FIG 5.

Nuclear translocation of PABPC and XBP1 exon skipping in parental RF and RRV and gene 7 monoreassortants. (A) Localization of PABPC in rotavirus-infected cells. MA104 cells infected (or mock infected) with bovine RF, rhesus RRV, or gene 7 monoreassortants for 9 h were fixed and incubated with NSP2- and PABPC-specific antibodies. Secondary antibodies coupled to the Alexa fluorophore stains NSP2 (red) and PABPC (green). Nuclei were stained blue with DAPI. (B) Quantification of nuclear PABC1 in rotavirus-infected cells. Images such as those shown in panel A were taken at 3, 6, and 9 h after infection (hpi) with the indicated virus and analyzed. The ratio of nuclear to total green (PABPC) fluorescence (corrected for background) is reported. The results are mean values ± the SEM of three fields with >50 cells. (C) Kinetics of XBP1 splicing. The XBP1 DNA products obtained by RT-PCR of RNA extracted from mock-infected cells or from cells infected (MOI of 10) for 3, 6, and 9 h with the indicated parental or monoreassortant virus were analyzed by agarose gel electrophoresis, together with the GAPDH PCR control. The sizes of the molecular weight markers (MW) are indicated in base pairs on the left.