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One at a time, live tracking of NGF axonal transport using quantum dots

Cui, Bianxiao and Wu, Chengbiao and Chen, Liang and Ramirez, Alfredo and Bearer, Elaine L. and Li, Wei-Ping and Mobley, William C. and Chu, Steven (2007) One at a time, live tracking of NGF axonal transport using quantum dots. Proceedings of the National Academy of Sciences of the United States of America, 104 (34). pp. 13666-13671. ISSN 0027-8424. PMCID PMC1959439. https://resolver.caltech.edu/CaltechAUTHORS:CUIpnas07

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[img] PDF (Fig. 6. Gel electrophoresis of BtNGF and QD-NGF.) - Supplemental Material
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[img] PDF (Fig. 7. Binding specificity of QD-NGF.) - Supplemental Material
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[img] PDF (Fig. 8. Electron microscopy of QD-NGF within transport vesicles in DRG neurons.) - Supplemental Material
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[img] Video (AVI) (Movie 1 (.avi). Live imaging of QD-NGF (NGF concentration, 1 nM) containing endosomes being retrogradely transported in the axons of DRG neurons.) - Supplemental Material
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[img] Video (AVI) (Movie 2 (.avi). Tracking the movement of a QD-NGF containing endosome.) - Supplemental Material
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[img] Video (AVI) (Movie 3 (.avi). Live imaging of QD-NGF (0.2 nM) containing endosomes being retrogradely transported in the axons of DRG neurons.) - Supplemental Material
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[img] Video (AVI) (Movie 4 (.avi). Live imaging of QD-NGF (2 nM) containing endosomes being retrogradely transported in the axons of DRG neurons.) - Supplemental Material
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[img] Video (AVI) (Movie 5 (.avi). Live imaging of QD-NGF (20 nM) containing endosomes being retrogradely transported in the axons of DRG neurons.) - Supplemental Material
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Abstract

Retrograde axonal transport of nerve growth factor (NGF) signals is critical for the survival, differentiation, and maintenance of peripheral sympathetic and sensory neurons and basal forebrain cholinergic neurons. However, the mechanisms by which the NGF signal is propagated from the axon terminal to the cell body are yet to be fully elucidated. To gain insight into the mechanisms, we used quantum dot-labeled NGF (QD-NGF) to track the movement of NGF in real time in compartmentalized culture of rat dorsal root ganglion (DRG) neurons. Our studies showed that active transport of NGF within the axons was characterized by rapid, unidirectional movements interrupted by frequent pauses. Almost all movements were retrograde, but short-distance anterograde movements were occasionally observed. Surprisingly, quantitative analysis at the single molecule level demonstrated that the majority of NGF-containing endosomes contained only a single NGF dimer. Electron microscopic analysis of axonal vesicles carrying QD-NGF confirmed this finding. The majority of QD-NGF was found to localize in vesicles 50–150 nm in diameter with a single lumen and no visible intralumenal membranous components. Our findings point to the possibility that a single NGF dimer is sufficient to sustain signaling during retrograde axonal transport to the cell body.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1959439/PubMed CentralArticle
https://doi.org/10.1073/pnas.0706192104DOIUNSPECIFIED
https://doi.org/10.1073/pnas.0706192104DOIUNSPECIFIED
Additional Information:© 2007 National Academy of Sciences of the USA. Contributed by Steven Chu, July 6, 2007 (received for review March 19, 2007). Published online on August 14, 2007, 10.1073/pnas.0706192104. B.C. thanks Drs. Harold Kim, Janice S. Valletta, Keith Weninger, and Wei-Hau Chang for their generous assistance for her research. E.L.B. thanks Jean Edens at Caltech for her skill with thin-sectioning for electron microscopy. This work was supported by National Science Grants PHY-0420752 and PHY-0647161, National Aeronautics and Space Administration Grant NNC04GB49G, National Institutes of Health (NIH) Grants NS24054, NS38869, AG16999, NS046810, NS05537, and GM47368, The Larry L. Hillblom Foundation, The Deane Johnson Fund, The Adler Foundation, Dart Neurosciences LLP, and The Moore Foundation. B.C. acknowledges support from the Pathway to Independence Career Award from NIH. Author contributions: B.C. and C.W. contributed equally to this work; B.C., C.W., W.C.M., and S.C. designed research; B.C., C.W., L.C., A.R., E.L.B., and W.-P.L. performed research; B.C. contributed new reagents/analytic tools; B.C. analyzed data; and B.C., C.W., and W.C.M. wrote the paper. The authors declare no conflict of interest. This article contains supporting information online at www.pnas.org/cgi/content/full/0706192104/DC1.
Funders:
Funding AgencyGrant Number
NSFPHY-0420752
NSFPHY-0647161
NASANNC04GB49G
NIHNS24054
NIHNS38869
NIHAG16999
NIHNS046810
NIHNS05537
NIHGM47368
Larry L. Hillblom FoundationUNSPECIFIED
Deane Johnson FundUNSPECIFIED
Adler FoundationUNSPECIFIED
Dart Neurosciences LLPUNSPECIFIED
Gordon and Betty Moore FoundationUNSPECIFIED
Subject Keywords:live imaging; nerve growth factor; single molecule imaging; NGF signaling; retrograde transport
Issue or Number:34
PubMed Central ID:PMC1959439
Record Number:CaltechAUTHORS:CUIpnas07
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:CUIpnas07
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9853
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:24 Mar 2008
Last Modified:03 Oct 2019 00:04

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