Myelin-associated glycoprotein (MAG) and Nogo are potent inhibitors of neurite outgrowth from a variety of neurons, and they have been identified as possible components of the central nervous system myelin that prevents axonal regeneration in the adult vertebrate central nervous system. The activation of RhoA and Rhokinase is reported to be an essential part of the signaling mechanism of these proteins. Here, we report that the collapsing response mediator protein-2 (CRMP-2) is phosphorylated by a Rho-kinasedependent mechanism downstream of MAG or Nogo-66. The overexpression of the nonphosphorylated form of CRMP-2 at threonine 555, which is the phosphorylation site for Rho-kinase, counteracts the inhibitory effect of MAG on the postnatal cerebellar neurons. Additionally, the expression of the dominant negative form of CRMP-2 or knockdown of the gene using small interference RNA (siRNA) mimics the effect of MAG in vitro. Consistent with the function of CRMP-2, which promotes microtubule assembly, microtubule levels are down-regulated in the cerebellar neurons that are stimulated with MAG in vitro. Reduction in the density of microtubules is also observed in the injured axons following the spinal cord injury, and this effect depends on the Rho-kinase activity. Our data suggest the important roles of CRMP-2 and microtubules in the inhibition of the axon regeneration by the myelin-derived inhibitors.Several myelin-derived proteins have been identified as components of the central nervous system myelin that prevents axonal regeneration in the adult vertebrate central nervous system. To date, three major inhibitors that are expressed by oligodendrocytes and myelinated fiber tracts have been identified (1). These are Nogo, myelin-associated glycoprotein (MAG), 2 and oligodendrocyte-myelin glycoprotein. All these proteins act on neurons through the p75 receptor (p75) (2-4) in complex with the Nogo receptor. One potential clue to understanding the signal transduction mechanism downstream of p75 and the Nogo receptor is found through observations that demonstrate the small GTPase RhoA as a key intracellular effector for growth inhibitory signaling by myelin. In its active GTP-bound form, RhoA rigidifies the actin cytoskeleton, thereby inhibiting axon elongation and mediating growth cone collapse. RhoA is activated by MAG, Nogo-66, and oligodendrocyte-myelin glycoprotein through a p75-dependent mechanism; thus, inhibiting neurite outgrowth from postnatal sensory neurons and cerebellar neurons (2-5). The regulation of RhoA activity by MAG and Nogo through p75 is mediated by the release of RhoA from Rho GDI, which suppresses the RhoA activity (6, 7).Although RhoA and one of its effectors, Rho-kinase, appear to play a key role in regulating axon growth, the mechanism by which the myelinderived proteins regulate axon outgrowth remains to be elucidated. The outgrowth of axons is based on the dynamic rearrangement of the cytoskeleton (8). Cues that influence the axon outgrowth are sensed by the growth cone, which is a highly mot...

Several myelin-derived proteins have been identified as components of central nervous system (CNS) myelin, which prevents axonal regeneration in the adult vertebrate CNS. The discovery of the receptor for these proteins was a major step toward understanding the failure of axon regeneration. The receptor complex consists of at least three elements: the p75 receptor (p75NTR), the Nogo receptor and LINGO-1. Downstream from the receptor complex, RhoA activation has been shown to be a key element of the signaling mechanism of these proteins. Rho activation arrests axon growth, and blocking Rho activation promotes axon regeneration in vivo. Recent studies have identified conventional protein kinase C as an additional necessary component for axon growth inhibition. Possible crosstalk downstream of these signals should be explored to clarify all the inhibitory signals and may provide an efficient molecular target against injuries to the CNS.

Local axon degeneration is a common pathological feature of many neurodegenerative diseases, whereas the underlying molecular mechanisms are largely unknown. In this study, we used the degeneration of transected axons, termed "Wallerian degeneration," as a model to examine the possible involvement of Rho. Nogo-66, a myelin-derived inhibitor of axon regeneration, significantly accelerated axon degeneration of the dorsal root ganglion explant in vitro, whereas inhibiting Rho-kinase activity abolished the effect. Rho activation was observed in the distal part of the injured axons after spinal cord injury. We demonstrate that degeneration of the injured cortico-spinal axons was significantly retarded by a Rho-kinase inhibitor in vivo. Our findings suggest that inhibiting the signaling pathway may retard axon degeneration in pathological conditions.Axon degeneration occurs frequently in many types of chronic neurodegenerative diseases and in injuries to axons caused by toxic, ischemic, or traumatic insults (1, 2). It may lead to separation of the neurons from their targets, resulting in the loss of neuronal function. Effort has focused on understanding the nature of neuronal cell death in these diseases (3); however, strategies designed to prevent neuronal cell death have resulted in only limited success in preventing or delaying neurodegeneration. Interfering with the process of axon degeneration may represent an additional and complementary therapeutic avenue for these diseases.The simplest model of axon degeneration known to date is the self-destructive process observed at the distal portion of a transected axon upon injury, termed "Wallerian degeneration." In vertebrates, the distal part of an axon can remain viable and conduct action potentials in vivo for up to a few days, after which they undergo rapid structural destruction in which the axolemma and axonal cytoskeleton are dismantled. Interestingly, the axons undergoing Wallerian degeneration do not seem to possess detectable activation of the caspase family cysteine proteases (4), suggesting that Wallerian degeneration and apoptosis may represent two distinct self-destruct programs. In Wallerian degeneration slow (Wld s ) mice, Wallerian degeneration in response to axonal injury is delayed because of a mutation that results in the overexpression of a chimeric protein (Wld s ) composed of the ubiquitin assembly protein, Ufd2a, and nicotinamide adenine dinucleotide (NAD) biosynthetic enzyme, Nmnat1. Increased Nmnat activity was recently shown to be responsible for the axon-sparing activity of Wld s protein (5). Downstream of Nmnat, Sirt1, a mammalian ortholog of Sir2, contributes to axonal protection. This discovery is a major step for understanding the molecular mechanism of Wallerian degeneration; however, the precise mechanism remains unknown.Given that Wallerian degeneration is an active process, we hypothesized that the self-destruction process of injured axons might be a type of cytoskeletal reorganization. Rho GTPases are a family of highly related prot...

The pan-neurotrophin receptor p75NTR belongs to a large family of receptors, which includes tumor necrosis factor receptors, Fas and approximately 25 other members. The p75NTR is the first receptor to be cloned molecularly. Recent years have seen the emergence of a consensus regarding the signaling pathways activated by p75NTR and its potential biological function, although receptor characterization had not been targeted for some years. We now know that p75NTR has surprisingly diverse effects, ranging from cell death to regulation of axon elongation. This diversity can be explained by the complex formation of p75NTR with other receptors and multiple signaling molecules that interact with the intracellular domain of p75NTR.

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