Expression of a glycosylated GFP as a bivalent reporter in exocytosis

Autor: Nadine Paris, Marianna Faraco, Giuseppe Dalessandro, Weronika Krzeszowiec, Gian Pietro Di Sansebastiano, Jean-Marc Neuhaus, Bruno Saint-Jean
Přispěvatelé: Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Physiologie et biotechnologie des Algues (PBA), Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER), Di.S.Te.B.A., Università del Salento, Laboratory of Cell and Molecular Biology, Université de Neuchâtel (UNINE), Paris, N, SAINT JEAN, B, Faraco, M, Krzeszowiec, W, Dalessandro, Giuseppe, NEUHAUS J., M, DI SANSEBASTIANO, Gian Pietro, Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)
Rok vydání: 2009
Předmět:
0106 biological sciences
Glycosylation
Mannose
Golgi Apparatus
Plant Science
01 natural sciences
Green fluorescent protein
chemistry.chemical_compound
Genes
Reporter
MESH: Microscopy
Confocal
MESH: Tobacco
Plant Proteins
chemistry.chemical_classification
0303 health sciences
MESH: Exocytosis
Microscopy
Confocal
biology
Protoplast
MESH: Plant Proteins
Protoplasts
General Medicine
MESH: Protoplasts
MESH: Glycosylation
Amino acid
secretion
Biochemistry
protoplast
symbols
lipids (amino acids
peptides
and proteins)
Glycan
glycosylation
Protein design
Green Fluorescent Proteins
GFP
Exocytosis
03 medical and health sciences
symbols.namesake
MESH: Golgi Apparatus
MESH: Green Fluorescent Proteins
Tobacco
[SDV.BV]Life Sciences [q-bio]/Vegetal Biology
Secretion
030304 developmental biology
MESH: Genes
Reporter
Golgi apparatus
Sialic acid
carbohydrates (lipids)
chemistry
biology.protein
Agronomy and Crop Science
010606 plant biology & botany
Zdroj: Plant Cell Reports
Plant Cell Reports, Springer Verlag, 2010, 29 (1), pp.79-86. ⟨10.1007/s00299-009-0799-7⟩
ISSN: 1432-203X
0721-7714
DOI: 10.1007/s00299-009-0799-7⟩
Popis: International audience; The complex-type N-linked glycans of plants differ markedly in structure from those of animals. Like those of insects and mollusks they lack terminal sialic acid(s) and may contain an alpha-(1,3)-fucose (Fuc) linked to the proximal GlcNAc residue and/or a beta-(1,2)-xylose (Xyl) residue attached to the proximal mannose (Man) of the glycan core. N-glycosylated GFPs were used in previous studies showing their effective use to report on membrane traffic between the ER and the Golgi apparatus in plant cells. In all these cases glycosylated tags were added at the GFP termini. Because of the position of the tag and depending on the sorting and accumulation site of these modified GFP, there is always a risk of processing and degradation, and this protein design cannot be considered ideal. Here, we describe the development of three different GFPs in which the glycosylation site is internally localized at positions 80, 133, or 172 in the internal sequence. The best glycosylation site was at position 133. This glycosylated GFPgl133 appears to be protected from undesired processing of the glycosylation site and represents a bivalent reporter for biochemical and microscopic studies. After experimental validation, we can conclude that amino acid 133 is an effective glycosylation site and that the GFPgl133 is a powerful tool for in vivo investigations in plant cell biology.
Databáze: OpenAIRE
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