Reinitiation and Other Unconventional Posttermination Events during Eukaryotic Translation

Skabkin, Maxim A.; Skabkina, Olga V.; Hellen, Christopher U.T.; Pestova, Tatyana V.

doi:10.1016/j.molcel.2013.05.026

Cited by 144 publications

(189 citation statements)

References 52 publications

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“…In eukaryotes (humans and mice), translation of approximately half of the transcripts are regulated by short upstream ORFs (uORFs) (62,63) and translation of the downstream ORF requires reinitiation. Recently, it was shown that after a termination event, 40S subunits, and probably also 80S ribosomes, could scan along the mRNA downstream or upstream to the next AUG position (64). Ribosome profiling methodology also revealed the possibility that 80S ribosomes scan downstream after translating a cistron in yeast (65).…”

Section: Discussionmentioning

confidence: 99%

70S-scanning initiation is a novel and frequent initiation mode of ribosomal translation in bacteria

Yamamoto

Wittek

Gupta

et al. 2016

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

106

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According to the standard model of bacterial translation initiation, the small ribosomal 30S subunit binds to the initiation site of an mRNA with the help of three initiation factors (IF1-IF3). Here, we describe a novel type of initiation termed "70S-scanning initiation," where the 70S ribosome does not necessarily dissociate after translation of a cistron, but rather scans to the initiation site of the downstream cistron. We detailed the mechanism of 70S-scanning initiation by designing unique monocistronic and polycistronic mRNAs harboring translation reporters, and by reconstituting systems to characterize each distinct mode of initiation. Results show that 70S scanning is triggered by fMet-tRNA and does not require energy; the Shine-Dalgarno sequence is an essential recognition element of the initiation site. IF1 and IF3 requirements for the various initiation modes were assessed by the formation of productive initiation complexes leading to synthesis of active proteins. IF3 is essential and IF1 is highly stimulating for the 70S-scanning mode. The task of IF1 appears to be the prevention of untimely interference by ternary aminoacyl (aa)-tRNA•elongation factor thermo unstable (EF-Tu)•GTP complexes. Evidence indicates that at least 50% of bacterial initiation events use the 70S-scanning mode, underscoring the relative importance of this translation initiation mechanism.protein synthesis | ribosomal functions | translational initiation | 30S-binding initiation | 70S-scanning initiation I t is textbook knowledge that 30S subunits initiate protein synthesis in bacteria; they recognize the initiation site of the mRNA composed of the Shine-Dalgarno (SD) sequence, the AUG codon, and fMet-tRNA, together with three initiation factors (IFs) forming the 30S initiation complex (30SIC). Association of the large 50S subunit triggers the release of the IFs, leading to the 70S initiation complex (70SIC) that enters the elongation phase of translation (reviewed in 1). We term this initiation path the "30S-binding mode" of bacterial initiation. After elongation and termination, it is thought that the ribosome dissociates into its subunits, thus providing 30S subunits for the next round of initiation.The functional role of IF2 is well defined. It can bind directly to the 30S, providing a docking site for fMet-tRNA (2), but it can also enter the 30S subunit as ternary complex fMet-tRNA•IF2•GTP (3). Both IF2 and IF3 are essential for viability. IF3 has a binding site at the 30S interface (4), which explains its antiassociation effect (5, 6), as well as its role in dissociation of the terminating 70S ribosome (7). However, the in vivo concentration of IF3 is 100-fold less (8) than required for full dissociation of 70S in vitro (4). Evidence for the presence of IF3 on 70S ribosomes was reported (9), indicating that the functional spectrum of IF3 is possibly not restricted to an antiassociation effect. Both IF3 and IF2 are also responsible for the fidelity of decoding the initiation AUG by fMet-tRNA Met f at the P site of 30S subun...

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Section: Discussionmentioning

confidence: 99%

70S-scanning initiation is a novel and frequent initiation mode of ribosomal translation in bacteria

Yamamoto

Wittek

Gupta

et al. 2016

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

106

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“…26,95 To get further insight into the involvement of ABCE1 in cell proliferation, we considered its potential role in the synthesis of proteins essential for cell cycle progression. The translation of certain transcripts is highly dependent on the initiation step, which is determined by both the activity of initiation factors and the 5 0 -UTR structure of mRNA.…”

Section: Discussionmentioning

confidence: 99%

ABCE1 is essential for S phase progression in human cells

et al. 2016

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ABCE1 is a highly conserved protein universally present in eukaryotes and archaea, which is crucial for the viability of different organisms. First identified as RNase L inhibitor, ABCE1 is currently recognized as an essential translation factor involved in several stages of eukaryotic translation and ribosome biogenesis. The nature of vital functions of ABCE1, however, remains unexplained. Here, we study the role of ABCE1 in human cell proliferation and its possible connection to translation. We show that ABCE1 depletion by siRNA results in a decreased rate of cell growth due to accumulation of cells in S phase, which is accompanied by inefficient DNA synthesis and reduced histone mRNA and protein levels. We infer that in addition to the role in general translation, ABCE1 is involved in histone biosynthesis and DNA replication and therefore is essential for normal S phase progression. In addition, we analyze whether ABCE1 is implicated in transcript-specific translation via its association with the eIF3 complex subunits known to control the synthesis of cell proliferation-related proteins. The expression levels of a few such targets regulated by eIF3A, however, were not consistently affected by ABCE1 depletion.

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“…Reinitiation of translation refers to the mechanism that allows the ribosome to bind and initiate at a downstream start codon after a stop codon has been met upstream [32][33][34][35] and reviewed in Jackson et al 36 A substantial number (445%) of mammalian genes contain at least one short upstream ORF (upORF) preceding the coding domain of the gene. 34,36 The process of reinitiation is often inefficient.…”

Section: Discussionmentioning

confidence: 99%

Reinitiation of mRNA translation in a patient with X-linked infantile spasms with a protein-truncating variant in ARX

et al. 2015

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Mutations in the Aristaless-related homeobox gene (ARX) lead to a range of X-linked intellectual disability phenotypes, with truncating variants generally resulting in severe X-linked lissencephaly with ambiguous genitalia (XLAG), and polyalanine expansions and missense variants resulting in infantile spasms. We report two male patients with early-onset infantile spasms in whom a novel c.34G4T (p.(E12*)) variant was identified in the ARX gene. A similar variant c.81C4G (p.(Y27*)), has previously been described in two affected cousins with early-onset infantile spasms, leading to reinitiation of ARX mRNA translation resulting in an N-terminal truncated protein. We show that the novel c.34G4T (p.(E12*)) variant also reinitiated mRNA translation at the next AUG codon (c.121-123 (p.M41)), producing the same N-terminally truncated protein. The production of both of these truncated proteins was demonstrated to be at markedly reduced levels using in vitro cell assays. Using luciferase reporter assays, we demonstrate that transcriptional repression capacity of ARX was diminished by both the loss of the N-terminal corepressor octapeptide domain, as a consequence of truncation, and the marked reduction in mutant protein expression. Our study indicates that premature termination mutations very early in ARX lead to reinitiation of translation to produce N-terminally truncated protein at markedly reduced levels of expression. We conclude that even low levels of N-terminally truncated ARX is sufficient to improve the patient's phenotype compared with the severe phenotype of XLAG that includes malformations of the brain and genitalia normally seen in complete loss-of-function mutations in ARX.

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Reinitiation and Other Unconventional Posttermination Events during Eukaryotic Translation

Cited by 144 publications

References 52 publications

70S-scanning initiation is a novel and frequent initiation mode of ribosomal translation in bacteria

70S-scanning initiation is a novel and frequent initiation mode of ribosomal translation in bacteria

ABCE1 is essential for S phase progression in human cells

Reinitiation of mRNA translation in a patient with X-linked infantile spasms with a protein-truncating variant in ARX

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