scholarly journals Structure of a human 48S translational initiation complex

Science ◽  
2020 ◽  
Vol 369 (6508) ◽  
pp. 1220-1227 ◽  
Author(s):  
Jailson Brito Querido ◽  
Masaaki Sokabe ◽  
Sebastian Kraatz ◽  
Yuliya Gordiyenko ◽  
J. Mark Skehel ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Messenger Rna ◽  
Initiation Factor ◽  
Start Codon ◽  
Exit Channel ◽  
Eukaryotic Initiation Factor ◽  
Initiation Complex ◽  
Translational Initiation ◽  
Cryo Electron Microscopy ◽  
Cap Binding Complex

A key step in translational initiation is the recruitment of the 43S preinitiation complex by the cap-binding complex [eukaryotic initiation factor 4F (eIF4F)] at the 5′ end of messenger RNA (mRNA) to form the 48S initiation complex (i.e., the 48S). The 48S then scans along the mRNA to locate a start codon. To understand the mechanisms involved, we used cryo–electron microscopy to determine the structure of a reconstituted human 48S. The structure reveals insights into early events of translation initiation complex assembly, as well as how eIF4F interacts with subunits of eIF3 near the mRNA exit channel in the 43S. The location of eIF4F is consistent with a slotting model of mRNA recruitment and suggests that downstream mRNA is unwound at least in part by being “pulled” through the 40S subunit during scanning.

scholarly journals Structures of ribosome-bound initiation factor 2 reveal the mechanism of subunit association

2016 ◽  
Vol 2 (3) ◽  
pp. e1501502 ◽  
Author(s):  
Thiemo Sprink ◽  
David J. F. Ramrath ◽  
Hiroshi Yamamoto ◽  
Kaori Yamamoto ◽  
Justus Loerke ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Protein Biosynthesis ◽  
Initiation Factor ◽  
Structural Basis ◽  
Initiation Complex ◽  
Cryo Electron Microscopy ◽  
Initiation Factor 2 ◽  
High Resolution Structure ◽  
Guanosine Triphosphatase ◽  
Resolution Structure

Throughout the four phases of protein biosynthesis—initiation, elongation, termination, and recycling—the ribosome is controlled and regulated by at least one specified translational guanosine triphosphatase (trGTPase). Although the structural basis for trGTPase interaction with the ribosome has been solved for the last three steps of translation, the high-resolution structure for the key initiation trGTPase, initiation factor 2 (IF2), complexed with the ribosome, remains elusive. We determine the structure of IF2 complexed with a nonhydrolyzable guanosine triphosphate analog and initiator fMet-tRNAiMet in the context of the Escherichia coli ribosome to 3.7-Å resolution using cryo-electron microscopy. The structural analysis reveals previously unseen intrinsic conformational modes of the 70S initiation complex, establishing the mutual interplay of IF2 and initator transfer RNA (tRNA) with the ribsosome and providing the structural foundation for a mechanistic understanding of the final steps of translation initiation.

scholarly journals The complete, functional and dynamic cycle of the bacterial Initiation Factor 3

2019 ◽  
Author(s):  
Jose A. Nakamoto ◽  
Roberto Spurio ◽  
Andrey L. Konevega ◽  
Attilio Fabbretti ◽  
Pohl Milón
Keyword(s):  
Complex Formation ◽  
Translation Initiation ◽  
Binding Sites ◽  
Mrna Translation ◽  
Initiation Factor ◽  
Start Codon ◽  
Initiation Complex ◽  
Terminal Domain ◽  
Dynamic Interplay ◽  
Dynamic Cycle

AbstractInitiation factor 3 (IF3) is an essential protein that enhances the fidelity and speed of bacterial initiation of mRNA translation. The dynamic interplay between the two independent IF3 domains, their alternative binding sites, and the mechanism that ensures translation initiation fidelity remains elusive. Here, we show that the functional positioning of IF3 domains occurs at velocities ranging over two orders of magnitude, driven by each 30S initiation ligand. IF1 and IF2 rapidly promote the accommodation of IF3 on the 30S platform with the C-terminal domain moving towards the P site. Reversion of this movement is triggered by decoding the mRNA start codon and rate limits translation initiation. Binding of the tRNA results in the concomitant accommodation of the N-terminal domain of IF3, largely dependent on the mRNA and initiator tRNA. 70S initiation complex formation promotes the closing and dissociation of IF3, recycling the factor for a new round of translation initiation. Altogether our results unveil the kinetic spectrum of IF3 conformations and highlight fundamental movements of the factor that ensure accurate translation initiation.

EARLY STEPS IN THE FORMATION OF A TRANSLATION INITIATION COMPLEX ON NEWLY TRANSCRIBED MESSENGER RNA

1973 ◽  
Vol 74 (Suppl) ◽  
pp. S33-S53 ◽  
Author(s):  
Michel Crépin ◽  
Jean-Claude Lelong ◽  
François Gros
Keyword(s):  
Translation Initiation ◽  
Transcription Initiation ◽  
Messenger Rna ◽  
Rna Synthesis ◽  
Initiation Factor ◽  
Initiation Complex ◽  
Initiator Trna ◽  
Transcription System ◽  
Run Off

ABSTRACT The aim of this study is to investigate how formation of a translation initiation complex affects the rate of transcription from a phage DNA template (λ plac or Ø 80 dlac DNA). Addition of "native" 30S ribosomes to a Ø 80 dlac in vitro transcription system (including limiting amounts of purified E. coli RNA polymerase and the requisite substrates) markedly enhances the rate of RNA synthesis. Factor free, 1 M NH4Cl washed 30S or 70S ribosomes show a weak albeit detectable activity, "run off 70S" or washed 50S being inefficient. Single addition of purified initiation factor IF3 greatly magnifies (about 2–3 fold) the stimulation obtained with washed 30S subunits, the effect being catalytic with respect to IF3; IF2 addition causes a weaker and stoichiometrical effect. Maximum stimulation (up to 6 fold) is achieved by the combined addition of washed ribosomes (30S or 70S), IF1, IF2, IF3 plus the highly purified fMet-tRNAfMet species. Under such conditions, very efficient initiator tRNA binding to nascent RNA does occur. Initiation factors show no activity in the absence of ribosomes. Kasugamycin greatly reduces the stimulation of RNA synthesis in the presence of the various translation elements. This system provides a new and very sensitive means to study the various factor dependent ribosome-messenger interactions even in the absence of initiator tRNA, thereby enabling one to analyze early translation initiation steps. Both the frequency of transcription initiation and the rate of RNA chain propagation appear to be enhanced when RNA synthesis and initiation of protein synthesis are coupled.

scholarly journals OseIF3h Regulates Plant Growth and Pollen Development at Translational Level Presumably through Interaction with OsMTA2

Plants ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1101
Author(s):  
Yuqing Huang ◽  
Peng Zheng ◽  
Xuejiao Liu ◽  
Hao Chen ◽  
Jumin Tu
Keyword(s):  
Plant Growth ◽  
Translation Initiation ◽  
Pollen Development ◽  
Messenger Rna ◽  
Protein Biosynthesis ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Sequencing Data ◽  
Knock Out ◽  
Translational Level

The initiation stage of protein biosynthesis is a sophisticated process tightly regulated by numerous initiation factors and their associated components. However, the mechanism underlying translation initiation has not been completely understood in rice. Here, we showed knock-out mutation of the rice eukaryotic translation initiation factor 3 subunit h (OseIF3h) resulted in plant growth retardation and seed-setting rate reduction as compared to the wild type. Further investigation demonstrated an interaction between OseIF3h and OsMTA2 (mRNA adenosine methylase 2), a rice homolog of METTL3 (methyltransferase-like 3) in mammals, which provided new insight into how N6-methyladenosine (m6A) modification of messenger RNA (mRNA) is engaged in the translation initiation process in monocot species. Moreover, the RIP-seq (RNA immunoprecipitation sequencing) data suggested that OseIF3h was involved in multiple biological processes, including photosynthesis, cellular metabolic process, precursor metabolites, and energy generation. Therefore, we infer that OseIF3h interacts with OsMTA2 to target a particular subset of genes at translational level, regulating plant growth and pollen development.

scholarly journals POLARITY IN SEGMENTS OF THE ESCHERICHIA COLI trp OPERON WITH DELETED INTRAOPERONIC TRANSLATIONAL INITIATION SIGNALS

Genetics ◽  
1975 ◽  
Vol 80 (4) ◽  
pp. 651-666
Author(s):  
Yasunobu Kano ◽  
Fumio Imamoto
Keyword(s):  
Escherichia Coli ◽  
Translation Initiation ◽  
Nonsense Mutation ◽  
Messenger Rna ◽  
Distal Part ◽  
High Efficiency ◽  
Translational Initiation ◽  
Transcriptional Termination ◽  
Trp Operon

ABSTRACT The effect of deletion of the operator-distal genes of the trp operon, including the trpE-trpD intercistronic punctuation point, on the degree of transcriptional polarity (in this case the effect of a nonsense mutation on the level of mRNA from the distal part of the very gene where the mutation is located) was investigated. Double mutants which contain a nonsense mutation and a deletion in trpE were constructed, and the degree of transcriptional polarity was estimated by the decrease in messenger RNA for the operator-distal trpE beyond the nonsense mutation, as well as by the production of truncated messenger RNA for the region of trpE proximal to the nonsense mutation. The content of mRNA of operator-distal trpE and the size of the mRNA of operator-proximal trpE of the double mutants show that transcriptional polarity is not relaxed as a function of distance of the nonsense mutation from the operator-distal end of the trpE segment (at which the subsequent high efficiency translational initiation signal has been deleted). These findings are consistent with the conclusion that the degree of polarity depends on the distance of the nonsense mutation fro mthe subsequent translation initiation signal, but not on its distance from the operator-distal end, including possible translational or transcriptional termination signals

scholarly journals Cap-binding protein (eukaryotic initiation factor 4E) and 4E-inactivating protein BP-1 independently regulate cap-dependent translation.

1996 ◽  
Vol 16 (10) ◽  
pp. 5450-5457 ◽  
Author(s):  
D Feigenblum ◽  
R J Schneider
Keyword(s):  
Heat Shock ◽  
Translation Initiation ◽  
Binding Protein ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Metaphase Arrest ◽  
Animal Cells ◽  
Eukaryotic Initiation Factor 4E ◽  
Dependent Protein

Cap-dependent protein synthesis in animal cells is inhibited by heat shock, serum deprivation, metaphase arrest, and infection with certain viruses such as adenovirus (Ad). At a mechanistic level, translation of capped mRNAs is inhibited by dephosphorylation of eukaryotic initiation factor 4E (eIF-4E) (cap-binding protein) and its physical sequestration with the translation repressor protein BP-1 (PHAS-I). Dephosphorylation of BP-I blocks cap-dependent translation by promoting sequestration of eIF-4E. Here we show that heat shock inhibits translation of capped mRNAs by simultaneously inducing dephosphorylation of eIF-4E and BP-1, suggesting that cells might coordinately regulate translation of capped mRNAs by impairing both the activity and the availability of eIF-4E. Like heat shock, late Ad infection is shown to induce dephosphorylation of eIF-4E. However, in contrast to heat shock, Ad also induces phosphorylation of BP-1 and release of eIF-4E. BP-1 and eIF-4E can therefore act on cap-dependent translation in either a mutually antagonistic or cooperative manner. Three sets of experiments further underscore this point: (i) rapamycin is shown to block phosphorylation of BP-1 without inhibiting dephosphorylation of eIF-4E induced by heat shock or Ad infection, (ii) eIF-4E is efficiently dephosphorylated during heat shock or Ad infection regardless of whether it is in a complex with BP-1, and (iii) BP-1 is associated with eIF-4E in vivo regardless of the state of eIF-4E phosphorylation. These and other studies establish that inhibition of cap-dependent translation does not obligatorily involve sequestration of eIF-4E by BP-1. Rather, translation is independently regulated by the phosphorylation states of eIF-4E and the 4E-binding protein, BP-1. In addition, these results demonstrate that BP-1 and eIF-4E can act either in concert or in opposition to independently regulate cap-dependent translation. We suggest that independent regulation of eIF-4E and BP-1 might finely regulate the efficiency of translation initiation or possibly control cap-dependent translation for fundamentally different purposes.

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