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Open Access 01-12-2009 | Research article

Upstream ORF affects MYCN translation depending on exon 1b alternative splicing

Authors: Roger Besançon, Sandrine Valsesia-Wittmann, Clara Locher, Céline Delloye-Bourgeois, Lydie Furhman, Giovani Tutrone, Christophe Bertrand, Anne-Catherine Jallas, Elisabeth Garin, Alain Puisieux

Published in: BMC Cancer | Issue 1/2009

Abstract

Background

The MYCN gene is transcribed into two major mRNAs: one full-length (MYCN) and one exon 1b-spliced (MYCN ^Δ1b) mRNA. But nothing is known about their respective ability to translate the MYCN protein.

Methods

Plasmids were prepared to enable translation from the upstream (uORF) and major ORF of the two MYCN transcripts. Translation was studied after transfection in neuroblastoma SH-EP cell line. Impact of the upstream AUG on translation was evaluated after directed mutagenesis. Functional study with the two MYCN mRNAs was conducted by a cell viability assay. Existence of a new protein encoded by the MYCN ^Δ1buORF was explored by designing a rabbit polyclonal antibody against a specific epitope of this protein.

Results

Both are translated, but higher levels of protein were seen with MYCN ^Δ1bmRNA. An upstream ORF was shown to have positive cis-regulatory activity on translation from MYCN but not from MYCN ^Δ1bmRNA. In transfected SH-EP neuroblastoma cells, high MYCN dosage obtained with MYCN ^Δ1bmRNA translation induces an antiapoptotic effect after serum deprivation that was not observed with low MYCN expression obtained with MYCN mRNA. Here, we showed that MYCNOT: MYCN Overlap Transcript, a new protein of unknown function is translated from the upstream AUG of MYCN ^Δ1bmRNA.

Conclusions

Existence of upstream ORF in MYCN transcripts leads to a new level of MYCN regulation. The resulting MYCN dosage has a weak but significant anti-apoptotic activity after intrinsic apoptosis induction.

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The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2407/9/445/prepub

Title: Upstream ORF affects MYCN translation depending on exon 1b alternative splicing
Authors: Roger Besançon
Sandrine Valsesia-Wittmann
Clara Locher
Céline Delloye-Bourgeois
Lydie Furhman
Giovani Tutrone
Christophe Bertrand
Anne-Catherine Jallas
Elisabeth Garin
Alain Puisieux
Publication date: 01-12-2009
Publisher: BioMed Central
Published in: BMC Cancer / Issue 1/2009
Electronic ISSN: 1471-2407
DOI: https://doi.org/10.1186/1471-2407-9-445

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Abstract

Background

Methods

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Conclusions

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