Novel Cell-autonomous Functions Of Neuronal Nogo-A

Nogo-A has been well-known as a myelin-derived neurite outgrowth inhibitor of thecentral nervous system (CNS) and the mechanism underlying axon regeneration is deeplyillustrated. However, accumulating evidence suggests that besides oligodendrocytes,Nogo-A is also expressed by sub-portions of neurons, especially in developing CNS.Although extensive studies have uncovered regulatory roles of Nogo-A in corticaldevelopment, precursor migration, growth cone activity, synaptic plasticity, neuriteoutgrowth and endomembrane system homeostasis, its cell-autonomous functions inneurons are largely uncharacterized. Furthermore, numerous investigations show that theexpression of neuronal Nogo-A obviously changes in various neurodegeneration diseasessuch as Seizure and Alzheimer's disease, however the relationship between the expressionof neuronal Nogo-A and these diseases is poorly understood. Our previous data show thatneuronal Nogo-A distributes at polyribosomes, rough endoplasmic reticulum (ER) andchromatins of the nucleus, bringing about its potent neuronal autonomous functions. In thepresent study, we try to explore the possible role of neuronal Nogo-A, and have got themain results as following:1. We systematically examine the expression pattern of neuronal Nogo-A usingimmunocytochemistry, subcellular separation and Western blot. Data show that, Nogo-Amainly localizes in cell body and neurites of cortical neurons, and living cell stainingshows amino-Nogo-A (186-1004aa) localizes intracellularly. In Cos7cell lines,overexpressed Nogo-A also distributes in ER, mitochondria, Golgi andmicrotubules-enriched regions. Interestingly, endogenous Nogo-A also localizes incentrosome of Neuro2A cell lines.2. TAT-mediated amino-Nogo-A protein transduction into cultured primary corticalneurons achieves an almost complete neuroprotection against oxidative stress induced byexogenous hydrogen peroxide (H2O2). Knockdown of neuronal Nogo-A results in more susceptibility to acute oxidative insults and markedly increases neuronal death. DHEexamination shows that amino-Nogo-A reduces reactive oxygen species (ROS) generationand then Western blot exhibits amino-Nogo-A can suppress extracellular signal-regulatedkinases (ERK) phosphorylation and decrease of Bcl-2induced by H2O2. Structure-functionmapping experiments reveal that, a novel region comprising residues290-562aa ofamino-Nogo-A is indispensable for preventing oxidative neuronal death. Furthermore, weget some evidence for the interaction between amino-Nogo-A and Prdx2through GST pulldown and Ni2+pull down. Moreover, mutagenesis analysis confirms that Cys residues424,464and559located in290-562aa are involved in interaction with peroxiredoxin2(Prdx2)and then reducing ROS generation.3. We discover the new functions of centrosome-localized Nogo-A. Endogenous andexogenous Nogo-A can regulate the translocation of acetylated α-tubulin in neurons andCos7cell lines individually. Down-regulation of Nogo-A leads to decreased neuronaldifferentiation rate and total neurites length in sodium valproate (VPA)-induced Neuro2Adifferentiation cell model. Thus, neuronal Nogo-A maybe positively regulate neuronaldifferentiation and neurites outgrowth through affecting centrosome-tubulin unit.In this study, our data suggest that neuronal Nogo-A might play a cell-autonomousrole in improving neuronal survival against oxidative insult through interaction with Prdx2and promoting neuronal differentiation and neurites outgrowth as centrosome-microtubuleassociated protein. These information can help us understand the multiple functions ofneuronal Nogo-A deeply and provide us new insights on therapeutic approach ofneurodegeneration diseases.