥ subunits (8, 9). EF-1␣ binds GTP and aminoacyl-tRNA and leads to the codon-dependent placement of the aminoacyl-tRNA at the acceptor site on the ribosome. After release of EF-1␣-GDP from the ribosome, the co...

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Nucleotide sequence of human elongation factor-1β cDNA

Cited by 38 publications

References 6 publications

Eukaryotic Elongation Factor 1A Interacts with Sphingosine Kinase and Directly Enhances Its Catalytic Activity

Eukaryotic Elongation Factor 1A Interacts with Sphingosine Kinase and Directly Enhances Its Catalytic Activity

Identification of the sites in the eukaryotic elongation factor 1α involved in the binding of elongation factor 1β and aminoacyl‐tRNA

Unbalanced expression of the different subunits of elongation factor 1 in diabetic skeletal muscle

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