RT Journal Article
SR Electronic
T1 Trk Activation of the ERK1/2 Kinase Pathway Stimulates Intermediate Chain Phosphorylation and Recruits Cytoplasmic Dynein to Signaling Endosomes for Retrograde Axonal Transport
JF The Journal of Neuroscience
JO J. Neurosci.
FD Society for Neuroscience
SP 15495
OP 15510
DO 10.1523/JNEUROSCI.5599-11.2012
VO 32
IS 44
A1 David J. Mitchell
A1 Kiev R. Blasier
A1 Erin D. Jeffery
A1 Mitchell W. Ross
A1 Ashok K. Pullikuth
A1 Dong Suo
A1 Juyeon Park
A1 W. Russell Smiley
A1 Kevin W.-H. Lo
A1 Jeffrey Shabanowitz
A1 Christopher D. Deppmann
A1 Jonathan C. Trinidad
A1 Donald F. Hunt
A1 Andrew D. Catling
A1 K. Kevin Pfister
YR 2012
UL http://www.jneurosci.org/content/32/44/15495.abstract
AB The retrograde transport of Trk-containing endosomes from the axon to the cell body by cytoplasmic dynein is necessary for axonal and neuronal survival. We investigated the recruitment of dynein to signaling endosomes in rat embryonic neurons and PC12 cells. We identified a novel phosphoserine on the dynein intermediate chains (ICs), and we observed a time-dependent neurotrophin-stimulated increase in intermediate chain phosphorylation on this site in both cell types. Pharmacological studies, overexpression of constitutively active MAP kinase kinase, and an in vitro assay with recombinant proteins demonstrated that the intermediate chains are phosphorylated by the MAP kinase ERK1/2, extracellular signal-regulated kinase, a major downstream effector of Trk. Live cell imaging with fluorescently tagged IC mutants demonstrated that the dephosphomimic mutants had significantly reduced colocalization with Trk and Rab7, but not a mitochondrial marker. The phosphorylated intermediate chains were enriched on immunoaffinity-purified Trk-containing organelles. Inhibition of ERK reduced the amount of phospho-IC and the total amount of dynein that copurified with the signaling endosomes. In addition, inhibition of ERK1/2 reduced the motility of Rab7- and TrkB-containing endosomes and the extent of their colocalization with dynein in axons. NGF-dependent survival of sympathetic neurons was significantly reduced by the overexpression of the dephosphomimic mutant IC-1B-S80A, but not WT IC-1B, further demonstrating the functional significance of phosphorylation on this site. These results demonstrate that neurotrophin binding to Trk initiates the recruitment of cytoplasmic dynein to signaling endosomes through ERK1/2 phosphorylation of intermediate chains for their subsequent retrograde transport in axons.