Doublecortin and JIP3 are neural-specific counteracting regulators of dynein-mediated retrograde trafficking
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In their paper, Rao, Li et al. explore the mechanisms by which the microtubule-associated protein, doublecortin (DCX), functions in regulating retrograde transport in neurons. They find that DCX affects the dynein-microtubule interaction to perturb its motion. Impressively, they reconstitute a dynein-dynactin-JIP3 complex, validating JIP3 as a bona fide adaptor, and show that DCX disrupts the transport of this processive complex. This mechanism will be useful in understanding how mutations in DCX cause lissencephaly and this paper will be of interest to those in the cytoskeletal and neurobiology fields.
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Abstract
Mutations in the microtubule (MT)-binding protein doublecortin (DCX) or in the MT-based molecular motor dynein result in lissencephaly. However, a functional link between DCX and dynein has not been defined. Here, we demonstrate that DCX negatively regulates dynein-mediated retrograde transport in neurons from Dcx -/y or Dcx -/y ;Dclk1 -/- mice by reducing dynein’s association with MTs and disrupting the composition of the dynein motor complex. Previous work showed an increased binding of the adaptor protein C-Jun-amino-terminal kinase-interacting protein 3 (JIP3) to dynein in the absence of DCX. Using purified components, we demonstrate that JIP3 forms an active motor complex with dynein and its cofactor dynactin with two dyneins per complex. DCX competes with the binding of the second dynein, resulting in a velocity reduction of the complex. We conclude that DCX negatively regulates dynein-mediated retrograde transport through two critical interactions by regulating dynein binding to MTs and regulating the composition of the dynein motor complex.
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eLife assessment
In their paper, Rao, Li et al. explore the mechanisms by which the microtubule-associated protein, doublecortin (DCX), functions in regulating retrograde transport in neurons. They find that DCX affects the dynein-microtubule interaction to perturb its motion. Impressively, they reconstitute a dynein-dynactin-JIP3 complex, validating JIP3 as a bona fide adaptor, and show that DCX disrupts the transport of this processive complex. This mechanism will be useful in understanding how mutations in DCX cause lissencephaly and this paper will be of interest to those in the cytoskeletal and neurobiology fields.
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Reviewer #1 (Public Review):
Mutations in Doublecortin (DCX), which is a microtubule-binding protein cause lissencephaly. This manuscript by Rao et al. demonstrates the mechanism by which DCX affects retrograde transport in the neurons. Authors show that DCX functions to affect dynein transport in the axon via two different mechanisms - 1. By regulating dynein-microtubule interaction and 2. By regulating the interaction of JIP3 to the dynein motor complex. Interestingly, they have also shown the formation of the dynein-dynactin-JIP3 complex and reconstituted its motility in vitro. Authors demonstrate DCX regulation by affecting the recruitment of the second dynein in the dynein-dynactin-JIP3 complex to affect dynein velocity. Because DCX also regulates Kinesin-3 mediated transport, this work uncovers the role of DCX in regulating …
Reviewer #1 (Public Review):
Mutations in Doublecortin (DCX), which is a microtubule-binding protein cause lissencephaly. This manuscript by Rao et al. demonstrates the mechanism by which DCX affects retrograde transport in the neurons. Authors show that DCX functions to affect dynein transport in the axon via two different mechanisms - 1. By regulating dynein-microtubule interaction and 2. By regulating the interaction of JIP3 to the dynein motor complex. Interestingly, they have also shown the formation of the dynein-dynactin-JIP3 complex and reconstituted its motility in vitro. Authors demonstrate DCX regulation by affecting the recruitment of the second dynein in the dynein-dynactin-JIP3 complex to affect dynein velocity. Because DCX also regulates Kinesin-3 mediated transport, this work uncovers the role of DCX in regulating opposite polarity motors during neuronal growth. Overall the manuscript is well written, the work is original, experiments are performed carefully and most of the findings justify the conclusions drawn by the authors.
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Reviewer #2 (Public Review):
The authors have tried to provide a molecular mechanism for the observation that the lack of DCX increases run lengths of retrogradely moving cargo. The authors show a direct interaction of DCX with Dynein and that this direct interaction is the key means by which to regulate dynein-dependent retrograde run lengths of cargo. DCX seems to have a dual role - on microtubules where it appears to prevent attachment of dynein to microtubules. DCX also appears to reduce JIP3 binding to dynein.
A major strength is that they have used a combination of approaches including in vitro motility assays to support their arguments.
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