Author: van der Schaar, H. M.; Melia, C. E.; van Bruggen, J. A. C.; Strating, J. R. P. M.; van Geenen, M. E. D.; Koster, A. J.; Bárcena, M.; van Kuppeveld, F. J. M.
Title: Illuminating the Sites of Enterovirus Replication in Living Cells by Using a Split-GFP-Tagged Viral Protein Document date: 2016_7_6
ID: 1aptufp6_34
Snippet: Enteroviruses can infect both polarized and nonpolarized cells. The use of CVB3 encoding split-GFP-tagged 3A in distinct cell types relies on the cellular expression of GFP(S1-10). As an alternative to delivering the GFP(S1-10) gene via transduction, we introduced the coding sequence of GFP(S1-10) into the viral genome together with 3A(S11aa2). This new recombinant CVB3 encoding both GFP fragments also induced GFP fluorescent foci that colocalize.....
Document: Enteroviruses can infect both polarized and nonpolarized cells. The use of CVB3 encoding split-GFP-tagged 3A in distinct cell types relies on the cellular expression of GFP(S1-10). As an alternative to delivering the GFP(S1-10) gene via transduction, we introduced the coding sequence of GFP(S1-10) into the viral genome together with 3A(S11aa2). This new recombinant CVB3 encoding both GFP fragments also induced GFP fluorescent foci that colocalized with 3A. While progeny virion production by this virus was delayed, the viral RNA levels were very similar to the levels in CVB3-3A(S11aa2) infection of BGM(GFPS1-10) cells, implying that the "two-fragment virus" retained its ability to form ROs. Hence, this virus would be suitable for studying enterovirus ROs in physiologically more relevant cell types, including pancreatic cells (14) and 3D-cultured CaCo-2 cells (63), without the need for ectopic expression of GFP(S1-10).
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