scholarly journals The DEAD-Box RNA Helicase Ded1p Affects and Accumulates inSaccharomyces cerevisiaeP-Bodies

2008 ◽  
Vol 19 (3) ◽  
pp. 984-993 ◽  
Author(s):  
Carla Beckham ◽  
Angela Hilliker ◽  
Anne-Marie Cziko ◽  
Amine Noueiry ◽  
Mani Ramaswami ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Processing Bodies ◽  
Body Formation ◽  
P Bodies ◽  
Dead Box ◽  
Inhibition Of Growth ◽  
Messenger Ribonucleoprotein ◽  
Positive Effect

Recent results suggest that cytoplasmic mRNAs can form translationally repressed messenger ribonucleoprotein particles (mRNPs) capable of decapping and degradation, or accumulation into cytoplasmic processing bodies (P-bodies), which can function as sites of mRNA storage. The proteins that function in transitions between the translationally repressed mRNPs that accumulate in P-bodies and mRNPs engaged in translation are largely unknown. Herein, we demonstrate that the yeast translation initiation factor Ded1p can localize to P-bodies. Moreover, depletion of Ded1p leads to defects in P-body formation. Overexpression of Ded1p results in increased size and number of P-bodies and inhibition of growth in a manner partially suppressed by loss of Pat1p, Dhh1p, or Lsm1p. Mutations that inactivate the ATPase activity of Ded1p increase the overexpression growth inhibition of Ded1p and prevent Ded1p from localizing in P-bodies. Combined with earlier work showing Ded1p can have a positive effect on translation, these results suggest that Ded1p is a bifunctional protein that can affect both translation initiation and P-body formation.

scholarly journals Fal1p is an essential DEAD-box protein involved in 40S-ribosomal-subunit biogenesis in Saccharomyces cerevisiae.

1997 ◽  
Vol 17 (12) ◽  
pp. 7283-7294 ◽  
Author(s):  
D Kressler ◽  
J de la Cruz ◽  
M Rojo ◽  
P Linder
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Translation Initiation ◽  
18S Rrna ◽  
Amino Acid Level ◽  
Ribosomal Subunit ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Northern Hybridization ◽  
Ribosomal Subunits ◽  
Dead Box

A previously uncharacterized Saccharomyces cerevisiae gene, FAL1, was found by sequence comparison as a homolog of the eukaryotic translation initiation factor 4A (eIF4A). Fal1p has 55% identity and 73% similarity on the amino acid level to yeast eIF4A, the prototype of ATP-dependent RNA helicases of the DEAD-box protein family. Although clearly grouped in the eIF4A subfamily, the essential Fal1p displays a different subcellular function and localization. An HA epitope-tagged Fal1p is localized predominantly in the nucleolus. Polysome analyses in a temperature-sensitive fal1-1 mutant and a Fal1p-depleted strain reveal a decrease in the number of 40S ribosomal subunits. Furthermore, these strains are hypersensitive to the aminoglycoside antibiotics paromomycin and neomycin. Pulse-chase labeling of pre-rRNA and steady-state-level analysis of pre-rRNAs and mature rRNAs by Northern hybridization and primer extension in the Fal1p-depleted strain show that Fal1p is required for pre-rRNA processing at sites A0, A1, and A2. Consequently, depletion of Fal1p leads to decreased 18S rRNA levels and to an overall deficit in 40S ribosomal subunits. Together, these results implicate Fal1p in the 18S rRNA maturation pathway rather than in translation initiation.

Acidic stress induces the formation of P-bodies, but not stress granules, with mild attenuation of bulk translation in Saccharomyces cerevisiae

2012 ◽  
Vol 446 (2) ◽  
pp. 225-233 ◽  
Author(s):  
Aya Iwaki ◽  
Shingo Izawa
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Lactic Acid ◽  
Stress Granules ◽  
Glucose Deprivation ◽  
Acidic Stress ◽  
Processing Bodies ◽  
Body Formation ◽  
P Bodies ◽  
Glucose Depletion ◽  
Messenger Ribonucleoprotein

The stress response of eukaryotic cells often causes an attenuation of bulk translation activity and the accumulation of non-translating mRNAs into cytoplasmic mRNP (messenger ribonucleoprotein) granules termed cytoplasmic P-bodies (processing bodies) and SGs (stress granules). We examined effects of acidic stress on the formation of mRNP granules compared with other forms of stress such as glucose deprivation and a high Ca2+ level in Saccharomyces cerevisiae. Treatment with lactic acid clearly caused the formation of P-bodies, but not SGs, and also caused an attenuation of translation initiation, albeit to a lesser extent than glucose depletion. P-body formation was also induced by hydrochloric acid and sulfuric acid. However, lactic acid in SD (synthetic dextrose) medium with a pH greater than 3.0, propionic acid and acetic acid did not induce P-body formation. The results of the present study suggest that the assembly of yeast P-bodies can be induced by external conditions with a low pH and the threshold was around pH 2.5. The P-body formation upon acidic stress required Scd6 (suppressor of clathrin deficiency 6), a component of P-bodies, indicating that P-bodies induced by acidic stress have rules of assembly different from those induced by glucose deprivation or high Ca2+ levels.

scholarly journals eIF4B stimulates translation of long mRNAs with structured 5′ UTRs and low closed-loop potential but weak dependence on eIF4G

2016 ◽  
Vol 113 (38) ◽  
pp. 10464-10472 ◽  
Author(s):  
Neelam Dabas Sen ◽  
Fujun Zhou ◽  
Michael S. Harris ◽  
Nicholas T. Ingolia ◽  
Alan G. Hinnebusch
Keyword(s):  
Translation Initiation ◽  
Closed Loop ◽  
Ribosome Profiling ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Rna Helicases ◽  
Eukaryotic Translation Initiation Factor ◽  
Eukaryotic Translation ◽  
Messenger Ribonucleoprotein ◽  
Eukaryotic Translation Initiation

DEAD-box RNA helicases eukaryotic translation initiation factor 4A (eIF4A) and Ded1 promote translation by resolving mRNA secondary structures that impede preinitiation complex (PIC) attachment to mRNA or scanning. Eukaryotic translation initiation factor 4B (eIF4B) is a cofactor for eIF4A but also might function independently of eIF4A. Ribosome profiling of mutants lacking eIF4B or with impaired eIF4A or Ded1 activity revealed that eliminating eIF4B reduces the relative translational efficiencies of many more genes than does inactivation of eIF4A, despite comparable reductions in bulk translation, and few genes display unusually strong requirements for both factors. However, either eliminating eIF4B or inactivating eIF4A preferentially impacts mRNAs with longer, more structured 5′ untranslated regions (UTRs). These findings reveal an eIF4A-independent role for eIF4B in addition to its function as eIF4A cofactor in promoting PIC attachment or scanning on structured mRNAs. eIF4B, eIF4A, and Ded1 mutations also preferentially impair translation of longer mRNAs in a fashion mitigated by the ability to form closed-loop messenger ribonucleoprotein particles (mRNPs) via eIF4F–poly(A)-binding protein 1 (Pab1) association, suggesting cooperation between closed-loop assembly and eIF4B/helicase functions. Remarkably, depleting eukaryotic translation initiation factor 4G (eIF4G), the scaffold subunit of eukaryotic translation initiation factor 4F (eIF4F), preferentially impacts short mRNAs with strong closed-loop potential and unstructured 5′ UTRs, exactly the opposite features associated with hyperdependence on the eIF4B/helicases. We propose that short, highly efficient mRNAs preferentially depend on the stimulatory effects of eIF4G-dependent closed-loop assembly.

Investigation of the conserved glutamate immediately following the DEAD box in eukaryotic translation initiation factor 4AI

2014 ◽  
Vol 92 (1) ◽  
pp. 33-42
Author(s):  
Krishnaben Patel ◽  
Grishma K. Shah ◽  
Sai Shilpa Kommaraju ◽  
Woon-Kai Low
Keyword(s):  
Atpase Activity ◽  
Translation Initiation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Eukaryotic Translation Initiation Factor ◽  
Eukaryotic Translation ◽  
Dependent Atpase ◽  
Binding Domains ◽  
Dead Box ◽  
Eukaryotic Translation Initiation

The DExD-box family (DEAD-box) of proteins was surveyed for eukaryotic translation initiation factor 4A-specific sequences surrounding the DEAD box. An eIF4A-unique glutamate residue (E186 in eIF4AI) was identified immediately following the D-E-A-D sequence in eIF4AI, II, and III that was found to be conserved from yeast to Man. Mutation to a selection of alternative amino acids was performed within recombinant eIF4AI expressed in Escherichia coli and mutant proteins were surveyed for RNA-dependent ATPase activity. The mutants were also investigated for changes in activity in the presence of the two eIF4AI-binding domains of eIF4GI as well as for co-purification ability to these two domains. The E186 residue was found to be of significance for RNA-dependent ATPase activity for eIF4AI alone and in the presence of eIF4AI-binding domains of eIF4GI through point-mutation analysis. Furthermore, binding interactions between eIF4AI and eIF4GI domains were also significantly influenced by mutation of E186, as observed through co-purification assays. Thus, this residue appears to be of functional significance for eIF4A.

scholarly journals A role for the eIF4E-binding protein 4E-T in P-body formation and mRNA decay

2005 ◽  
Vol 170 (6) ◽  
pp. 913-924 ◽  
Author(s):  
Maria A. Ferraiuolo ◽  
Sanjukta Basak ◽  
Josee Dostie ◽  
Elizabeth L. Murray ◽  
Daniel R. Schoenberg ◽  
...  
Keyword(s):  
Mrna Decay ◽  
Binding Protein ◽  
Initiation Factor ◽  
Binding Motif ◽  
Processing Bodies ◽  
Mrna Decapping ◽  
P Bodies ◽  
Eukaryotic Initiation Factor 4E ◽  
Messenger Ribonucleoprotein ◽  
Bona Fide

4E-transporter (4E-T) is one of several proteins that bind the mRNA 5′cap-binding protein, eukaryotic initiation factor 4E (eIF4E), through a conserved binding motif. We previously showed that 4E-T is a nucleocytoplasmic shuttling protein, which mediates the import of eIF4E into the nucleus. At steady state, 4E-T is predominantly cytoplasmic and is concentrated in bodies that conspicuously resemble the recently described processing bodies (P-bodies), which are believed to be sites of mRNA decay. In this paper, we demonstrate that 4E-T colocalizes with mRNA decapping factors in bona fide P-bodies. Moreover, 4E-T controls mRNA half-life, because its depletion from cells using short interfering RNA increases mRNA stability. The 4E-T binding partner, eIF4E, also is localized in P-bodies. 4E-T interaction with eIF4E represses translation, which is believed to be a prerequisite for targeting of mRNAs to P-bodies. Collectively, these data suggest that 4E-T interaction with eIF4E is a priming event in inducing messenger ribonucleoprotein rearrangement and transition from translation to decay.

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