Recent studies have suggested that neuronal death in Alzheimer's disease or ischemia could arise from dysfunction of the endoplasmic reticulum (ER). Although caspase-12 has been implicated in ER stress-induced apoptosis and amyloid-β (Aβ)–induced apoptosis in rodents, it is controversial whether similar mechanisms operate in humans. We found that human caspase-4, a member of caspase-1 subfamily that includes caspase-12, is localized to the ER membrane, and is cleaved when cells are treated with ER stress-inducing reagents, but not with other apoptotic reagents. Cleavage of caspase-4 is not affected by overexpression of Bcl-2, which prevents signal transduction on the mitochondria, suggesting that caspase-4 is primarily activated in ER stress-induced apoptosis. Furthermore, a reduction of caspase-4 expression by small interfering RNA decreases ER stress-induced apoptosis in some cell lines, but not other ER stress-independent apoptosis. Caspase-4 is also cleaved by administration of Aβ, and Aβ-induced apoptosis is reduced by small interfering RNAs to caspase-4. Thus, caspase-4 can function as an ER stress-specific caspase in humans, and may be involved in pathogenesis of Alzheimer's disease.

Repulsive guidance molecule (RGM) is a protein implicated in both axonal guidance and neural tube closure. We report RGMa as a potent inhibitor of axon regeneration in the adult central nervous system (CNS). RGMa inhibits mammalian CNS neurite outgrowth by a mechanism dependent on the activation of the RhoA–Rho kinase pathway. RGMa expression is observed in oligodendrocytes, myelinated fibers, and neurons of the adult rat spinal cord and is induced around the injury site after spinal cord injury. We developed an antibody to RGMa that efficiently blocks the effect of RGMa in vitro. Intrathecal administration of the antibody to rats with thoracic spinal cord hemisection results in significant axonal growth of the corticospinal tract and improves functional recovery. Thus, RGMa plays an important role in limiting axonal regeneration after CNS injury and the RGMa antibody offers a possible therapeutic agent in clinical conditions characterized by a failure of CNS regeneration.

Netrin-1 induces repulsive axon guidance by binding to the mammalian Unc5 receptor family (Unc5A-Unc5D). Mouse genetic analysis of selected members of the Unc5 family, however, revealed essential functions independent of Netrin-1, suggesting the presence of other ligands. Unc5B was recently shown to bind fibronectin and leucine-rich transmembrane protein-3 (FLRT3), although the relevance of this interaction for nervous system development remained unclear. Here, we show that the related Unc5D receptor binds specifically to another FLRT protein, FLRT2. During development, FLRT2/3 ectodomains (ECDs) are shed from neurons and act as repulsive guidance molecules for axons and somata of Unc5-positive neurons. In the developing mammalian neocortex, Unc5D is expressed by neurons in the subventricular zone (SVZ), which display delayed migration to the FLRT2-expressing cortical plate (CP). Deletion of either FLRT2 or Unc5D causes a subset of SVZ-derived neurons to prematurely migrate towards the CP, whereas overexpression of Unc5D has opposite effects. Hence, the shed FLRT2 and FLRT3 ECDs represent a novel family of chemorepellents for Unc5-positive neurons and FLRT2/ Unc5D signalling modulates cortical neuron migration.

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...