Extended Data Figure 9: Protein VII is necessary and sufficient for chromatin retention of HMGB1 in human and mouse cells.
From: A core viral protein binds host nucleosomes to sequester immune danger signals
a, b, Replication of Ad5-flox-VII virus on 293 or 293-Cre cells. Quantitative PCR analysis of viral genomic DNA over a time course of infection (a) shows the DBP gene is increasing exponentially in 293 and 293-Cre cells when infected with Ad5-flox-VII virus. In contrast, PCR for the protein VII gene (b) demonstrates deletion in 293-Cre cells (n = 2 biological replicates, mean ± s.d.). c, Salt fractionation of 293-Cre cells infected with wild-type Ad5, indicating that the Cre recombinase does not interfere with the ability of protein VII to retain HMGB1 in the high-salt chromatin fraction. Protein VII is also necessary for the chromatin retention of HMGB2. d, THP-1 cells transduced to express protein-VII–GFP results in chromatin distortion and HMGB1 retention in chromatin. Immunofluorescence of transduced PMA-treated THP-1 cells showing protein-VII–GFP (green), HMGB1 (red) and DNA (grey, blue in merge). e, Transduction to express protein-VII–GFP is sufficient to relocalize mouse HMGB1 in mouse embryonic fibroblast (MEF) cells. f, Salt fractionation of mouse embryonic fibroblast cells transduced to express protein-VII–GFP. Human Ad5 protein VII is sufficient to retain mouse HMGB1 in the high-salt fraction in MEF cells. The control vector expressing GFP alone does not have this effect.
