In vivo analysis of protein translation activity in sea urchin eggs and embryos

Héloïse Chassé (1, 2, 3) , Sandrine Boulben (1, 2, 3) , Virginie Glippa (1, 2, 3) , Florian Pontheaux (3, 4, 2) , Patrick Cormier (3, 4, 2) , Julia Morales (2, 3, 4)

1 LBI2M - Laboratoire de Biologie Intégrative des Modèles Marins
2 CNRS - Centre National de la Recherche Scientifique
3 SU - Sorbonne Université
4 LBI2M - Laboratoire de Biologie Intégrative des Modèles Marins

Protein synthesis is a major regulatory step of gene expression in different physiological processes including development. Translation of proteins in sea urchin is stimulated upon fertilization, and is necessary for cell cycle progression and development. Translational control is exerted through multifactorial mechanisms, including mRNA recruitment into polysomes and increased rates of translational activity. In this chapter, we review the methods used in sea urchin eggs and embryos to analyze translation activity in vivo both from perspectives of the proteins and of the mRNAs. First, we describe methods to quantify or visualize newly synthesized proteins with radioactive and non-radioactive labeling techniques. Next we present the polysome isolation and profiling on sucrose gradients, allowing the identification of translated mRNAs. Finally, we outline a procedure to follow the translation of a reporter luciferase protein from an mRNA microinjected into the egg.

Mots clés: protein synthesis; translation; polysomes; polysome gradient; mRNA; luciferase reporter; microinjection

Domaines: Sciences du Vivant [q-bio]

Héloïse Chassé, Sandrine Boulben, Virginie Glippa, Florian Pontheaux, Patrick Cormier, et al.. In vivo analysis of protein translation activity in sea urchin eggs and embryos. Echinoderms, Part B, 151, 2019, Methods in Cell Biology, ⟨10.1016/bs.mcb.2018.10.008⟩. ⟨hal-02018207⟩