Abstract
CNS neurons are generally incapable of regenerating their axons after injury due to several intrinsic and extrinsic factors, including the presence of axon growth inhibitory molecules. One such potent inhibitor of CNS axon regeneration is Reticulon (RTN) 4 or Nogo-A. Here, we focused on RTN3 as its contribution to CNS axon regeneration is currently unknown. We found that RTN3 expression correlated with an axon regenerative phenotype in dorsal root ganglion neurons (DRGN) after injury to the dorsal columns, a well-characterised model of spinal cord injury. Overexpression of RTN3 promoted disinhibited DRGN neurite outgrowth in vitro and dorsal column axon regeneration/sprouting and electrophysiological, sensory and locomotor functional recovery after injury in vivo. Knockdown of protrudin, however, ablated RTN3-enhanced neurite outgrowth/axon regeneration in vitro and in vivo. Moreover, overexpression of RTN3 in a second model of CNS injury, the optic nerve crush injury model, enhanced retinal ganglion cell (RGC) survival, disinhibited neurite outgrowth in vitro and survival and axon regeneration in vivo, an effect that was also dependent on protrudin. These results demonstrate that RTN3 enhances neurite outgrowth/axon regeneration in a protrudin-dependent manner after both spinal cord and optic nerve injury.
Highlights
We used Quantitative RT-PCR (qRT-PCR) to demonstrate that RTN3 levels were upregulated by 4.2 ± 0.05- and 6.2 ± 0.04-fold in regenerating sciatic nerve (SN) and preconditioning SN (pSN) + dorsal column (DC) dorsal root ganglion neurons (DRGN) compared to those harvested after DC injury (Figure 1B)
In regenerating SN and pSN + DC DRGN, RTN3 immunoreactivity was widely observed in the cytoplasm and cell membranes of DRGN after DC injury (Figure 1E). These results demonstrated that elevated levels of RTN3 correlated with an axon regenerative phenotype and that RTN3 was mainly localised in the cytoplasm of DRGN
We show that RTN3 levels correlated with an axon regenerative phenotype and that overexpression of RTN3 enhanced DRGN neurite outgrowth, which was dependent on protrudin function
Summary
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Reticulons (RTNs) are membrane-bound proteins of which four have been identified in mammals and termed RTN1, 2, 3 and 4 (Nogo). RTNs are characterised by highly conserved reticulon homology domains (RHD); approximately 150–200 amino acids are located in the RTN C-terminal, which are involved in the improvement of the localisation and function of these proteins. RTNs are found in a variety of tissues including the brain, kidney, spleen and liver, playing diverse roles such as ER shaping and morphology, membrane trafficking and apoptosis. RTNs are found in neurodegenerative diseases such as Alzheimer’s, amyotrophic lateral sclerosis and multiple sclerosis [1]. RTN1, 2 and 4 have three splice isoforms A, B and C whilst RTN3 only has two spliced isoforms RTN3A and
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