TY - JOUR
T1 - Intercellular Extensions Are Induced by the Alphavirus Structural Proteins and Mediate Virus Transmission
AU - Martinez, Maria Guadalupe
AU - Kielian, Margaret
N1 - Publisher Copyright:
© 2016 Martinez, Kielian.
PY - 2016/12/15
Y1 - 2016/12/15
N2 - Alphaviruses are highly organized enveloped RNA viruses with an internal nucleocapsid surrounded by a membrane containing the E2 and E1 transmembrane proteins. Alphavirus budding takes place at the plasma membrane and requires the interaction of the cytoplasmic domain of E2 with the capsid protein. Here we used WT alphaviruses and Sindbis virus in which E2 was fused to a fluorescent protein to characterize virus exit from host cells. Our results show that alphavirus infection induced striking modifications of the host cell cytoskeleton and resulted in the formation of stable intercellular extensions that emanated exclusively from the infected cell. The intercellular extensions were long (> 10 μM), contained actin and tubulin, and formed flattened contacts with neighboring cells, but did not mediate membrane or cytoplasmic continuity between cells. Receptor down-regulation studies indicated that formation of stable extensions did not require the virus receptor, and that extensions promoted cell-to-cell virus transmission to receptor-depleted cells. Virus mutant experiments demonstrated that formation of extensions required the E2-capsid interaction but not active particle budding, while intercellular transmission of infection required the production of fusion-active virus particles. Protein expression studies showed that even in the absence of virus infection, the viral structural proteins alone induced intercellular extensions, and that these extensions were preferentially targeted to non-expressing cells. Together, our results identify a mechanism for alphavirus cell-to-cell transmission and define the key viral protein interactions that it requires.
AB - Alphaviruses are highly organized enveloped RNA viruses with an internal nucleocapsid surrounded by a membrane containing the E2 and E1 transmembrane proteins. Alphavirus budding takes place at the plasma membrane and requires the interaction of the cytoplasmic domain of E2 with the capsid protein. Here we used WT alphaviruses and Sindbis virus in which E2 was fused to a fluorescent protein to characterize virus exit from host cells. Our results show that alphavirus infection induced striking modifications of the host cell cytoskeleton and resulted in the formation of stable intercellular extensions that emanated exclusively from the infected cell. The intercellular extensions were long (> 10 μM), contained actin and tubulin, and formed flattened contacts with neighboring cells, but did not mediate membrane or cytoplasmic continuity between cells. Receptor down-regulation studies indicated that formation of stable extensions did not require the virus receptor, and that extensions promoted cell-to-cell virus transmission to receptor-depleted cells. Virus mutant experiments demonstrated that formation of extensions required the E2-capsid interaction but not active particle budding, while intercellular transmission of infection required the production of fusion-active virus particles. Protein expression studies showed that even in the absence of virus infection, the viral structural proteins alone induced intercellular extensions, and that these extensions were preferentially targeted to non-expressing cells. Together, our results identify a mechanism for alphavirus cell-to-cell transmission and define the key viral protein interactions that it requires.
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U2 - 10.1371/journal.ppat.1006061
DO - 10.1371/journal.ppat.1006061
M3 - Article
C2 - 27977778
AN - SCOPUS:85007543330
SN - 1553-7366
VL - 12
JO - PLoS pathogens
JF - PLoS pathogens
IS - 12
M1 - e1006061
ER -