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Ndel1 promotes axon regeneration via intermediate filaments.

Toth C, Shim SY, Wang J, Jiang Y, Neumayer G, Belzil C, Liu WQ, Martinez J, Zochodne D, Nguyen MD - PLoS ONE (2008)

Bottom Line: Consistent with a role for Ndel1 in the conditioning lesion-induced neurite outgrowth of Dorsal Root Ganglion (DRG) neurons, the long lasting in vivo formation of the neuronal Ndel1/Vimentin complex was associated with robust axon regeneration.Furthermore, local silencing of Ndel1 in transected axons by siRNA severely reduced the extent of regeneration in vivo.Thus, Ndel1 promotes axonal regeneration; activating this endogenous repair mechanism may enhance neuroregeneration during acute and chronic axonal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Neurosciences, University of Calgary, Hotchkiss Brain Institute, Calgary, Canada.

ABSTRACT
Failure of axons to regenerate following acute or chronic neuronal injury is attributed to both the inhibitory glial environment and deficient intrinsic ability to re-grow. However, the underlying mechanisms of the latter remain unclear. In this study, we have investigated the role of the mammalian homologue of aspergillus nidulans NudE, Ndel1, emergently viewed as an integrator of the cytoskeleton, in axon regeneration. Ndel1 was synthesized de novo and upregulated in crushed and transected sciatic nerve axons, and, upon injury, was strongly associated with neuronal form of the intermediate filament (IF) Vimentin while dissociating from the mature neuronal IF (Neurofilament) light chain NF-L. Consistent with a role for Ndel1 in the conditioning lesion-induced neurite outgrowth of Dorsal Root Ganglion (DRG) neurons, the long lasting in vivo formation of the neuronal Ndel1/Vimentin complex was associated with robust axon regeneration. Furthermore, local silencing of Ndel1 in transected axons by siRNA severely reduced the extent of regeneration in vivo. Thus, Ndel1 promotes axonal regeneration; activating this endogenous repair mechanism may enhance neuroregeneration during acute and chronic axonal degeneration.

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Silencing Ndel1 by siRNA reduces lesion-conditioned neurite outgrowth in DRG neurons.(A) Co-localization of Ndel1 and Vimentin in the cell body and neurites of nearly all Vimentin-expressing DRG neurons. (B) Reduction of the conditioning lesion-induced neurite outgrowth of isolated DRG neurons treated with Ndel1 siRNA. Co-treatment with Ndel1 and Vimentin siRNAs does not further impede outgrowth. 5 to 7 different DRG cultures were performed and the total number of neurons analyzed is indicated. Typical examples of NF-positive DRG neuron with or without neurites are depicted. Note that only DRG neurons with a normal nuclear morphology were counted. * P (T<t) two tails: <0.001, ** P (T<t) two tails: <0.0001. Red: NF; blue: DAPI.
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pone-0002014-g005: Silencing Ndel1 by siRNA reduces lesion-conditioned neurite outgrowth in DRG neurons.(A) Co-localization of Ndel1 and Vimentin in the cell body and neurites of nearly all Vimentin-expressing DRG neurons. (B) Reduction of the conditioning lesion-induced neurite outgrowth of isolated DRG neurons treated with Ndel1 siRNA. Co-treatment with Ndel1 and Vimentin siRNAs does not further impede outgrowth. 5 to 7 different DRG cultures were performed and the total number of neurons analyzed is indicated. Typical examples of NF-positive DRG neuron with or without neurites are depicted. Note that only DRG neurons with a normal nuclear morphology were counted. * P (T<t) two tails: <0.001, ** P (T<t) two tails: <0.0001. Red: NF; blue: DAPI.

Mentions: To examine the role of Ndel1 in axon outgrowth following injury, we first investigated the conditioning lesion response of isolated DRG neurons treated with an Alexa 488-conjugated Ndel1 or control siRNA. We previously generated a specific siRNA against Ndel1 to knockdown the protein in primary neurons and mouse embryonic brains [18], [19]. An Alexa 488-conjugated scrambled sequence of siRNA without homology to any known protein was used as control. Sciatic nerve transections were performed 7 days prior to culture of neurons from L4/L5/L6 DRGs. Prelesioned conditioned neurons treated with Alexa 488-conjugated Ndel1 siRNA for 24 hours in culture incorporated the oligonucleotides (data not shown). Approximately 60% of cultured DRG neurons expressed Vimentin while nearly all Vimentin-positive DRG neurons express Ndel1. In these neurons, the two proteins co-localize in the cell body and neurites (Fig. 5A). Importantly, treatment of lesion-induced DRG neurons with Ndel1 siRNA reduced the number of neurons undergoing neurite outgrowth up to 50% as determined by staining with NF antibodies (Fig. 5B). Furthermore, co-treatment of lesion-induced DRG neurons with Ndel1 and Vimentin siRNAs did not further reduce the number of neurons extending neurites (Fig. 5B). These results indicated that Ndel1 is important for lesion-induced neurite outgrowth of DRG neurons in vitro. Based on data indicating that the two proteins associate together during neurite outgrowth [23], these results suggested that Ndel1 works upstream or at least serially with neuronal Vimentin in the same pathway.


Ndel1 promotes axon regeneration via intermediate filaments.

Toth C, Shim SY, Wang J, Jiang Y, Neumayer G, Belzil C, Liu WQ, Martinez J, Zochodne D, Nguyen MD - PLoS ONE (2008)

Silencing Ndel1 by siRNA reduces lesion-conditioned neurite outgrowth in DRG neurons.(A) Co-localization of Ndel1 and Vimentin in the cell body and neurites of nearly all Vimentin-expressing DRG neurons. (B) Reduction of the conditioning lesion-induced neurite outgrowth of isolated DRG neurons treated with Ndel1 siRNA. Co-treatment with Ndel1 and Vimentin siRNAs does not further impede outgrowth. 5 to 7 different DRG cultures were performed and the total number of neurons analyzed is indicated. Typical examples of NF-positive DRG neuron with or without neurites are depicted. Note that only DRG neurons with a normal nuclear morphology were counted. * P (T<t) two tails: <0.001, ** P (T<t) two tails: <0.0001. Red: NF; blue: DAPI.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2291557&req=5

pone-0002014-g005: Silencing Ndel1 by siRNA reduces lesion-conditioned neurite outgrowth in DRG neurons.(A) Co-localization of Ndel1 and Vimentin in the cell body and neurites of nearly all Vimentin-expressing DRG neurons. (B) Reduction of the conditioning lesion-induced neurite outgrowth of isolated DRG neurons treated with Ndel1 siRNA. Co-treatment with Ndel1 and Vimentin siRNAs does not further impede outgrowth. 5 to 7 different DRG cultures were performed and the total number of neurons analyzed is indicated. Typical examples of NF-positive DRG neuron with or without neurites are depicted. Note that only DRG neurons with a normal nuclear morphology were counted. * P (T<t) two tails: <0.001, ** P (T<t) two tails: <0.0001. Red: NF; blue: DAPI.
Mentions: To examine the role of Ndel1 in axon outgrowth following injury, we first investigated the conditioning lesion response of isolated DRG neurons treated with an Alexa 488-conjugated Ndel1 or control siRNA. We previously generated a specific siRNA against Ndel1 to knockdown the protein in primary neurons and mouse embryonic brains [18], [19]. An Alexa 488-conjugated scrambled sequence of siRNA without homology to any known protein was used as control. Sciatic nerve transections were performed 7 days prior to culture of neurons from L4/L5/L6 DRGs. Prelesioned conditioned neurons treated with Alexa 488-conjugated Ndel1 siRNA for 24 hours in culture incorporated the oligonucleotides (data not shown). Approximately 60% of cultured DRG neurons expressed Vimentin while nearly all Vimentin-positive DRG neurons express Ndel1. In these neurons, the two proteins co-localize in the cell body and neurites (Fig. 5A). Importantly, treatment of lesion-induced DRG neurons with Ndel1 siRNA reduced the number of neurons undergoing neurite outgrowth up to 50% as determined by staining with NF antibodies (Fig. 5B). Furthermore, co-treatment of lesion-induced DRG neurons with Ndel1 and Vimentin siRNAs did not further reduce the number of neurons extending neurites (Fig. 5B). These results indicated that Ndel1 is important for lesion-induced neurite outgrowth of DRG neurons in vitro. Based on data indicating that the two proteins associate together during neurite outgrowth [23], these results suggested that Ndel1 works upstream or at least serially with neuronal Vimentin in the same pathway.

Bottom Line: Consistent with a role for Ndel1 in the conditioning lesion-induced neurite outgrowth of Dorsal Root Ganglion (DRG) neurons, the long lasting in vivo formation of the neuronal Ndel1/Vimentin complex was associated with robust axon regeneration.Furthermore, local silencing of Ndel1 in transected axons by siRNA severely reduced the extent of regeneration in vivo.Thus, Ndel1 promotes axonal regeneration; activating this endogenous repair mechanism may enhance neuroregeneration during acute and chronic axonal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Neurosciences, University of Calgary, Hotchkiss Brain Institute, Calgary, Canada.

ABSTRACT
Failure of axons to regenerate following acute or chronic neuronal injury is attributed to both the inhibitory glial environment and deficient intrinsic ability to re-grow. However, the underlying mechanisms of the latter remain unclear. In this study, we have investigated the role of the mammalian homologue of aspergillus nidulans NudE, Ndel1, emergently viewed as an integrator of the cytoskeleton, in axon regeneration. Ndel1 was synthesized de novo and upregulated in crushed and transected sciatic nerve axons, and, upon injury, was strongly associated with neuronal form of the intermediate filament (IF) Vimentin while dissociating from the mature neuronal IF (Neurofilament) light chain NF-L. Consistent with a role for Ndel1 in the conditioning lesion-induced neurite outgrowth of Dorsal Root Ganglion (DRG) neurons, the long lasting in vivo formation of the neuronal Ndel1/Vimentin complex was associated with robust axon regeneration. Furthermore, local silencing of Ndel1 in transected axons by siRNA severely reduced the extent of regeneration in vivo. Thus, Ndel1 promotes axonal regeneration; activating this endogenous repair mechanism may enhance neuroregeneration during acute and chronic axonal degeneration.

Show MeSH
Related in: MedlinePlus