Dynein mediates retrograde neurofilament transport within axons and anterograde delivery of NFs from perikarya into axons: Regulation by multiple phosphorylation events

We examined the respective roles of dynein and kinesin in axonal transport of neurofilaments (NFs). Differentiated NB2a/d1 cells were transfected with green fluorescent protein‐NF‐M (GFP‐M) and dynein function was inhibited by co‐transfection with a construct expressing myc‐tagged dynamitin, or by i...

Full description

Saved in:

Bibliographic Details
Published in:	Cell motility and the cytoskeleton Vol. 63; no. 5; pp. 266 - 286
Main Authors:	Motil, Jennifer, Chan, Walter K.-H., Dubey, Maya, Chaudhury, Pulkit, Pimenta, Aurea, Chylinski, Teresa M., Ortiz, Daniela T., Shea, Thomas B.
Format:	Journal Article
Language:	English
Published:	Hoboken Wiley Subscription Services, Inc., A Wiley Company 01-05-2006
Subjects:	Animals axonal transport Axonal Transport - drug effects Axons - chemistry Axons - metabolism Biological Transport - drug effects dynein Dyneins - antagonists & inhibitors Dyneins - genetics Dyneins - physiology Flavonoids - pharmacology Green Fluorescent Proteins - analysis Intermediate Filaments - metabolism kinesin Kinesin - metabolism Kinesin - physiology Mice Microtubules - drug effects Microtubules - metabolism mirotubules Mitogen-Activated Protein Kinase Kinases - antagonists & inhibitors Neurites - chemistry Neurites - metabolism neurofilaments Phosphorylation Protein Kinase Inhibitors - pharmacology Tumor Cells, Cultured vinblastine Vincristine - pharmacology
Online Access:	Get full text
Tags:	Add Tag No Tags, Be the first to tag this record!

Description
Summary:	We examined the respective roles of dynein and kinesin in axonal transport of neurofilaments (NFs). Differentiated NB2a/d1 cells were transfected with green fluorescent protein‐NF‐M (GFP‐M) and dynein function was inhibited by co‐transfection with a construct expressing myc‐tagged dynamitin, or by intracellular delivery of purified dynamitin and two antibodies against dynein's cargo domain. Monitoring of the bulk distribution of GFP signal within axonal neurites, recovery of GFP signal within photobleached regions, and real‐time monitoring of individual NFs/punctate structures each revealed that pertubation of dynein function inhibited retrograde transport and accelerated anterograde, confirming that dynein mediated retrograde axonal transport, while intracellular delivery of two anti‐kinesin antibodies selectively inhibited NF anterograde transport. In addition, dynamitin overexpression inhibited the initial translocation of newly‐expressed NFs out of perikarya and into neurites, indicating that dynein participated in the initial anterograde delivery of NFs into neurites. Delivery of NFs to the axon hillock inner plasma membrane surface, and their subsequent translocation into neurites, was also prevented by vinblastine‐mediated inhibition of microtubule assembly. These data collectively suggest that some NFs enter axons as cargo of microtubues that are themselves undergoing transport into axons via dynein‐mediated interactions with the actin cortex and/or larger microtubules. C‐terminal NF phosphorylation regulates motor association, since anti‐dynein selectively coprecipitated extensively phosphorylated NFs, while anti‐kinesin selectively coprecipitated less phosphorylated NFs. In addition, however, the MAP kinase inhibitor PD98059 also inhibited transport of a constitutively‐phosphorylated NF construct, indicating that one or more additional, non‐NF phosphorylation events also regulated NF association with dynein or kinesin. Cell Motil. Cytoskeleton 2006. © 2006 Wiley‐Liss, Inc.
Bibliography:	istex:6E3B023C2E85497AB426C5CB58DF5DFDBDE0C73E ArticleID:CM20122 ark:/67375/WNG-5703NP76-Q National Science Foundation ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0886-1544 1097-0169
DOI:	10.1002/cm.20122