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ncb1544 - LETTERS Retroviruses can establish filopodial...

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LETTERS 310 NATURE CELL BIOLOGY VOLUME 9 | NUMBER 3 | MARCH 2007 Retroviruses can establish filopodial bridges for efficient cell-to-cell transmission Nathan M. Sherer 1,3 , Maik J. Lehmann 1,4 , Luisa F. Jimenez-Soto 1,5 , Christina Horensavitz 2 , Marc Pypaert 2 and Walther Mothes 1,6 The spread of retroviruses between cells is estimated to be 2–3 orders of magnitude more efficient when cells can physically interact with each other 1,2 . The underlying mechanism is largely unknown, but transfer is believed to occur through large-surface interfaces, called virological or infectious synapses 3–6 . Here, we report the direct visualization of cell-to-cell transmission of retroviruses in living cells. Our results reveal a mechanism of virus transport from infected to non-infected cells, involving thin filopodial bridges. These filopodia originate from non- infected cells and interact, through their tips, with infected cells. A strong association of the viral envelope glycoprotein (Env) in an infected cell with the receptor molecules in a target cell generates a stable bridge. Viruses then move along the outer surface of the filopodial bridge toward the target cell. Our data suggest that retroviruses spread by exploiting an inherent ability of filopodia to transport ligands from cell to cell. To study the spread of retroviruses between living cells, we used the murine leukemia virus (MLV) as a model. MLV was fluorescently labelled in infected cells by expressing a CFP-fusion with the capsid protein Gag (MLV Gag–CFP), as well as an envelope protein (Env) carrying a YFP- insertion (MLV Env–YFP) 7,8 . Infected cells were then cocultured with non-infected target cells expressing a CFP fusion with the MLV receptor mCAT1 (mCAT1–CFP) 8 . Infected cells were readily identified by the presence of retroviral particles, observed as punctae displaying both YFP and CFP fluorescence (Fig. 1A). Receptor-expressing target cells were characterized by homogeneous CFP fluorescence at the plasma mem- brane. Strikingly, essentially all virus particles moving from infected to target cells migrated along thin, elongated filopodia (Fig. 1A and see Supplementary Information, Movie 1). The particles moved unidirec- tionally at an average rate of 0.7 μm min –1 ( n = 117) and required, on average, approximately 18 min to move from one cell to the other (Fig. 1B, C). Identical observations were made when target cells were labelled with mCAT1–YFP (see Supplementary Information, Movie 2). Filopodial bridges were only observed between infected and non- infected cells. They averaged 5.8 μm in length ( n = 59) and were long lived (Fig. 1D, E; observed up to the maximum imaging time of 4 h). In contrast, normal filopodia of target cells that did not connect with an infected cell were significantly shorter (average length, 2.37 μm; n = 60) and highly dynamic, rapidly undergoing cycles of growth and retraction (Fig. 1D, E). To visualize moving viral particles at higher resolution, cells were cocultured on a coverslip with a lettered grid, and a filopodial
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This note was uploaded on 11/01/2010 for the course PRC 1234 taught by Professor All during the Spring '10 term at HKU.

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ncb1544 - LETTERS Retroviruses can establish filopodial...

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