Functional characterization of the alpha glucosidase II (GSII) in the green microalga Chlamydomonas reinhardtii
Résumé
The N-glycosylation pathway of proteins is one of the most important co- and posttranslational modification of proteins in Eukaryotes1. Glycoproteins are produced firstly in the endoplasmic reticulum (ER) where molecular actors (such as glucosidases I and II, lectins) are responsible for the proper folding and 3D conformation of proteins before their transfer into the Golgi apparatus2. In a context of bioproduction of therapeutic recombinant glycoproteins, new cheaper expression systems emerged, such as microalgae3,4. The growing interest of using C. reinhardtii, a green microalga, as an emerging cell factory for the industrial production of recombinant glycoproteins require overall an in-depth understanding and analysis of protein N-glycosylation in this organism. Despite a relatively well-investigated N-glycosylation pathway in C. reinhardtii5,6,7,8,9, many glyco-enzymes involved in the N-glycosylation pathway in the ER are still not characterized. In this work, functional characterization of the putative alpha-glucosidase II (GSII) was conducted using combinatory approaches on C. reinhardtii insertional mutant cells in order to evaluate the impact of the GSII gene deletion compared to the WT cells and to give new insights on the ER N-glycosylation pathway in this microalga. Accumulation of free oligosaccharides (fOSs) and N-glycans such as Glc2Man5GlcNAc2 are observed in the GSII mutant cells. Moreover, an overexpression of both Cal and PDI genes, respectively involved in the quality control of misfolded proteins and UPR pathway, was observed at 6 days of culture growth in addition to an accumulation of reactive oxygen species (ROS), suggesting a physiological stress in the microalgae mutant cells.
Domaines
Sciences du Vivant [q-bio]Origine | Fichiers produits par l'(les) auteur(s) |
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