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Contribution to the understanding of translation under hypoxia in Drosophila melanogaster

par Fosseprez, Gauthier
Président du jury Andris, Fabienne
Promoteur Gueydan, Cyril
Co-Promoteur Kruys, Véronique
Publication Non publié, 2023-09-06

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Résumé :

During their growth, cells must face different kind of stress such as nutrient starvation, DNA damage or even hypoxia. Global translation is usually inhibited under these conditions. However, translation of specific mRNAs is undertaken to assure cellular response. As the canonical pathway is inhibited, specific translation is guaranteed through alternative mechanisms. Such mechanisms rely, among others, on the intervention of paralogs of the canonical cap-recognition initiation factor, which is eIF4E1. One of these paralogs is eIF4E2, called eIF4EHP in Drosophila melanogaster, which has been shown to act both as a translational inhibitor and activator. Alternative translational mechanisms can also occur through the presence in a DRACH motif of the N6-methyladenosine (m6A) modification on mRNA. During hypoxic stress, the hypoxia inducible factors (HIFs) are important transcription factors assuring the cells hypoxic response. In D. melanogaster, the Similar (Sima) protein is the unique HIFs- homologue. Under hypoxia, Ldh mRNA level, and subsequent protein level, is increased and our laboratory has shown that its hypoxic translation is eIF4EHP- and CA-rich 3’UTR-dependent.Because D. melanogaster's m6A DRACH motif is similar to Ldh CA-rich translation activating motif and m6A pathway has been shown to impact translation, assessment of its implication in the expression of Ldh under hypoxia was first realised. Results showed that the m6A pathway seems to influence Ldh production both in normoxia and hypoxia, which could occur through Ythdc1-mediated transcription repression, Ythdf-mediated mRNA degradation or Mettl3-mediated translation. We then wanted to assess whether or not hypoxic Sima mRNA translation was similar to the recently described mechanisms of hypoxic Ldh mRNA translation. Data showed that hypoxic translation of Sima mRNA seems to be eIF4EHP-dependent without excluding the implication of other unknown mechanisms. However, although Sima mRNA 3’UTR seems to promote hypoxic translation, results showed that it is eIF4EHP-independent, in opposition with Ldh mRNA. Finally, characterization of eIF4EHP isoforms revealed that they present a differential translation profile and interactome. Indeed, -RA isoform translation is reduced under hypoxia while -RB is maintained. Furthermore, sequence alignment and structural data showed that -RA and -RB isoforms differ in their dorsal region, shown to be the interaction site between human eIF4EHP and GIGYF protein. Yeast two-hybrid experiment revealed that D. melanogaster -RA isoform interacts with GIGYF protein in opposition of -RB. Altogether, these observations suggest that the described dual role of eIF4EHP as a translational repressor and activator could be explained through the observed differences between eIF4EHP-RA and -RB isoforms.