Translation of Eukaryotic Translation Initiation Factor 4GI (eIF4GI) Proceeds from Multiple mRNAs Containing a Novel Cap-dependent Internal Ribosome Entry Site (IRES) That Is Active during Poliovirus Infection

Byrd, Marshall P.; Zamora, Miguel; Lloyd, Richard E.

doi:10.1074/jbc.m414014200

Cited by 56 publications

(53 citation statements)

References 58 publications

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“…Curiously, however, the internal ribosome entry site-mediated translation of eIF4GI still demonstrates cap-dependence. 15 It has also been suggested that rapamycin causes decreased transcription of the eIF4G gene and, additionally, can enhance the rate of proteolysis of the initiation factor. 16,17 We cannot formally rule out the possibility that rapamycin regulates the expression of eIF4GI by such mechanisms.…”

Section: Discussionmentioning

confidence: 99%

Regulation of translation factors eIF4GI and 4E-BP1 during recovery of protein synthesis from inhibition by p53

Constantinou

Clemens

2006

Cell Death Differ

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Activation of the tumour suppressor protein p53 rapidly inhibits protein synthesis. This is associated with dephosphorylation and cleavage of initiation factor eIF4GI and the eIF4E-binding protein 4E-BP1. When the activation of p53 is reversed within 16 h 4E-BP1 becomes rephosphorylated, the level of intact eIF4GI slowly increases and protein synthesis gradually recovers. The recovery of protein synthesis is partially blocked by rapamycin and wortmannin but not by the protein kinase inhibitors PD98059 and CGP74514A. Both rapamycin and wortmannin, but not PD98059 or CGP74514A, delay the reappearance of eIF4GI. In contrast, full-length 4E-BP1 rapidly becomes rephosphorylated and this process is partially inhibited by rapamycin, PD98059 and CGP74514A. Thus, activation of p53 results in the inhibition of distinct rapamycin-and wortmannin-sensitive pathways that target eIF4GI, and rapamycin-sensitive and -insensitive pathways that target 4E-BP1. Following inactivation of p53 the gradual recovery is determined largely by the kinetics of restoration of eIF4GI rather than by the rephosphorylation of full-length 4E-BP1. These findings suggest that the ability of cells to rephosphorylate 4E-BP1, resynthesise eIF4GI and restore the rate of protein synthesis after inactivation of p53 is an important aspect of recovery following the relief of physiological stress. Activation of the tumour suppressor protein p53 causes a rapid inhibition of protein synthesis which precedes the induction of a cell cycle block and subsequent apoptosis. 1-3The inhibition of translation is accompanied by early dephosphorylation and cleavage of the eukaryotic initiation factor (eIF)4E-binding protein 4E-BP1 as well as cleavage of polypeptide chain initiation factor eIF4GI and dephosphorylation of this protein at Ser 1108 . 3 The cleaved products of 4E-BP1 and eIF4GI, which are generated by a caspaseindependent mechanism, associate with eIF4E and there is a correspondingly decreased extent of interaction of eIF4E with full-length eIF4GI. These changes are likely to play an important role in impairing the initiation of protein synthesis.In this study, we have examined whether the inhibition of protein synthesis can be reversed when a temperaturesensitive form of p53 is inactivated, prior to the irreversible commitment to cell death, 2 by return of the cells to the non-permissive temperature (381C). We have also investigated the requirement for various signalling pathways that are known to regulate protein synthesis and have analysed the integrity and state of phosphorylation of eIF4GI and 4E-BP1 during the recovery phase. The results indicate a multifactorial mode of regulation of translation following the inactivation of p53 that is partially dependent on the activities of the protein kinase mammalian target of rapamycin (mTOR) and phosphatidylinositol-3-kinase (PI3k) but is independent of the protein kinases Mek1/2 and cdc2. Furthermore, our findings suggest that the rate of recovery of protein synthesis is limited by the kinetics of reappearance ...

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Section: Discussionmentioning

confidence: 99%

Regulation of translation factors eIF4GI and 4E-BP1 during recovery of protein synthesis from inhibition by p53

Constantinou

Clemens

2006

Cell Death Differ

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“…Indeed it has been shown that in a Krebs-2 in vitro system, or in an RNA transfection assay, an mRNA with the 900 nt (60% GC) LINE-1 5 0 -UTR, is translated by the cap-dependent scanning mechanism at 50% relative efficiency compared to the b-globin 5 0 -UTR (Dmitriev et al 2007). It should also be noted that some long 5 0 -UTRs with apparent IRES activity have subsequently been shown to be incompletely spliced variants with retained introns (Baranick et al 2008), for example the original eIF4GI 5 0 -UTR studied by Gan and Rhoads (1996), in contrast to the eIF4GI 5 0 -UTRs described by Johannes and Sarnow (1998) and Byrd et al (2005).…”

Section: Cellular Mrna Iressmentioning

confidence: 99%

The Current Status of Vertebrate Cellular mRNA IRESs

Jackson

2013

Cold Spring Harbor Perspectives in Biology

137

130

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Internal ribosome entry sites/segments (IRESs) were first discovered over 20 years ago in picornaviruses, followed by the discovery of two other types of IRES in hepatitis C virus (HCV), and the dicistroviruses, which infect invertebrates. In the meantime, reports of IRESs in eukaryotic cellular mRNAs started to appear, and the list of such putative IRESs continues to grow to the point in which it now stands at 100, 80% of them in vertebrate mRNAs. Despite initial skepticism from some quarters, there now seems universal agreement that there is genuine internal ribosome entry on the viral IRESs. However, the same cannot be said for cellular mRNA IRESs, which continue to be shrouded in controversy. The aim of this article is to explain why vertebrate mRNA IRESs remain controversial, and to discuss ways in which these controversies might be resolved.

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“…The 3Ј ss in the putative eIF4G IRES is known to be used in formation of several of the eIF4G transcript variants (SI Fig. 12 A) (25). In transcript variant 5, which contains the proposed IRES and is transcribed from its own promoter, the 3Ј ss is not thought to be used.…”

Section: Six Putative Cellular Iress Exhibit Little Activity In a Dicmentioning

confidence: 99%

Splicing mediates the activity of four putative cellular internal ribosome entry sites

Baranick¹,

Lemp²,

Nagashima³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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A growing number of cellular mRNAs are thought to possess internal ribosome entry sites (IRESs), sequences that permit translation of a transcript independent of its 5 end and cap structure. Although dicistronic assays are the canonical method of testing sequences for IRES activity, they may produce false-positive results if unanticipated monocistronic RNAs arise from the dicistronic construct used. Using a dicistronic reporter system and a green fluorescent protein-tagged retrovirus to evaluate six previously reported cellular IRESs, we found that four contain 3 splice sites whose activity was required for apparent IRES function and which resulted in formation of monocistronic transcripts by splicing. Bioinformatic analysis revealed that the 3 splice sites identified in three of these putative IRESs are used in their native mRNAs and that the fourth is likely an artifactual sequence created during cDNA cloning. Our findings demonstrate a need for reexamination of other reported cellular IRESs by using careful RNA structural analysis to rule out splicing as the source of perceived IRES activity.T he vast majority of eukaryotic mRNAs are translated via a mechanism in which the 40S ribosomal subunit engages the mRNA at its methylguanosine-capped 5Ј end (1). Upon associating with the transcript, these subunits are believed then to scan in the 5Ј to 3Ј direction for an appropriately situated AUG at which to begin translation (2, 3). A smaller number of mRNAs are translated by a 5Ј end-and cap-independent mechanism wherein ribosomes are recruited to the transcript at an interior location through an internal ribosome entry site (IRES).IRESs were first discovered in the picornaviruses encephalomyocarditis virus (EMCV) and poliovirus (4, 5). The RNA of these viruses possesses very long 5Ј UTRs bearing many unutilized upstream AUGs (uAUGs) and, unlike cellular mRNA, is uncapped (6). Soon after the identification of picornaviral IRESs, a number of cellular mRNAs were also reported to contain IRESs. To date, at least 85 cellular IRESs have been described (7). The experimental grounds on which proof of most cellular IRESs rest, however, has been the subject of dispute (8-10).A primary criticism of the data presented as establishing the existence of cellular IRESs concerns the plasmid-based dicistronic assay, the standard method of ascertaining IRES activity. In this assay, the candidate sequence is inserted between two reporter genes (5) so that both the upstream and downstream cistron are transcribed on the same RNA. If the test insert causes increased expression of the downstream cistron relative to the upstream cistron, the result is considered evidence for internal ribosome entry. However, the generation of even low levels of monocistronic RNAs from dicistronic constructs has the potential to falsely indicate IRES activity (8, 9, 11). One way that such RNAs could arise is through splicing of the dicistronic transcript due to the presence of a 3Ј splice site (ss) in the test sequence [see supporting information (SI) Fig....

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Translation of Eukaryotic Translation Initiation Factor 4GI (eIF4GI) Proceeds from Multiple mRNAs Containing a Novel Cap-dependent Internal Ribosome Entry Site (IRES) That Is Active during Poliovirus Infection

Cited by 56 publications

References 58 publications

Regulation of translation factors eIF4GI and 4E-BP1 during recovery of protein synthesis from inhibition by p53

Regulation of translation factors eIF4GI and 4E-BP1 during recovery of protein synthesis from inhibition by p53

The Current Status of Vertebrate Cellular mRNA IRESs

Splicing mediates the activity of four putative cellular internal ribosome entry sites

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