Internalization of a GFP-tetanus toxin C-terminal fragment fusion protein at mature mouse neuromuscular junctions.
The distribution, dynamics, internalization, and retrograde axonal traffic of a fusion protein composed of green fluorescent protein (GFP)and the atoxic C-terminal fragment of tetanus toxin (TTC) were studied after its in vivo injection. Confocal microscopy and immuno-gold electron microscopy revealed that the fusion protein (GFP-TTC) rapidly clustered in motor nerve terminals of the neuromuscular junction. Clathrin-coated pits, and axolemma infoldings located between active zones appeared to be involved in the internalization of the fusion protein. Biochemical analysis of detergent-extracted neuromuscular preparations showed that the GFP-TTC fusion protein was associated with lipid microdomains. We suggest that GFP-TTC clustering in these lipid microdomains favors the recruitment of other proteins involved in its endocytosis and internalization in motor nerve terminals. During its retrograde trafficking, GFP-TTC accumulated indifferent axonal compartments than those used by cholera toxin B-subunit suggesting that these two proteins are transported by different pathways and cargos.
Roux
Sylvie
S
Unité d'Embryologie Moléculaire, Institut Pasteur, Unités de Recherche Associées 2578, Centre National de la Recherche Scientifique, Paris, France. sroux@nbcm.cnrs-gif.fr
Colasante
Cesare
C
Saint Cloment
Cécile
C
Barbier
Julien
J
Curie
Thomas
T
Girard
Emmanuelle
E
Molgó
Jordi
J
Brûlet
Philippe
P
D000818:Animals; D001370:Axonal Transport / Q000502:physiology*; D002772:Cholera Toxin / Q000378:metabolism; D022163:Clathrin-Coated Vesicles / Q000378:metabolism / Q000648:ultrastructure; D004705:Endocytosis / Q000502:physiology*; D005260:Female; D049452:Green Fluorescent Proteins / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism / Q000648:ultrastructure; D051379:Mice; D009046:Motor Neurons / Q000378:metabolism / Q000648:ultrastructure; D009469:Neuromuscular Junction / Q000378:metabolism* / Q000648:ultrastructure; D010446:Peptide Fragments / Q000378:metabolism; D017729:Presynaptic Terminals / Q000378:metabolism / Q000648:ultrastructure; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013570:Synaptic Membranes / Q000378:metabolism* / Q000648:ultrastructure; D013744:Tetanus Toxin / Q000378:metabolism; D014056:Tomography, X-Ray
D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D013744:Tetanus Toxin; C040635:tetanus toxin fragment C; D049452:Green Fluorescent Proteins; D002772:Cholera Toxin