Borna disease virus phosphoprotein binds a neurite outgrowth factor, amphoterin/HMG-1

The Borna disease virus (BDV) p24 phosphoprotein is an abundant protein in BDV-infected cultured cells and animal brains. Therefore, there is a possib ility that binding of the p24 protein to cellular factor(s) induces functio nal alterations of infected neural cells in the brain. To identify a cellul ar protein(s) that interacts with BDV p24 protein, we performed far-Western blotting with extracts from various cell lines. Using recombinant p24 prot ein as a probe, we detected a 30-kDa protein in all cell lines examined. Bi nding between the 30-kDa and BDV p24 proteins was also demonstrated using B DV p24 affinity and ion-exchange chromatography columns. Microsequence anal ysis of the purified 30-kDa protein revealed that its N terminus showed com plete homology with rat amphoterin protein, which is a neurite outgrowth fa ctor abundant in the brain during development. Mammalian two-hybrid and imm unoprecipitation analyses also confirmed that amphoterin is a specific targ et for the p24 protein in vivo. Furthermore, we showed that infection by BD V, as well as purified p24 protein in the medium, significantly decreased c ell process outgrowth of cells grown on laminin, indicating the functional inhibition of amphoterin by interaction with the p24 protein. Immunohistoch emical analysis revealed decreased levels of amphoterin protein at the lead ing edges of BDV-infected cells. Moreover, the expression of the receptor f or advanced glycation end products, of which the extracellular moiety is a receptor for amphoterin, was not significantly activated in BDV-infected ce lls during the process of extension, suggesting that the secretion of ampho terin from the cell surface is inhibited by the binding of the p24 protein. These results suggested that BDV infection may cause direct damage in the developing brain by inhibiting the function of amphoterin due to binding by the p24 phosphoprotein.