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© The Rockefeller University Press, 0021-9525/1999//671 $5.00
The Journal of Cell Biology, Volume 147, Number 3, , 1999 671-682

A Novel Virus–Host Cell Membrane Interaction

: Membrane Voltage–Dependent Endocytic-like Entry of Bacteriophage 6 Nucleocapsid

Minna M. Poranen^a,b, Rimantas Daugelaviius^b,c, Päivi M. Ojala^a,b, Michael W. Hess^a, and Dennis H. Bamford^a,b

^a Institute of Biotechnology, FIN-00014, University of Helsinki, Helsinki, Finland
^b Department of Biosciences, FIN-00014, University of Helsinki, Helsinki, Finland
^c Department of Biochemistry and Biophysics, Vilnius University, LT-2009 Vilnius, Lithuania
Viikki Biocenter, P.O. Box 56 (Viikinkaari 5), FIN-00014, University of Helsinki, Finland.358-9-191 59098358-9-191 59100

gen_phag{at}cc.helsinki.fi

Studies on the virus–cell interactions have proven valuable in elucidating vital cellular processes. Interestingly, certain virus–host membrane interactions found in eukaryotic systems seem also to operate in prokaryotes (Bamford, D.H., M. Romantschuk, and P.J. Somerharju, 1987. EMBO (Eur. Mol. Biol. Organ.) J. 6:1467–1473; Romantschuk, M., V.M. Olkkonen, and D.H. Bamford. 1988. EMBO (Eur. Mol. Biol. Organ.) J. 7:1821–1829). 6 is an enveloped double-stranded RNA virus infecting a gram-negative bacterium. The viral entry is initiated by fusion between the virus membrane and host outer membrane, followed by delivery of the viral nucleocapsid (RNA polymerase complex covered with a protein shell) into the host cytosol via an endocytic-like route. In this study, we analyze the interaction of the nucleocapsid with the host plasma membrane and demonstrate a novel approach for dissecting the early events of the nucleocapsid entry process. The initial binding of the nucleocapsid to the plasma membrane is independent of membrane voltage () and the K⁺ and H⁺ gradients. However, the following internalization is dependent on plasma membrane voltage (), but does not require a high ATP level or K⁺ and H⁺ gradients. Moreover, the nucleocapsid shell protein, P8, is the viral component mediating the membrane–nucleocapsid interaction.

Key Words: endocytosis • cell energetics • dsRNA virus entry • prokaryote • phi6

© 1999 The Rockefeller University Press

1.used in this paper: , membrane voltage; p, proton motive force; pH, transmembrane pH gradient; AP, apyrase; CCCP, carbonyl cyanide m-chlorophenyl hydrazone; CF, chemical fixation; ds, dou-

ble-stranded; DCCD, N,N'-dicyclohexylcarbodiimide; FCCP, carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone; FS, freeze substitution; GD, gramicidin D; IC, infective center; MN, monensin; NC, nucleocapsid; NG, nigericin; OM, outer membrane; p.i., postinfection; PM, plasma membrane; PMB, polymyxin B; RF, rapid freezing; TPP+, tetraphenylphosphonium

P.M. Ojala's present address is Cell Cycle Laboratory, Haartman Institute, University of Helsinki, Finland.

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