Borna Disease Virus Requires Cholesterol in both Cellular Membrane and Viral Envelope for Efficient Cell Entry

Abstract

Borna disease virus (BDV), the prototypic member of the family Bornaviridae within the order Mononegavirales, provides an important model for the investigation of viral persistence within the central nervous system (CNS) and of associated brain disorders. BDV is highly neurotropic and enters its target cell via receptor-mediated endocytosis, a process mediated by the virus surface glycoprotein (G), but the cellular factors and pathways determining BDV cell tropism within the CNS remain mostly unknown. Cholesterol has been shown to influence viral infections via its effects on different viral processes, including replication, budding, and cell entry. In this work, we show that cell entry, but not replication and gene expression, of BDV was drastically inhibited by depletion of cellular cholesterol levels. BDV G-mediated attachment to BDV-susceptible cells was cholesterol independent, but G localized to lipid rafts (LR) at the plasma membrane. LR structure and function critically depend on cholesterol, and hence, compromised structural integrity and function of LR caused by cholesterol depletion likely inhibited the initial stages of BDV cell internalization. Furthermore, we also show that viral-envelope cholesterol is required for BDV infectivity.Borna disease virus (BDV) is an enveloped virus with a nonsegmented negative-strand RNA genome whose organization (3Ј-N-p10/P-M-G-L-5Ј) is characteristic of mononegaviruses (6, 28, 46, 48). However, based on its unique genetics and biological features, BDV is considered to be the prototypic member of a new virus family, Bornaviridae, within the order Mononegavirales (8,28,46,49).BDV can infect a variety of cell types in cell culture but in vivo exhibits exquisite neurotropism and causes central nervous system (CNS) disease in different vertebrate species, which is frequently manifested in behavioral abnormalities (19,33,44,53). Both host and viral factors contribute to a variable period of incubation and heterogeneity in the symptoms and pathology associated with BDV infection (14,16,29,42,44). BDV provides an important model for the investigation of both immune-mediated pathological events associated with virus-induced neurological disease and mechanisms whereby noncytolytic viruses induce neurodevelopmental and behavioral disturbances in the absence of inflammation (15,18,41). Moreover, serological data and molecular epidemiological studies suggest that BDV, or a BDV-like virus, can infect humans and that it might be associated with certain neuropsychiatric disorders (17, 24), which further underscores the interest in understanding the mechanisms underlying BDV persistence in the CNS and its effect on brain cell functions. The achievement of these goals will require the elucidation of the determinants of BDV cell tropism within the CNS.

Molecular Chaperone BiP Interacts with Borna Disease Virus Glycoprotein at the Cell Surface

“…1B, GP1-TAP reduced BDV infection by 40% compared to levels for mock-treated cells. This result was consistent with earlier reports showing that recombinant GP1 protein binds to the cell surface and inhibits BDV infection (6,20).…”

“…1B, GP1-TAP reduced BDV infection by 40% compared to levels for mock-treated cells. This result was consistent with earlier reports showing that recombinant GP1 protein binds to the cell surface and inhibits BDV infection (6,20).To investigate the host factor(s) that mediates the interaction of GP1 with the cell surface, a combination of tandem affinity purification (TAP) and liquid chromatography tandem mass spectrometry analyses was designed (13). We transfected GP1-TAP into OL cells and then purified GP1 from cell homogenates using a TAP strategy.…”

“…On the other hand, a BDV-nonpermissive cell line, HeLa, appeared to express BiP on the cell surface, suggesting that the cell surface BiP may not be necessarily involved in the infectivity of BDV. A recent study by Clemente et al (6) revealed that following initial attachment to the cell surface, BDV is recruited to the plasma membrane lipid raft (LR) prior to internalization of the particles. The study suggested that BDV may use the LR as a platform to interact with additional host cell factor(s) required for efficient BDV internalization.…”

Cell Entry of Borna Disease Virus Follows a Clathrin-Mediated Endocytosis Pathway That Requires Rab5 and Microtubules

“…Previous studies have documented that BDV enters cells via receptor-mediated endocytosis (19) and that cholesterol plays a critical role in BDV cell entry (9). However, many of the details related to BDV cell entry have not been investigated yet.…”

“…This endocytotic pathway is strictly dependent on recruitment of lipid rafts to the cell surface, an event mediated by cholesterol. In this regard, we have recently documented the requirements of cholesterol and structural integrity of cell surface lipid rafts for efficient cell entry of BDV (9).…”