αvβ5 integrin mediates the effect of vitronectin on the initial stage of differentiation in mouse cerebellar granule cell precursors

Vitronectin (VN), one of the extracellular matrix proteins, controls the maturation of cerebellar granule cells (CGCs) through the promotion of the initial differentiation stage progress. However, the receptors of VN in the initial differentiation stage of CGC precursors (CGCPs) have not been clarified. In this study, we characterized the receptor candidates for VN in CGCPs. First, we confirmed that alpha v beta 3 and alpha v beta 5 integrins, which are receptor candidates for VN, were co-localized with VN in the developing cerebellum and primary cultured CGCPs. Next, the knockdown (KD) of alpha v, beta 3, and beta 5 integrins with small interference RNA (siRNA) for each integrin reduced the ratio of Tuj1, a final differentiation marker, -positive CGCPs. We further studied whether alpha v beta 3 and alpha v beta 5 integrins control the initial differentiation stage. The KD of alpha v and beta 5, but not beta 3, integrins significantly increased the ratio of transient axonal glycoprotein 1 (TAG1), an initial differentiation marker,-positive CGCPs, whereas the KD of alpha v and beta 3 integrins, not beta 5 integrin, stimulated the proliferation of CGCPs. Overexpression of beta 5 integrin stimulated the progress of the initial differentiation stage as well. To confirm the interaction between alpha v beta 5 integrin and VN, VN was added to beta 5 integrin-KD CGCPs. The promotion of the progress of initial differentiation by VN was abrogated by beta 5 integrin KD using small hairpin RNA (shRNA). Taken together, our results indicated that alpha v beta 5 integrin, as the very receptor of VN, is responsible for the progress of the initial differentiation stage in mouse CGCPs. (C) 2018 Elsevier B.V. All rights reserved.