An approach to analyse the specific impact of rapamycin on mRNA-ribosome association

Article Article scientifique Article An approach to analyse the specific impact of rapamycin on mRNA-ribosome association eng 2008 author Genolet Raphaël 631340 author Araud Tanguy 629127 author Maillard Laetitia 636230 author Jaquier-Gubler Pascale 23175 author Curran Joseph 22465 10.1186/1755-8794-1-33 18673536 1755-8794 http://www.biomedcentral.com/1755-8794/1/33 BACKGROUND: Recent work, using both cell culture model systems and tumour derived cell lines, suggests that the differential recruitment into polysomes of mRNA populations may be sufficient to initiate and maintain tumour formation. Consequently, a major effort is underway to use high density microarray profiles to establish molecular fingerprints for cells exposed to defined drug regimes. The aim of these pharmacogenomic approaches is to provide new information on how drugs can impact on the translational read-out within a defined cellular background. METHODS: We describe an approach that permits the analysis of de-novo mRNA-ribosome association in-vivo during short drug exposures. It combines hypertonic shock, polysome fractionation and high-throughput analysis to provide a molecular phenotype of translationally responsive transcripts. Compared to previous translational profiling studies, the procedure offers increased specificity due to the elimination of the drugs secondary effects (e.g. on the transcriptional read-out). For this pilot "proof-of-principle" assay we selected the drug rapamycin because of its extensively studied impact on translation initiation. RESULTS: High throughput analysis on both the light and heavy polysomal fractions has identified mRNAs whose re-recruitment onto free ribosomes responded to short exposure to the drug rapamycin. The results of the microarray have been confirmed using real-time RT-PCR. The selective down-regulation of TOP transcripts is also consistent with previous translational profiling studies using this drug. CONCLUSION: The technical advance outlined in this manuscript offers the possibility of new insights into mRNA features that impact on translation initiation and provides a molecular fingerprint for transcript-ribosome association in any cell type and in the presence of a range of drugs of interest. Such molecular phenotypes defined pre-clinically may ultimately impact on the evaluation of a particular drug in a living cell. Dewey 616 54 Département de microbiologie et médecine moléculaire 21B false 58 Caractérisation génétique et biochimique du complexe Ccr4-Not conservé. 81 365 Régulation traductionnelle dans les cellules de mammifère 631 BMC medical genomics 1 33 18e8df1e-a208-4e88-bc0a-4a9b6a1fcb3d Article (Accepted version) Public 18673536.pdf 928941 application/pdf 55b1d4568be815248aef24bd889437c38205f304f5b6cb6c82d78784f2b5f548 openAccess Faculté de médecine / Section de médecine fondamentale / Département de microbiologie et médecine moléculaire

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An approach to analyse the specific impact of rapamycin on mRNA-ribosome association	BMC medical genomics			2008	674	527

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Maillard, Laetitia