Cholesterol-Rich Microdomains as Docking Platforms for Respiratory Syncytial Virus in Normal Human Bronchial Epithelial Cells

Abstract

Respiratory syncytial virus (RSV) is one of the major causes of respiratory infections in children, and it is the main pathogen causing bronchiolitis in infants. The binding and entry mechanism by which RSV infects respiratory epithelial cells has not yet been determined. In this study, the earliest stages of RSV infection in normal human bronchial epithelial cells were probed by tracking virions with fluorescent lipophilic dyes in their membranes. Virions colocalized with cholesterol-containing plasma membrane microdomains, identified by their ability to bind cholera toxin subunit B. Consistent with an important role for cholesterol in RSV infection, cholesterol depletion profoundly inhibited RSV infection, while cholesterol repletion reversed this inhibition. Merger of the outer leaflets of the viral envelope and the cell membrane appeared to be triggered at these sites. Using small-molecule inhibitors, RSV infection was found to be sensitive to Pak1 inhibition, suggesting the requirement of a subsequent step of cytoskeletal reorganization that could involve plasma membrane rearrangements or endocytosis. It appears that RSV entry depends on its ability to dock to cholesterol-rich microdomains (lipid rafts) in the plasma membrane where hemifusion events begin, assisted by a Pak1-dependent process.

Respiratory Syncytial Virus: Infection, Detection, and New Options for Prevention and Treatment

“…1). This internalization occurs when the RSV envelope fuses with the host cell at or near the plasma membrane on lipid raft platforms (96), and fusion is triggered in a pH-independent manner. There was a recent report showing that prior to fusion, RSV is internalized in an actin-dependent process called macropinocytosis which also appears to require the host cell protein Rab5 in HeLa cells (95).…”

Cholesterol Induces Uneven Curvature of Asymmetric Lipid Bilayers

“…Particularly, the respiratory syncytial virus (RSV), which causes respiratory infections in children, binds to cholesterol-rich lipid rafts in the plasma membrane of human bronchial epithelial cells [30]. Moreover, the release of RSV virus particles from infected cells also requires cholesterol-rich lipid rafts [31].…”

Cholesterol is required for stability and infectivity of influenza A and respiratory syncytial viruses

“…First, we investigated whether depletion of cellular cholesterol by Gem and Lov inhibits virus entry process in human airway cells, because a previous study suggested that lipid raft serves as an entry platform for RSV (San-Juan-Vergara et al, 2012). Cells treated with Gem (400 μM) and Lov (40 μM) for 24 h were infected with the viruses at MOI of 0.5 for 24 h. Cells were then fixed and the number of infected cells were determined by immunostaining using specific anti-NP monoclonal antibodies (mAb).…”

“…These results highlighted the difference in requirement of raft integrity for virus budding and release among respiratory RNA viruses. All of these enveloped viruses are known to utilize ordered lipid raft domains to localize essential viral components during assembly (Nayak et al, 2009; Nayak et al, 2004; Oomens et al, 2006; San-Juan-Vergara et al, 2012). Cholesterol seems to be required for cell surface accumulation of SeV and hPIV1, but not for IAV and RSV glycoproteins (Fig.…”