Apical recycling systems regulate directional budding of respiratory syncytial virus from polarized epithelial cells

Abstract

Respiratory syncytial virus (RSV) is the major viral cause of serious lower respiratory tract illness in infants and young children worldwide. RSV infection is limited to the superficial layers of the respiratory epithelium in immunocompetent individuals. Consistent with this in vivo observation, we and others have found that RSV buds preferentially from the apical surface of infected polarized epithelial cells. In contrast, directional budding is not observed in nonpolarized human epithelial cells. These findings suggest that RSV uses specific cellular trafficking pathways to accomplish viral replication. The host cell proteins that regulate directional budding of RSV are undefined. Apical sorting of cellular proteins in polarized epithelial cells involves the apical recycling endosome (ARE). To investigate whether ARE-mediated protein sorting plays a role during RSV replication, we expressed a fragment of the myosin Vb tail that functions as a dominant negative inhibitor of ARE-mediated protein sorting in polarized Madin-Darby canine kidney cells. When these cells were infected with RSV, a >9,000-fold reduction in viral yield was observed. A similar effect on virus replication was observed when a carboxyl-terminal fragment of another AREassociated protein, the Rab11 family interacting protein 1, was expressed in Madin-Darby canine kidney cells. These data suggest that RSV requires proper ARE-mediated protein sorting for efficient egress from the apical surface of polarized epithelial cells.

The Human Respiratory Syncytial Virus Matrix Protein Is Required for Maturation of Viral Filaments

“…Several cytoskeletal proteins, including actin isoforms, cofilin, and filamin-1, were detected by mass spectrometry analyses of purified viral filaments (although purity was difficult to assess due to the cell-associated nature of filaments). There is ample evidence of a role for polymerized actin and actinregulatory proteins in assembly (4,6,11,22,30,32,49,53,59). However, the role of polymerized actin seems to be limited to the actual budding/release event (11,30,32), which was shown previously to occur at sites of lipid rafts (8,9,22,27,31,36,42,49,50).…”

“…Several reports have implicated cytoskeletal elements in the processes that lead to virus egress, in particular microtubules, myosin V, actin, and actin-regulatory proteins such as profilin and RhoA (30,32,59). In addition, a role for an actin/myosinbased motility system in HRSV exit has been proposed (6,53,59), and there is strong evidence for the involvement of lipid rafts (8,9,22,27,31,36,49,50,53).…”

The histopathology of fatal untreated human respiratory syncytial virus infection

“…This may be related to the apical budding and entry of RSV from polarized epithelial cells. 16 In the alveoli, the dominant cell type infected was the type 1 pneumocyte. Although some cuboidal cells were immunostain-positive, it is difficult to know whether these represented terminally differentiated type 2 pneumocytes or reparative type 2 cells.…”

RAB11A Is Essential for Transport of the Influenza Virus Genome to the Plasma Membrane

“…The Rab11 pathway previously has been implicated in the budding of respiratory syncytial virus from the apical plasma membrane of infected cells (6,49). Moreover, Rab11 has been reported to colocalize with Sendai virus (SeV) vRNPs (11) although the requirement for Rab11 expression and function in their transport has not been explored.…”