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.

scholarly journals Effects of starvation, diabetes and acute insulin treatment on the regulation of polypeptide-chain initiation in rat skeletal muscle

1984 ◽  
Vol 223 (3) ◽  
pp. 687-696 ◽  
Author(s):  
C S Harmon ◽  
C G Proud ◽  
V M Pain
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Polypeptide Chain ◽  
Ribosomal Subunit ◽  
Diabetic Rats ◽  
Initiation Factor ◽  
Chain Initiation ◽  
Low Concentrations ◽  
40S Ribosomal Subunit

The rate of protein synthesis in skeletal muscle is greatly decreased in response to diabetes and starvation. Analysis of polyribosome profiles indicates that polypeptide-chain initiation is impaired under these conditions. To identify the step in initiation that is affected, we assayed the incorporation of [35S]methionyl-tRNAfMet into [35S]methionyl-tRNAfMet . 40S-ribosomal-subunit initiation complexes in cell-free extracts based on postmitochondrial supernatants prepared from gastrocnemius muscle. Extracts from either starved or diabetic rats were 30-40% less active in forming these complexes compared with those derived from fed or insulin-maintained controls respectively. This change could be reversed by treatment of either starved or diabetic rats with insulin in vivo 30 min before death. Formation of 40S initiation complexes by extracts from either fed or starved rats could be stimulated by the addition of exogenous purified initiation factor eIF-2, but extracts from starved or diabetic rats were more sensitive than controls to stimulation by low concentrations of the factor. These results provide evidence for the acute regulation by insulin of protein synthesis in skeletal muscle at the level of polypeptide-chain initiation, and suggest that in this tissue, as in certain other eukaryotic systems, control of initiation appears to be mediated by changes in the activity of initiation factor eIF-2.

Nuclear localization of initiation factor 2 in neuron primary cultures

1995 ◽  
Vol 53 ◽  
pp. 774-775
Author(s):  
F.J. Martinez Alonso ◽  
M.V. Toledo Lobo ◽  
S. Rodriguez Martínez ◽  
F.M. Muñoz Postigo ◽  
J.J. López-Fando Castro
Keyword(s):  
Protein Synthesis ◽  
Translational Control ◽  
Control Function ◽  
Primary Cultures ◽  
Ribosomal Subunit ◽  
Initiation Factor ◽  
40S Ribosomal Subunit ◽  
Initiation Factor 2 ◽  
Immunocytochemical Techniques ◽  
Embryo Brain

The dominant mechanism that controls protein synthesis is the phosphorylation/dephosphorylation of initiation and elongation factors, with a translational control function. Each phase of protein synthesis is promoted by some of these factors that transiently interact with ribosomes, mRNAs and aminoacyltRNAs. Eukaryotic initiation factor-2 (eIF2, 130 kD) is one of these proteins and it is composed of three subunits: alpha, beta and gamma. eIF2 forms a ternary complex (GTP-eIF2-Met tRNAi) that can then interact with the 40S ribosomal subunit which in turn binds mRNA and the 60S ribosomal subunit to form the 80S initiation complex. The relation between eIF2 and the ribosomes is then a well established aspect of protein synthesis, but there are no previous studies about the distribution of eIF2 within the cell.Using immunocytochemical techniques, we show the distribution of eIF2 within the cell found in primary cultures of rat embryo brain neurons, in which eIF2 and eIF2-kinases have been identified. Primary culture neuron cells were grown in D15 and N2 mediums for 8 days.

scholarly journals Translation initiation at non-AUG codons mediated by weakened association of eukaryotic initiation factor (eIF) 2 subunits

2002 ◽  
Vol 367 (2) ◽  
pp. 359-368 ◽  
Author(s):  
Nilce N. HASHIMOTO ◽  
Larissa S. CARNEVALLI ◽  
Beatriz A. CASTILHO
Keyword(s):  
Translation Initiation ◽  
Ribosomal Subunit ◽  
Initiation Factor ◽  
Temperature Sensitive ◽  
Amino Acid Residues ◽  
Eukaryotic Initiation Factor ◽  
Strength Of Interaction ◽  
40S Ribosomal Subunit ◽  
Initiator Codon

The heterotrimeric eukaryotic initiation factor (eIF) 2 binds the initiator methionyl-tRNA in a GTP-dependent mode and delivers it to the 40S ribosomal subunit. In the present study, we have identified amino acid residues in eIF2β required for binding to eIF2γ in yeast. Alteration of six residues in the central region of eIF2β abolished this interaction, as determined by GST-pull down and two-hybrid assays, and leads to cell lethality. Substitution of 131Tyr and 132Ser by alanine residues (131YS), although abolishing the binding to eIF2γ in these assays, resulted in a functional but defective protein in vivo, imparting a temperature-sensitive growth phenotype to cells. A dramatically weakened association of this mutant protein with eIF2γ in vivo was shown by co-immunoprecipitation. The 131YS mutation in eIF2β allows translation to initiate at non-AUG codons, as defined by the ability of cells carrying an initiator codon mutation in the HIS4 mRNA to grow in the absence of histidine. The combination of this mutation with the 264Ser→Tyr alteration, a previously isolated suppressor of initiator codon mutations which has been shown to increase the spontaneous GTP hydrolysis in the ternary complex, caused a recessive lethality, suggesting additive defects. Thus the impaired interaction of these two subunits represents a novel type of defect in eIF2 function, providing in vivo evidence that the strength of interaction between eIF2β and eIF2γ defines the correct usage of the AUG codon for translation initiation.

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.

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