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mRNA translation under hypoxia relies on specific 3’UTR sequences and cap-binding factor eIF4EHP in Drosophila melanogaster

par Hody, Clara
Président du jury Droogmans, Louis
Promoteur Gueydan, Cyril
Co-Promoteur Defrance, Matthieu
Publication Non publié, 2024-03-15

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Résumé :

The molecular mechanisms governing gene expression and corresponding protein synthesis in a global context of stress-induced translation inhibition remain an open question in metazoans. Using reporter gene approach and gene inactivation strategies, we identified a mechanism, responsible for LDH production under hypoxia and depending on the recognition of a specific CA-rich region in Ldh 3’UTR and an alternative cap recognition factor eIF4EHP. To better characterize the overall role of eIF4EHP in hypoxic translation, high-throughput data were generated in hypoxic S2 cells (RNA seq and polysome profiling) to identify the messengers comprising the hypoxic transcriptome and translatome. Analysis of the variations in total mRNA and polysomal mRNA between normoxic to hypoxic transitions revealed that hypoxia deeply impacts both the transcriptome and translatome of S2 cells, in similar proportions. Furthermore, analysis of the variations in mRNA translation efficiency revealed the existence of specific translation mechanisms promoting mRNA recruitment on the polysomes, independently of their total abundance, under hypoxic conditions. The messengers with an increased total or polysomal abundance reflect functions induced by hypoxia, such as glycolysis, correct protein folding mediated by chaperone proteins or protection against oxidative stress. On the other hand, messengers encoding functions linked to ribogenesis or translation are repressed in hypoxia. Comparative Clip-seq analysis of hypoxic wild-type and eIF4EHP-KO S2 cells identified specifically bound by eIF4EHP. eIF4EHP-bound mRNAs have higher translational efficiency, suggesting a global role of eIF4EHP as a translation activator. eIF4EHP upregulates translation of messengers involved in signaling pathways, such as MAPK or TGFβ pathways, rather than in metabolic adaptations. Similarly to our observation related to Ldh mRNA, CA-rich motifs were identified in the 3'UTR of a large family of mRNAs with improved translation efficiency and bound by eIF4EHP target mRNAs, reinforcing the hypothesis of a global regulatory mechanism of translation in hypoxia by eIF4EHP. AU-rich motifs were also identified in the 3'UTRs of eIF4EHP bound messengers and therefore appear to promote recognition by eIF4EHP. Those motifs are also enriched in mRNA targets with a decreased total abundance and could be associated with a degradation mechanism under hypoxia. We also observed that eIF4EHP binds its own messenger suggesting the existence of an autoregulatory loop controlling eIF4EHP synthesis. eIF4EHP is expressed in two distinct isoforms in S2 cells showing different expression profiles in hypoxia. Based on structural differences and preliminary analysis of proteins partners, the isoform mRNAs have diverging functions. Taken together, we have identified a functional group of mRNAs escaping the translational blockade induced by hypoxic stress in Drosophila S2 cells. We have identified cis- and trans-acting factors that allows this translation on specific and global scales. Given the conservation between D. melanogaster and H. sapiens genomes, the global mechanism of translation regulation identified in this work could have been maintained across evolution.