Specific binding of eukaryotic initiation factor 2 to satellite tobacco necrosis virus RNA at a 5′-terminal sequence comprising the ribosome binding site

Kaempfer, Raymond; Emmelo, John van; Fiers, Walter

doi:10.1073/pnas.78.3.1542

Cited by 53 publications

(23 citation statements)

References 26 publications

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“…Eukaryotic initiation factor eIF4B has been observed to bind to sequences in satellite tobacco necrosis virus RNA containing the AUG initiation codon (2) and was found to bind preferentially to AUG triplets (14). Initiation factor eIF2 can interact with various RNA molecules that contain AUG codons (23,35); however, the exact binding sites for eIF2 on these RNAs have not been reported. More recently, Dasso et al provided evidence that selection of the 5Ј-proximal start site in influenza virus N2 RNA can be influenced by eIF2 levels (8).…”

Section: Discussionmentioning

confidence: 99%

Evidence for Involvement of trans-Acting Factors in Selection of the AUG Start Codon during Eukaryotic Translational Initiation

McBratney

Sarnow

1996

Molecular and Cellular Biology

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The molecular mechanism with which an appropriate AUG codon is selected as the start site for translational initiation by eukaryotic ribosomes is not known. By using a cell-free translation system, small RNA molecules containing single AUG codons, surrounded by various nucleotide sequences, were tested for their abilities to interfere with the translation of a reporter mRNA. RNAs containing the AUG in an ACCAUGG context (Kozak consensus sequence) were able to inhibit translation of the reporter mRNA. In contrast, RNAs containing the AUG in a less favorable context for start site selection (for example, CAGAUGC) had no effect on the translation of the reporter mRNA. The effect mediated by the ACCAUGG-containing RNAs was not due to sequestration of ribosomal subunits or to particular structural features in these RNAs. To identify potential trans-acting factors that might be preferentially bound by ACCAUGG-containing RNAs, ACCAUGG-and CAGAUGC-containing RNAs with a single 4-thiouridine residue at the AUG were incubated with partially fractionated extracts, and AUG-binding proteins were identified after irradiation of the complexes with UV light and subsequent analysis by gel electrophoresis. The analysis of such complexes in competition experiments revealed that proteins, approximately 50 and 100 kDa in size, were found to bind directly at the AUG codon embedded in the ACCAUGG motif. One of these proteins has been identified as the La autoantigen. These findings indicate that trans-acting factors may play a role in AUG start site selection during translational initiation.

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Section: Discussionmentioning

confidence: 99%

Evidence for Involvement of trans-Acting Factors in Selection of the AUG Start Codon during Eukaryotic Translational Initiation

McBratney

Sarnow

1996

Molecular and Cellular Biology

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“…First, capping of full-length STNV RNA does not enhance initiation complex formation whereas capping of 5'-terminal fragments has a stimulatory effect (26). Second, the preferential binding of eIF-2 to full-length and nearly full-length STNV (14) suggests that eIF-2 recognizes a conformation present only in larger molecules. A secondary structure model for STNV-1 RNA in which nt 24 to 55 are interacting with nt 979 to 1009 was proposed (14).…”

mentioning

confidence: 99%

The 3' untranslated region of satellite tobacco necrosis virus RNA stimulates translation in vitro.

Danthinne¹,

Seurinck²,

Meulewaeter³

et al. 1993

Mol. Cell. Biol.

102

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The RNA of satellite tobacco necrosis virus (STNV) is a monocistronic messenger that lacks both a 5' cap structure and a 3' poly(A) tail. We show that in a cell-free translation system derived from wheat germ, STNV RNA lacking the 600-nucleotide trailer is translated an order of magnitude less efficiently than full-size RNA.Deletion analyses positioned the translational enhancer domain (TED) within a conserved hairpin structure immediately downstream from the coat protein cistron. TED enhances translation when fused to a heterologous mRNA, but the level of enhancement depends on the nature of the 5' untranslated sequence and is maximal in combination with the STNV leader. The STNV leader and TED have two regions of complementarity. One of the complementary regions in TED resembles picornavirus box A, which is involved in cap-independent translation but which is located upstream of the coding region. In contrast, the STNV-1 and -2 trailers represent half of the genome and share 64% nucleotide sequence conservation (4). STNV RNA is characterized by the absence of both a cap structure at the 5' end and a poly(A) tail at the 3' end (11). This peculiarity has two implications with regard to translation.First, STNV RNA apparently uses a translational initiation mechanism that bypasses the common cap recognition step. This mechanism may be quite different from that used by other uncapped viral RNAs like those of picornaviruses, comoviruses, and potyviruses (3, 27, 28). Picornaviral RNAs have long 5' untranslated sequences ranging from 600 to 1,200 nt, which carry multiple noninitiating AUG codons and which can fold into highly conserved secondary structures involved in translation initiation (reference 27 and references therein). In contrast, the STNV leader sequence is very short, and its predicted hairpin structure is not as extensive and stable (4). Interestingly, translation of STNV RNA in vitro, as of poliovirus, is stimulated by the cap binding factor eukaryotic initiation factor 4F (eIF-4F) (1, 6).Second, the absence of a poly(A) tail poses the question of how STNV RNA substitutes for the major cytoplasmic functions associated with this structure, i.e., the control of mRNA stability and the modulation of translation efficiency. The poly(A) tail, complexed with poly(A)-binding protein, is believed to enhance the formation of 80S translational initi-* Corresponding author. ation complexes, presumably through promoting some sort of interaction between 5'-and 3'-proximal elements of the mRNA (20). The exact role of the STNV trailer in viral replication and translation is not known, but its regulatory function is suggested by its sequence conservation, which exceeds that of the coat protein coding region. Phylogenetic comparison between the STNV-1 and STNV-2 trailer sequences revealed the presence of three pseudoknotted structures which are also found in several other nonpolyadenylated plant viral RNAs such as that of tobacco mosaic virus (TMV) (4, 23). In the latter case, it was shown that the pseudoknot domain can ...

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“…7). It has also been reported that eIF2 interacts directly with the 5Ј end of STNV RNA, and this may also play a role in the recruitment of a 40 S ribosome to the 5Ј end (69). However, eIF2 did not bind specifically to the 5Ј-UTR of STNV-1 in the nitrocellulose binding assay (data not shown).…”

Section: Stnv-1 Ted Functions As a Cap Mimic In The 3ј-utr-mentioning

confidence: 76%

A Novel Interaction of Cap-binding Protein Complexes Eukaryotic Initiation Factor (eIF) 4F and eIF(iso)4F with a Region in the 3′-Untranslated Region of Satellite Tobacco Necrosis Virus

Gazo

Murphy

Gatchel

et al. 2004

Journal of Biological Chemistry

108

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Satellite tobacco necrosis virus (STNV)RNA is naturally uncapped at its 5 end and lacks polyadenylation at its 3 end. Despite lacking these two hallmarks of eukaryotic mRNAs, STNV-1 RNA is translated very efficiently. A ϳ130-nucleotide translational enhancer (TED), located 3 to the termination codon, is necessary for efficient cap-independent translation of STNV-1 RNA. The STNV-1 TED RNA fragment binds to the eukaryotic cap-binding complexes, initiation factor (eIF) 4F and eIF(iso)4F, as measured by nitrocellulose binding and fluorescence titration. STNV-1 TED is a potent inhibitor of in vitro translation when added in trans. This inhibition is reversed by the addition of eIF4F or eIF(iso)4F, and the subunits of eIF4F and eIF(iso)4F cross-link to STNV-1 TED, providing additional evidence that these factors interact directly with STNV-1 TED. Deletion mutagenesis of the STNV-1 TED indicates that a minimal region of ϳ100 nucleotides is necessary to promote cap-independent translation primarily through interaction with the cap binding subunits (eIF4E or eIF(iso)4E) of eIF4F or eIF(iso)4F.Initiation of protein synthesis in eukaryotic cells is a complicated process requiring 8 -10 initiation factors for proper alignment of the initiation codon of messenger RNA on the 40 S ribosome and subsequent joining of the 60 S ribosome (for review, see Refs. 1-6). For eukaryotes the first step in this process is the recognition of the m 7 GpppX cap structure at the 5Ј end of mRNA by eIF4E, 1 the cap-binding protein component of the eIF4F complex. The eIF4G component of eIF4F then acts as a scaffold for the assembly of other initiation factors (for review, see Refs. 7-11). Initiation factors eIF4A, eIF3, and poly(A)-binding protein (PABP) are known to interact with eIF4G during this process (for review, see Refs. 7, 9, and 10). PABP binds the poly(A) tail present at the 3Ј end of most cellular mRNAs, and the interaction of PABP with eIF4G implies that the 5Ј and 3Ј ends of the cellular mRNA are brought into close proximity during the initiation process (for review, see Refs. 9 and 12). This assembly of factors on the mRNA presumably functions in an ATP-dependent unwinding of secondary structure in the 5Ј-UTR of the mRNA before binding and scanning of the 40 S ribosome to find the correct initiation codon (5).Plant eIF4F consists of two subunits, a small cap-binding protein, eIF4E (26 kDa), and a large subunit, eIF4G (180 kDa). Higher plants possess an isozyme form of eIF4F, termed eIF(iso) 4F (13, 14). eIF(iso)4F, like eIF4F, consists of two subunits, eIF(iso)4E (28 kDa), and a large subunit, eIF(iso)4G (86 kDa). eIF(iso)4F has functions similar to eIF4F in the initiation of translation of plant mRNAs (14). The amino acid sequence of plant eIF(iso)4E is ϳ50% similar to plant eIF4E and retains all the conserved tryptophan residues found in cap-binding proteins (15,16). The large subunit, eIF(iso)4G, is significantly smaller than plant eIF4G (86 versus 180 kDa, (15)), sharing ϳ30 -40% similarity with central and C-terminal regions o...

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Specific binding of eukaryotic initiation factor 2 to satellite tobacco necrosis virus RNA at a 5′-terminal sequence comprising the ribosome binding site

Cited by 53 publications

References 26 publications

Evidence for Involvement of trans-Acting Factors in Selection of the AUG Start Codon during Eukaryotic Translational Initiation

Evidence for Involvement of trans-Acting Factors in Selection of the AUG Start Codon during Eukaryotic Translational Initiation

The 3' untranslated region of satellite tobacco necrosis virus RNA stimulates translation in vitro.

A Novel Interaction of Cap-binding Protein Complexes Eukaryotic Initiation Factor (eIF) 4F and eIF(iso)4F with a Region in the 3′-Untranslated Region of Satellite Tobacco Necrosis Virus

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