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bioRxiv · 10.1101/2021.03.18.436003

Elimination of GPI2 suppresses glycosylphosphatidylinositol GlcNAc transferase activity and alters GPI glycan modification in Trypanosoma brucei

Jenni, A.·Knüsel, S.·Nagar, R.·Benninger, M.·Häner, R.·Ferguson, M. A. J.·Roditi, I.·Menon, A. K.·Bütikofer, P.
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Abstract

The biosynthesis of glycosylphosphatidylinositol (GPI) membrane protein anchors is initiated in the endoplasmic reticulum by transfer of GlcNAc from the sugar nucleotide UDP-GlcNAc to phosphatidylinositol. The reaction is catalyzed by GPI GlcNAc transferase, a multi-subunit complex comprising the catalytic subunit Gpi3/PIG-A, as well as at least five other subunits including the hydrophobic protein Gpi2 which is essential for activity in yeast and mammals, but whose function is not known. Here we exploited Trypanosoma brucei (Tb), an early diverging eukaryote and important model organism, to investigate the function of Gpi2. We generated trypanosomes that lack TbGPI2 and found that in TbGPI2-null parasites (i) GPI GlcNAc transferase activity is reduced but not lost, in contrast with the situation in yeast and human cells, (ii) the GPI GlcNAc transferase complex persists, but its architecture is affected, with loss of at least the TbGPI1 subunit, and (iii) the GPI anchors of the major surface proteins are underglycosylated when compared with their wild-type counterparts, indicating the importance of TbGPI2 for reactions that are expected to occur in the Golgi apparatus. Additionally, TbGPI2-null parasites were unable to perform social motility, a form of collective migration on agarose plates. Immunofluorescence microscopy localized TbGPI2 to the endoplasmic reticulum as expected, but also to the Golgi apparatus, suggesting that in addition to its expected function as a subunit of the GPI GlcNAc transferase complex, TbGPI2 may have an enigmatic non-canonical role in Golgi-localized GPI anchor modification in trypanosomes.

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Jenni, A., Knüsel, S., Nagar, R., Benninger, M., Häner, R., Ferguson, M. A. J., Roditi, I., Menon, A. K., Bütikofer, P.. 2021-03-19. Elimination of GPI2 suppresses glycosylphosphatidylinositol GlcNAc transferase activity and alters GPI glycan modification in Trypanosoma brucei. https://doi.org/10.1101/2021.03.18.436003

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