Initiation of translation of the cauliflower mosaic virus genome from a polycistronic mRNA: evidence from deletion mutagenesis

Dixon, Linda K.; Höhn, Thomas

doi:10.1002/j.1460-2075.1984.tb02203.x

Cited by 141 publications

(72 citation statements)

References 30 publications

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“…In this paper, we extend our earlier analysis of the expression of DHFR with SV40-dhfr recombinant viruses by altering the sequence upstream of the dhfr initiator AUG to reveal signals required for efficient internal translation initiation. The re-sults reinforce the suggestion that translation of mammalian mRNAs can be reinitiated at internal AUG codons if translation that had been initiated upstream is terminated in the vicinity of the internal AUG. Our results are also consistent with those recently reported by others (6,10,11,20).…”

supporting

confidence: 94%

“…The mobility shifts observed on SDS-polyacrylamide gels were consistent with the predicted changes in the molecular 100 100 SVGT7dhfr7 SVGTdhfr22x VOL. 6,1986 on May 7, 2018 by guest http://mcb.asm.org/ Downloaded from sizes of these proteins. SVGT7dhfr22X produced both the fusion and normal forms of DHFR at high levels, apparently because termination of VP2/3 translation occurred only 21 nucleotides downstream of the alternative initiation site.…”

Section: Resultsmentioning

confidence: 99%

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Effect of upstream reading frames on translation efficiency in simian virus 40 recombinants.

Peabody¹,

Subramani²,

Berg³

1986

Mol. Cell. Biol.

130

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In a previous report (S. Subramani, R. Mulligan, and P. Berg, Mol. Cell. Biol. 1:854-864, 1981), it was shown that mouse dihydrofolate reductase (DHFR) could be efficiently expressed from simian virus 40 recombinant viruses containing the DHFR cDNA in different locations in the viral late region. This was true even in the case of the SVGT7dhfr26 recombinant, which had the DHFR coding sequence 700 to 800 nucleotides from the 5' end of the mRNA, where it was preceded by the VP2 and VP3 initiator AUGs and a number of other noninitiator AUGs. To investigate the process of internal translation initiation in mammalian cells, we constructed a series of SVGT7dhfr recombinants in which the upstream VP2 and VP3 reading frame was terminated in various positions relative to the DHFR initiation codon. The efficient production of DHFR in infected CV1 cells depended on having the terminators of the VP2-VP3 reading frame positioned upstream or nearby downstream from the DHFR initiation codon. These results reinforce the notion that mammalian ribosomes are capable of translational reinitiation.

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supporting

confidence: 94%

Section: Resultsmentioning

confidence: 99%

Effect of upstream reading frames on translation efficiency in simian virus 40 recombinants.

Peabody¹,

Subramani²,

Berg³

1986

Mol. Cell. Biol.

130

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“…However, the translational start does not contain the consensus purine at the -3 position. Although polycistronic messages are common in procaryotes, in eucaryotes they are known only in viral systems (3,8,15,19 (47). However, blot analysis with Antp probes of RNA from embryonic, larval, and pupal stages demonstrated the activity of both promoters at each stage ( Fig.…”

Section: Discussionmentioning

confidence: 99%

Multiple transcripts from the Antennapedia gene of Drosophila melanogaster.

Stroeher¹,

Jørgensen²,

Garber³

1986

Mol. Cell. Biol.

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“…On the other hand, the EBNA2 protein is expressed inefficiently despite a lengthy intercistronic gap between the leader ORF and the EBNA2 coding sequence (58,69). Reinitiation is believed to be the natural mode of translation of caulimovirus genes (12), which are closely spaced (21); thus the requirements for reinitiation in plants might differ from those in mammals.…”

Section: Discussionmentioning

confidence: 99%

Effects of intercistronic length on the efficiency of reinitiation by eucaryotic ribosomes.

Kozak¹

1987

Mol. Cell. Biol.

502

433

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Simian virus 40-based plasmids that direct the synthesis of preproinsulin during short-term transfection of COS cells have been used to probe the mechanism of reinitiation by eucaryotic ribosomes. Earlier studies from several laboratories had established that the ability of ribosomes to reinitiate translation at an internal AUG codon depends on having a terminator codon in frame with the preceding AUG triplet and upstream from the intended restart site. In the present studies, the position of the upstream terminator codon relative to the preproinsulin restart site has been systematically varied. The efficiency of reinitiation progressively improved as the intercistronic sequence was lengthened. When the upstream "minicistron" terminated 79 nucleotides before the preproinsulin start site, the synthesis of proinsulin was as efficient as if there were no upstream AUG codons. A mechanism is postulated that might account for this result, which is somewhat surprising inasmuch as bacterial ribosomes reinitiate less efficiently as the intercistronic gap is widened.Eucaryotic ribosomes usually initiate at the AUG codon that lies closest to the 5' end of the mRNA (31). That tendency has been rationalized by postulating that the small ribosomal subunit binds initially at the capped 5' end of the mRNA and subsequently migrates to the AUG initiator codon, which is recognized more or less efficiently depending on nearby sequences. From a comparison of several hundred vertebrate mRNAs, GCCACCAUGG has been proposed as the consensus sequence for initiation in higher eucaryotes (30, 33, Kozak, submitted for publication). The contribution of every nucleotide in that motif has been confirmed by mutagenesis (36; Kozak, J. Mol. Biol., in press). The most highly conserved nucleotides are the purine, most often A, in position -3 (i.e., three nucleotides upstream from the AUG codon) and the G in position +4; the aforementioned mutational analyses confirmed that those two positions have the strongest influence on initiation. Thus, a potential initiator codon can usually be designated as "strong" or "weak" by considering only those positions.There are ways for eucaryotic ribosomes to initiate at internal sites in certain mRNAs, but those sites can be reached only by advancing from the 5' end; never, it seems, by direct internal binding. An early indication of the constraint against direct internal binding was the inability of eucaryotic ribosomes to bind to circular RNA templates (27,28), and a considerable body of other evidence has since been adduced (see Discussion). One mechanism by which ribosomes can reach an internal AUG codon is "leaky scanning," which occurs when the 5'-proximal AUG codon lies in an unfavorable context for initiation. Data consistent with leaky scanning have come from manipulating the sequences of synthetic constructs (35,36,40) and from analyzing naturally occurring viral mRNAs that are bifunctional (reviewed in references 38 and 39). A second mechanism for reaching an internal AUG codon operates when the upstre...

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Initiation of translation of the cauliflower mosaic virus genome from a polycistronic mRNA: evidence from deletion mutagenesis

Cited by 141 publications

References 30 publications

Effect of upstream reading frames on translation efficiency in simian virus 40 recombinants.

Effect of upstream reading frames on translation efficiency in simian virus 40 recombinants.

Multiple transcripts from the Antennapedia gene of Drosophila melanogaster.

Effects of intercistronic length on the efficiency of reinitiation by eucaryotic ribosomes.

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