Vitronectin, an extracellular matrix protein, controls the differentiation of cerebellar granule cell precursors (CGCPs) via αvβ5 integrin, particularly in the initial stage of differentiation to granule cells. In this study, we determined whether vitronectin regulates axon specification in this initial differentiation stage of CGCPs. First, we analyzed whether vitronectin deficiency, β5 integrin knockdown (KD), and β5 integrin overexpression affect axon specification of primary cultured CGCPs. Vitronectin deficiency and β5 integrin KD inhibited axon formation, while vitronectin administrated- and β5 integrin overexpressed-neurons formed multiple axons. Moreover, KD of β5 integrin suppressed vitronectin-induced multiple axon formation. These findings indicate that vitronectin contributes to regulating axon specification via αvβ5 integrin in CGCPs. Next, we determined the signaling pathway involved in regulating vitronectin-induced axon specification. Wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3K), inhibited vitronectin-induced multiple axon specification, and lithium chloride, an inhibitor of glyocogen synthase kinase 3 beta (GSK3β), attenuated the inhibitory effect of vitronectin-KO and β5 integrin KD on the specification of CGCPs. In addition, vitronectin induced the phosphorylation of protein kinase B (Akt) and GSK3β in neuroblastoma Neuro2a cells. Taken together, our results indicate that vitronectin plays an important factor in axon formation process in CGCPs via a β5 integrin/PI3K/GSK3β pathway.
Keywords: Axon specification; Cerebellar granule cell; GSK3; Integrin; PI3K; Vitronectin.
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