Figure 6
From: Role of SUMO-1 and SUMO interacting motifs in rhesus TRIM5α-mediated restriction
The SIM1 and SIM2 mutations disrupt rhTRIM5α localization to nuclear bodies containing PML/TRIM19 andSUMO-1. A. HeLa cells stably expressing wild typeYFP-rhTRIM5α or SIM mutants were treated with LMB for 4 hours. Following treatment the cells were fixed and imaged. Z-stack images were collected with a DeltaVision microscope equipped with a digital camera using a 1.4-numerical aperture (NA) 100× objective lens, and were deconvolved with SoftWoRx deconvolution software. Individual channel images were superimposed to create the merged panels. B. HeLa cells expressing wild type YFP-rhTRIM5a or SIM mutants were treated with LMB for 4hours. Following treatment the cells were fixed and stained with an anti-SUMO-1 antibody. Z-stack images were collected as described in A. Deconvolved images were analyzed for YFP-rhTRIM5a maximum fluorescence intensity (MFI) in SUMO-1nuclear bodies by the use of the Surface Finder function in the Imaris software(Bitplane). For each SUMO-1 puncta, the MFI of YFP-rhTRIM5a mutants was determined and the data was plotted in GraphPad Prism 5® software.