TY - JOUR
T1 - Long-distance axonal transport of AAV9 is driven by dynein and kinesin-2 and is trafficked in a highly motile Rab7-positive compartment
AU - Castle, Michael J.
AU - Perlson, Eran
AU - Holzbaur, Erika L.F.
AU - Wolfe, John H.
N1 - Funding Information:
We thank Margaret Maronski and Marc Dichter (University of Pennsylvania) for providing dissociated rat cortical neurons, Rui Xiao (University of Pennsylvania) for assistance with statistical analysis, Luk Vandenberghe (Harvard University) for AAV-related technical advice and discussion, and Jessica Melin (Stanford Microfluidics Foundry) for microfluidic mold fabrication and design assistance. All research was conducted in Philadelphia, PA, USA. This work was supported by NINDS grants T32NS007413, R01NS038690, and R01NS060698.
PY - 2014/3
Y1 - 2014/3
N2 - Adeno-associated virus (AAV) vectors can move along axonal pathways after brain injection, resulting in transduction of distal brain regions. This can enhance the spread of therapeutic gene transfer and improve treatment of neurogenetic disorders that require global correction. To better understand the underlying cellular mechanisms that drive AAV trafficking in neurons, we investigated the axonal transport of dye-conjugated AAV9, utilizing microfluidic primary neuron cultures that isolate cell bodies from axon termini and permit independent analysis of retrograde and anterograde axonal transport. After entry, AAV was trafficked into nonmotile early and recycling endosomes, exocytic vesicles, and a retrograde-directed late endosome/lysosome compartment. Rab7-positive late endosomes/lysosomes that contained AAV were highly motile, exhibiting faster retrograde velocities and less pausing than Rab7-positive endosomes without virus. Inhibitor experiments indicated that the retrograde transport of AAV within these endosomes is driven by cytoplasmic dynein and requires Rab7 function, whereas anterograde transport of AAV is driven by kinesin-2 and exhibits unusually rapid velocities. Furthermore, increasing AAV9 uptake by neuraminidase treatment significantly enhanced virus transport in both directions. These findings provide novel insights into AAV trafficking within neurons, which should enhance progress toward the utilization of AAV for improved distribution of transgene delivery within the brain.
AB - Adeno-associated virus (AAV) vectors can move along axonal pathways after brain injection, resulting in transduction of distal brain regions. This can enhance the spread of therapeutic gene transfer and improve treatment of neurogenetic disorders that require global correction. To better understand the underlying cellular mechanisms that drive AAV trafficking in neurons, we investigated the axonal transport of dye-conjugated AAV9, utilizing microfluidic primary neuron cultures that isolate cell bodies from axon termini and permit independent analysis of retrograde and anterograde axonal transport. After entry, AAV was trafficked into nonmotile early and recycling endosomes, exocytic vesicles, and a retrograde-directed late endosome/lysosome compartment. Rab7-positive late endosomes/lysosomes that contained AAV were highly motile, exhibiting faster retrograde velocities and less pausing than Rab7-positive endosomes without virus. Inhibitor experiments indicated that the retrograde transport of AAV within these endosomes is driven by cytoplasmic dynein and requires Rab7 function, whereas anterograde transport of AAV is driven by kinesin-2 and exhibits unusually rapid velocities. Furthermore, increasing AAV9 uptake by neuraminidase treatment significantly enhanced virus transport in both directions. These findings provide novel insights into AAV trafficking within neurons, which should enhance progress toward the utilization of AAV for improved distribution of transgene delivery within the brain.
UR - http://www.scopus.com/inward/record.url?scp=84895461561&partnerID=8YFLogxK
U2 - 10.1038/mt.2013.237
DO - 10.1038/mt.2013.237
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AN - SCOPUS:84895461561
SN - 1525-0016
VL - 22
SP - 554
EP - 566
JO - Molecular Therapy
JF - Molecular Therapy
IS - 3
ER -