F-actin sequesters elongation factor 1alpha from interaction with aminoacyl-tRNA in a pH-dependent reaction.

Liu, Gang; Tang, Justin; Edmonds, Brian T.; Murray, John W.; Levin, S. G.; Condeelis, John S.

doi:10.1083/jcb.135.4.953

Cited by 115 publications

(107 citation statements)

References 76 publications

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“…Examples of proteins not directly related to elongation that are known to interact with eEF1A are filamentous actin (F-actin) 51 and the zinc finger protein ZPR1, which is associated with eEF1A and translocated to the nucleus upon treatment of quiescent mammalian cells with mitogens or epidermal growth factor. 52 On the other hand, rapid posttranscriptionally mediated up-regulation of eEF1A has been demonstrated following oxidative stress-induced apoptosis (e.g., upon treatment with hydrogen peroxide).…”

Section: Discussionmentioning

confidence: 99%

Structural Basis for the Binding of Didemnins to Human Elongation Factor eEF1A and Rationale for the Potent Antitumor Activity of These Marine Natural Products

et al. 2004

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Didemnins and tamandarins are closely related marine natural products with potent inhibitory effects on protein synthesis and cell viability. On the basis of available biochemical and structural evidence and results from molecular dynamics simulations, a model is proposed that accounts for the strong and selective binding of these compounds to human elongation factor eEF1A in the presence of GTP. We suggest that the p-methoxyphenyl ring of these cyclic depsipeptides is inserted into the same pocket in eEF1A that normally lodges either the 3′ terminal adenine of aminoacylated tRNA, as inferred from two prokaryotic EF-Tu‚GTP‚tRNA complexes, or the aromatic side chain of Phe/Tyr-163 from the nucleotide exchange factor eEF1BR, as observed in several X-ray crystal structures of a yeast eEF1A:eEF1BR complex. This pocket, which has a strong hydrophobic character, is formed by two protruding loops on the surface of eEF1A domain 2. Further stabilization of the bound depsipeptide is brought about by additional crucial interactions involving eEF1A domain 1 in such a way that the molecule fits snugly at the interface between these two domains. In the GDP-bound form of eEF1A, this binding site exists only as two separate halves, which accounts for the much greater affinity of didemnins for the GTP-bound form of this elongation factor. This binding mode is entirely different from those seen in the complexes of the homologous prokaryotic EF-Tu with kirromycin-type antibiotics or the cyclic thiazolyl peptide antibiotic GE2270A. Interestingly, the set of interactions used by didemnins to bind to eEF1A is also distinct from that used by eEF1BR or eEF1B , thus establishing a competition for binding to a common site that goes beyond simple molecular mimicry. The model presented here is consistent with both available biochemical evidence and known structure-activity relationships for these two classes of natural compounds and synthetic analogues and provides fertile ground for future research.

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Section: Discussionmentioning

confidence: 99%

Structural Basis for the Binding of Didemnins to Human Elongation Factor eEF1A and Rationale for the Potent Antitumor Activity of These Marine Natural Products

et al. 2004

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“…Therefore, it is of interest to know which domains interact with actin and/or mRNA to further define the relationship of EF1␣ binding to the actin cytoskeleton and the mRNA. Previous studies indicated that there is probably more than one actin-binding site on Dityostelium EF1␣ (Liu et al, 1996b). EF1␣ is an evolutionarily conserved protein family that has Ͼ80% sequence homology among eukaryotes.…”

Section: Domain III Contains the Dominant Actin-binding Site Of Ef1␣mentioning

confidence: 99%

“…Therefore, deletion Figure 4. SDS-PAGE gel images of recombinant full-length EF1␣, domains I, II, and III, were previously described (Liu et al, 1996b). Arrowhead, GST-fusion proteins; *, actin; S, supernatant; p, pellet.…”

Section: Deletion Of the C-terminal Actin-binding Site Of Ef1␣ Is Cormentioning

confidence: 99%

“…The effect of anti-C-terminal actin-binding site antibody (anti-403-456) on EF1␣'s cross-linking of actin filaments was analyzed using right angle light scattering as described previously (Liu et al, 1996b). Briefly, rabbit muscle G-actin was allowed to polymerize for 2 h at room temperature before it was used.…”

Section: Light-scattering Assaymentioning

confidence: 99%

“…Dictyostelium actin and EF1␣, rabbit skeletal muscle actin, GST fusion proteins of Dictyostelium and rat EF1␣, and corresponding truncations were prepared as described previously (Liu et al, 1996b). All of the GST fusion proteins were clarified by centrifugation at 320,000 ϫ g at 4°C for 30 min and stored in storage buffer (10 mM PIPES, 0.05% NaN 3 , 0.1 mM EDTA, 1 mM DTT, and 25% glycerol, pH 7.0) on ice for the short term or under liquid nitrogen for the long term.…”

Section: Protein Purificationmentioning

confidence: 99%

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Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Liu

Grant²,

Persky³

et al. 2002

MBoC

158

151

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The targeting of mRNA and local protein synthesis is important for the generation and maintenance of cell polarity. As part of the translational machinery as well as an actin/ microtubule-binding protein, elongation factor 1␣ (EF1␣) is a candidate linker between the protein translation apparatus and the cytoskeleton. We demonstrate in this work that EF1␣ colocalizes with ␤-actin mRNA and F-actin in protrusions of chicken embryo fibroblasts and binds directly to F-actin and ␤-actin mRNA simultaneously in vitro in actin cosedimentation and enzyme-linked immunosorbent assays. To investigate the role of EF1␣ in mRNA targeting, we mapped the two actin-binding sites on EF1␣ at high resolution and defined one site at the N-terminal 49 residues of domain I and the other at the C-terminal 54 residues of domain III. In vitro actin-binding assays and localization in vivo of recombinant full-length EF1␣ and its various truncates demonstrated that the C terminus of domain III was the dominant actin-binding site both in vitro and in vivo. We propose that the EF1␣-F-actin complex is the scaffold that is important for ␤-actin mRNA anchoring. Disruption of this complex would lead to delocalization of the mRNA. This hypothesis was tested by using two dominant negative polypeptides: the actin-binding domain III of EF1␣ and the EF1␣-binding site of yeast Bni1p, a protein that inhibits EF1␣ binding to F-actin and also is required for yeast mRNA localization. We demonstrate that either domain III of EF1␣ or the EF1␣-binding site of Bni1p inhibits EF1␣ binding to ␤-actin mRNA in vitro and causes delocalization of ␤-actin mRNA in chicken embryo fibroblasts. Taken together, these results implicate EF1␣ in the anchoring of ␤-actin mRNA to the protrusion in crawling cells.

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Association between elongation factor-1? and microtubules in vivo is domain dependent and conditional

Moore

Cyr

2000

Cell Motil. Cytoskeleton

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Although the precise definition for a microtubule‐associated protein (MAP) has been the subject of debate, elongation factor‐1α (EF‐1α) fits the most basic criteria for a MAP [Durso and Cyr, 1994a]. It binds, bundles, stabilizes, and promotes the assembly of microtubules in vitro, and localizes to plant microtubule arrays in situ. In this study, the in vitro and in vivo association of EF‐1α with microtubules was further investigated. Analysis of the in vitro binding data for EF‐1α and microtubules indicates that EF‐1α binds cooperatively to the microtubule lattice. In order to investigate the interaction of EF‐1α with microtubules in vivo, GFP fusions to EF‐1α or to EF‐1α truncates were transiently expressed in living plant cells. Using this method, two putative microtubule‐binding domains on EF‐1α were identified: one in the N‐terminal domain I and one in the C‐terminal domain III. The binding of domain I to microtubules in vivo, like the binding of full‐length EF‐1α, is conditional, and requires incubation in weak, lipophilic organic acids. The binding of domain III to microtubules in vivo, however, is not conditional, and occurs under normal cellular regimes. Furthermore, domain III stabilizes cortical microtubules as determined by their resistance to the anti‐microtubule herbicide, oryzalin. Because the accumulation of EF‐1α onto microtubules is unconditional in the absence of domain I, we hypothesize that domain I negatively regulates the accumulation of EF‐1α onto microtubules in vivo. This hypothesis is discussed in terms of possible regulatory mechanisms that could affect the accumulation of EF‐1α onto microtubules within living cells. Cell Motil. Cytoskeleton 45:279–292, 2000 © 2000 Wiley‐Liss, Inc.

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F-actin sequesters elongation factor 1alpha from interaction with aminoacyl-tRNA in a pH-dependent reaction.

Cited by 115 publications

References 76 publications

Structural Basis for the Binding of Didemnins to Human Elongation Factor eEF1A and Rationale for the Potent Antitumor Activity of These Marine Natural Products

Structural Basis for the Binding of Didemnins to Human Elongation Factor eEF1A and Rationale for the Potent Antitumor Activity of These Marine Natural Products

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Association between elongation factor-1? and microtubules in vivo is domain dependent and conditional

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