In vivo neuronal tracing with GFP-TTC gene delivery.
The retrograde transport and transynaptic transfer properties of the nontoxic tetanus toxin C-fragment (TTC) can be used to visualize specific neural pathways or to deliver biomolecules in the central nervous system (CNS). Here we tested different delivery techniques to explore the potential use of a new GFP-TTC fusion construct for use as a genetic tracer in vivo. Plasmids encoding GFP-TTC were targeted to brain regions using intracerebral grafted transfected cells or adenoviral transduction. Transport was monitored using GFP fluorescence. We show that following GFP-TTC synthesis in grafted transfected cells, the TTC fragment alone, with no signal peptide, is necessary and sufficient to provide secretion and uptake of the fusion protein into neighboring neurons around the injection site. Using an adenoviral vector to express the fusion protein into brain neurons, we show that transduced neurons can deliver the fusion protein specifically into connected neurons, demonstrating that synaptic transfer in the CNS can be visualized with GFP-TTC.
Kissa
Karima
K
Unité d'Embryologie Moléculaire, URA 1947 CNRS, Institut Pasteur, 25 rue du Dr Roux, F-75724 Paris Cedex 15, France.
Mordelet
Elodie
E
Soudais
Claire
C
Kremer
Eric J
EJ
Demeneix
Barbara A
BA
Brûlet
Philippe
P
Coen
Laurent
L
D000818:Animals; D001693:Biological Transport, Active; D001921:Brain / Q000601:surgery; D002460:Cell Line; D017690:Cell Transplantation; D002490:Central Nervous System / Q000502:physiology*; D018920:Coculture Techniques; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000493:pharmacokinetics; D007425:Intracellular Membranes / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics* / Q000493:pharmacokinetics; D008297:Male; D009474:Neurons / Q000502:physiology*; D010446:Peptide Fragments / Q000235:genetics* / Q000493:pharmacokinetics; D021382:Protein Sorting Signals / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D013744:Tetanus Toxin / Q000235:genetics* / Q000493:pharmacokinetics; D014018:Tissue Distribution; D014161:Transduction, Genetic
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D010446:Peptide Fragments; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D013744:Tetanus Toxin; C040635:tetanus toxin fragment C; D049452:Green Fluorescent Proteins