Modifications of eukaryotic initiation factor 4F (eIF4F) in adult cardiocytes by adenoviral gene transfer: differential effects on eIF4F activity and total protein synthesis rates

2001 ◽  
Vol 356 (2) ◽  
pp. 557-566 ◽  
Author(s):  
Atif N. SAGHIR ◽  
William J. TUXWORTH ◽  
Curt H. HAGEDORN ◽  
Paul J. McDERMOTT
Keyword(s):  
Protein Synthesis ◽  
Gene Transfer ◽  
Complex Formation ◽  
Binding Affinity ◽  
Total Protein ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Adenoviral Gene Transfer ◽  
Eif4f Complex ◽  
Rate Limiting

In adult feline cardiocytes, increases in eukaryotic initiation factor 4F (eIF4F) activity are correlated with accelerated rates of total protein synthesis produced in response to increased load. Adenoviral gene transfer was employed to increase either eIF4F complex formation or the phosphorylation of eIF4E on Ser-209. To simulate load, cardiocytes were electrically stimulated to contract (2Hz, 5ms pulses). Non-stimulated cardiocytes were used as controls. Adenovirus-mediated overexpression of wild-type eIF4E increased the total eIF4E pool by 120–140% above endogenous levels after 24h and produced a corresponding increase in eIF4F content. However, it did not accelerate total protein synthesis rates in quiescent cardiocytes; neither did it potentiate the increase produced by contraction. To modify the affinity of eIF4F, cardiocytes were infected with a mutant (eIF4E/W56F) with a decreased binding affinity for the mRNA cap. Overexpression of eIF4E/W56F increased the quantity of eIF4F but the rate of total protein synthesis was decreased in quiescent and contracting cardiocytes. Overexpression of a mutant that blocked eIF4E phosphorylation (eIF4E/S209A) increased the quantity of eIF4F without any significant effect on total protein synthesis rates in quiescent or contracting cardiocytes. Overexpression of the eIF4E kinase Mnk-1 increased eIF4E phosphorylation without a corresponding increase in eIF4F complex formation or in the rate of total protein synthesis. We conclude the following: (1) eIF4F assembly is increased by raising eIF4E levels via adenoviral gene transfer; (2) the cap binding affinity of eIF4F is a rate-limiting determinant for total protein synthesis rates; and (3) increases in the quantity of eIF4F alone or in eIF4E phosphorylation are not sufficient to accelerate total protein synthesis rates.

scholarly journals Cellular stress in Xenopus kidney cells enhances the phosphorylation of eukaryotic translation initiation factor (eIF)4E and the association of eIF4F with poly(A)-binding protein

1999 ◽  
Vol 342 (3) ◽  
pp. 519-526 ◽  
Author(s):  
Christopher S. FRASER ◽  
Virginia M. PAIN ◽  
Simon J. MORLEY
Keyword(s):  
Protein Synthesis ◽  
Complex Formation ◽  
Map Kinase ◽  
Kinase Inhibitor ◽  
Binding Protein ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Kidney Cells ◽  
Eif4f Complex ◽  
Cellular Stresses

Eukaryotic initiation factor (eIF) 4E binds to the 5′-cap structure of eukaryotic mRNA and has a central role in the control of cell proliferation. We have shown previously that the stimulation of cultured Xenopus kidney cells with serum resulted in the activation of protein synthesis, enhanced phosphorylation of eIF4E and increased binding of the adapter protein, eIF4G, and poly(A)-binding protein (PABP) to eIF4E to form the functional initiation factor complex, eIF4F/PABP. We now show that cellular stresses such as arsenite, anisomycin and heat shock also result in increased phosphorylation of eIF4E, eIF4F complex formation and the association of PABP with eIF4G, in conditions under which the rate of protein synthesis is severely inhibited. In contrast with reported effects on mammalian cells, the stress-induced increase in eIF4F complex formation occurs in the absence of changes in the association of eIF4E with its binding proteins 4E-BP1 or 4E-BP2. The stress-induced changes in eIF4E phosphorylation were totally abrogated by the p38 mitogen-activated protein (MAP) kinase inhibitor SB203580, and were partly inhibited by the phosphoinositide 3-kinase inhibitor LY294002 and the mammalian target of rapamycin (mTOR) inhibitor rapamycin. However, eIF4E phosphorylation was unaffected by extracellular signal-regulated protein kinase (MAP kinase) inhibitor PD98059. These results indicate that cellular stresses activate multiple signalling pathways that converge at the level of eIF4F complex formation to influence the interactions between eIF4E, eIF4G and PABP.

scholarly journals Fission Yeast Asc1 Stabilizes the Interaction between Eukaryotic Initiation Factor 3a and Rps0A/uS2 for Protein Synthesis

2019 ◽  
Vol 39 (19) ◽  
Author(s):  
Yi-Ting Wang ◽  
Yu-Chen Chien ◽  
Wan-Yi Hsiao ◽  
Chien-Chia Wang ◽  
Shao-Win Wang
Keyword(s):  
Protein Synthesis ◽  
Complex Formation ◽  
Fission Yeast ◽  
Ribosomal Subunit ◽  
Trna Synthetase ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Aminoacyl Trna Synthetase ◽  
Preinitiation Complex ◽  
40S Ribosomal Subunit

ABSTRACT Aminoacyl-tRNA synthetase cofactors play important roles in coordinating aminoacylation and translation. In this study, we describe an additional function of the fission yeast aminoacyl-tRNA synthetase cofactor 1 (Asc1) in translation. We found that Asc1 directly binds and stabilizes the interaction between small ribosomal protein Rps0A/uS2 and eukaryotic initiation factor 3a (eIF3a). In the absence of Asc1, the interaction between eIF3a and Rps0A/uS2 was compromised. The interaction between Rps0A/uS2 and eIF3a mediated the 40S ribosomal subunit binding of eIF3 in 43S preinitiation complex formation to stimulate translation initiation. Keeping with this idea, in an asc1 mutant, the association of mRNA with the 40S ribosomal subunit was defective and protein synthesis was compromised. To show that Asc1 is directly involved in translation, we demonstrate that the addition of recombinant Asc1 is able to rescue the translation defect of the asc1 mutant in a cell-free system. Furthermore, this function of Asc1 is likely to be evolutionarily conserved, as a similar interaction with eIF3a and Rps0A/uS2 could be identified in the budding yeast Saccharomyces cerevisiae and human aminoacyl-tRNA synthetase cofactors. Together, these results identify a function of aminoacyl-tRNA synthetase cofactors in translation preinitiation complex formation, which adds significantly to the expanded functions associated with aminoacyl-tRNA synthetases and their cofactors.

Feeding stimulates protein synthesis in muscle and liver of neonatal pigs through an mTOR-dependent process

2000 ◽  
Vol 279 (5) ◽  
pp. E1080-E1087 ◽  
Author(s):  
Scot R. Kimball ◽  
Leonard S. Jefferson ◽  
Hahn V. Nguyen ◽  
Agus Suryawan ◽  
Jill A. Bush ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Complex Formation ◽  
Mammalian Target Of Rapamycin ◽  
Specific Inhibitor ◽  
Initiation Factor ◽  
Ribosomal Protein S6 ◽  
Eif4f Complex ◽  
Dependent Process ◽  
Stimulation Of

Protein synthesis is repressed in both skeletal muscle and liver after a short-term fast and is rapidly stimulated in response to feeding. Previous studies in rats and pigs have shown that the feeding-induced stimulation of protein synthesis is associated with activation of the 70-kDa ribosomal protein S6 kinase (S6K1) as well as enhanced binding of eukaryotic initiation factor eIF4E to eIF4G to form the active eIF4F complex. In cells in culture, hormones and nutrients regulate both of these events through a protein kinase termed the mammalian target of rapamycin (mTOR). In the present study, the involvement of mTOR in the feeding-induced stimulation of protein synthesis in skeletal muscle and liver was examined. Pigs at 7 days of age were fasted for 18 h, and then one-half of the animals were fed. In addition, one-half of the animals in each group were administered rapamycin (0.75 mg/kg) 2 h before feeding. The results reveal that treating 18-h fasted pigs with rapamycin, a specific inhibitor of mTOR, before feeding prevented the activation of S6K1 and the changes in eIF4F complex formation observed in skeletal muscle and liver after feeding. Rapamycin also ablated the feeding-induced stimulation of protein synthesis in liver. In contrast, in skeletal muscle, rapamycin attenuated, but did not prevent, the stimulation of protein synthesis in response to feeding. The results suggest that feeding stimulates hepatic protein synthesis through an mTOR-dependent process involving enhanced eIF4F complex formation and activation of S6K1. However, in skeletal muscle, these two processes may account for only part of the stimulation of protein synthesis, and thus additional steps may be involved in the response.

Blocking eukaryotic initiation factor 4F complex formation does not inhibit the mTORC1-dependent activation of protein synthesis in cardiomyocytes

2009 ◽  
Vol 296 (2) ◽  
pp. H505-H514 ◽  
Author(s):  
Brandon P. H. Huang ◽  
Yanni Wang ◽  
Xuemin Wang ◽  
Zhuren Wang ◽  
Christopher G. Proud
Keyword(s):  
Protein Synthesis ◽  
Complex Formation ◽  
Mammalian Target Of Rapamycin ◽  
Initiation Factor ◽  
Minor Role ◽  
Eukaryotic Initiation Factor ◽  
Eukaryotic Initiation Factor 4E ◽  
Mtorc1 Signaling ◽  
A Minor ◽  
Rapid Activation

Activation of the mammalian target of rapamycin complex 1 (mTORC1) causes the dissociation of eukaryotic initiation factor 4E complex (eIF4E)-binding protein 1 (4E-BP1) from eIF4E, leading to increased eIF4F complex formation. mTORC1 positively regulates protein synthesis and is implicated in several diseases including cardiac hypertrophy, a potentially fatal disorder involving increased cardiomyocyte size. The importance of 4E-BP1 in mTORC1-regulated protein synthesis was investigated by overexpressing 4E-BP1, which blocks eIF4F formation in isolated primary cardiomyocytes without affecting other targets for mTORC1 signaling. Interestingly, blocking eIF4F formation did not impair the degree of activation of overall protein synthesis by the hypertrophic agent phenylephrine (PE), which, furthermore, remained dependent on mTORC1. Overexpressing 4E-BP1 also only had a small effect on PE-induced cardiomyocyte growth. Overexpressing 4E-BP1 did diminish the PE-stimulated synthesis of luciferase encoded by structured mRNAs, confirming that such mRNAs do require eIF4F for their translation in cardiomyocytes. These data imply that the substantial inhibition of cardiomyocyte protein synthesis and growth caused by inhibiting mTORC1 cannot be attributed to the activation of 4E-BP1 or loss of eIF4F complexes. Our data indicate that increased eIF4F formation plays, at most, only a minor role in the mTORC1-dependent activation of overall protein synthesis in these primary cells but is required for the translation of structured mRNAs. Therefore, other mTORC1 targets are more important in the inhibition by rapamycin of the rapid activation of protein synthesis and of cell growth.

scholarly journals The eIF4E-binding proteins are modifiers of cytoplasmic eIF4E relocalization during the heat shock response

2009 ◽  
Vol 296 (5) ◽  
pp. C1207-C1217 ◽  
Author(s):  
R. Sukarieh ◽  
N. Sonenberg ◽  
J. Pelletier
Keyword(s):  
Protein Synthesis ◽  
Binding Proteins ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Eukaryotic Initiation Factor 4E ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Repressor Proteins ◽  
Shock Response ◽  
Rate Limiting

Stress granules (SGs) arise as a consequence of cellular stress, contain stalled translation preinitiation complexes, and are associated with cell survival during environmental insults. SGs are dynamic entities with proteins relocating into and out of them during stress. Among the repertoire of proteins present in SGs is eukaryotic initiation factor 4E (eIF4E), a translation factor required for cap-dependent translation and that regulates a rate-limiting step for protein synthesis. Herein, we demonstrate that localization of eIF4E to SGs is dependent on the presence of a family of repressor proteins, eIF4E-binding proteins (4E-BPs). Our results demonstrate that 4E-BPs regulate the SG localization of eIF4E.

Cytoskeletal proteins associate with components of the ribosomal maturation and translation apparatus in Xenopus stage I oocytes

Zygote ◽  
2014 ◽  
Vol 23 (5) ◽  
pp. 669-682 ◽  
Author(s):  
Loredana Chierchia ◽  
Margherita Tussellino ◽  
Domenico Guarino ◽  
Rosa Carotenuto ◽  
Nadia DeMarco ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Elongation Factor ◽  
Cytoskeletal Proteins ◽  
Stage I ◽  
Initiation Factor ◽  
Perinuclear Region ◽  
Eukaryotic Initiation Factor ◽  
Translation Apparatus ◽  
Ribosomal Stalk

SummaryActin-based cytoskeleton (CSK) and microtubules may bind to RNAs and related molecules implicated in translation. However, many questions remain to be answered regarding the role of cytoskeletal components in supporting the proteins involved in steps in the maturation and translation processes. Here, we performed co-immunoprecipitation and immunofluorescence to examine the association between spectrins, keratins and tubulin and proteins involved in 60S ribosomal maturation and translation in Xenopus stage I oocytes, including ribosomal rpl10, eukaryotic initiation factor 6 (Eif6), thesaurins A/B, homologs of the eEF1α elongation factor, and P0, the ribosomal stalk protein. We found that rpl10 and eif6 cross-reacted with the actin-based CSK and with tubulin. rpl10 co-localizes with spectrin, particularly in the perinuclear region. eif6 is similarly localized. Given that upon ribosomal maturation, the insertion of rpl10 into the 60S subunit occurs simultaneously with the release of eif6, one can hypothesise that actin-based CSK and microtubules provide the necessary scaffold for the insertion/release of these two molecules and, subsequently, for eif6 transport and binding to the mature 60S subunit. P0 and thesaurins cross-reacted with only spectrin and cytokeratins. Thesaurins aggregated at the oocyte periphery, rendering this a territory favourable site for protein synthesis; the CSK may support the interaction between thesaurins and sites of the translating ribosome. Moreover, given that the assembly of the ribosome stalk, where P0 is located, to the 60S subunit is essential for the release of eif6, it can be hypothesised that the CSK can facilitate the binding of the stalk to the 60S.

Sign in / Sign up

Export Citation Format

Share Document