Journal of Clinical Virology 37 Suppl. 1 (2006) S63–S68 www.elsevier.com/locate/jcv > Human herpesvirus 6B inhibits cell proliferation by a p53-independent pathway Bodil Øster, Maja D. Kaspersen, Emil Kofod-Olsen, Bettina Bundgaard, Per H¨ ollsberg* Institute of Medical Microbiology and Immunology, University of Aarhus, Aarhus, Denmark Abstract Background: Various forms of cellular stress can activate the tumour suppressor protein p53, an important regulator of cell cycle arrest, apoptosis, and cellular senescence. Cells infected by human herpesvirus 6B (HHV-6B) accumulate aberrant amounts of p53. Objectives: The aim of this study was to investigate the role of p53 accumulation in the HHV-6B-induced cell cycle arrest. Study design: The role of p53 was studied using the p53 inhibitor pifithrin-a, and cells genetically deficient in functional p53 by homologous recombination. Results: In response to HHV-6B infection, epithelial cells were arrested in the G1/S phase of the cell cycle concomitant with an aberrant accumulation of p53. However, the known p53-induced mediator of cell cycle arrest, p21, was not upregulated. Approximately 90% of the cells expressed HHV-6B p41, indicative of viral infection. The presence of pifithrin-a, a p53 inhibitor, did not reverse the HHV-6B-induced cell cycle block. In support of this, HHV-6B infection of p53 −/ − cells induced a cell cycle block before S-phase with kinetics similar to or faster than that observed by infection in wt cells. Conclusions: HHV-6B infection inhibited host cell proliferation concomitantly with p53 accumulation, but importantly the block in cell cycle occurred by a pathway independent of p53. © 2006 Elsevier B.V. All rights reserved. Keywords: HHV-6B; p53; Cell cycle; Epithelial cells 1. Introduction Human herpesvirus 6B (HHV-6B) differs in its clinical, biological and antigenic features from that of HHV-6A despite a 90% sequence identity (Dominguez et al., 1999; Isegawa et al., 1999). In the western world, HHV-6B infects most children before the age of two, an infection that is symptomatic in more than 90% of children giving rise to exanthem subitum (Braun et al., 1997; Zerr et al., 2005). Like other human herpesviruses, HHV-6B persists in the host in a latent or persistent state throughout life. During infections, HHV-6B interacts intimately with the host cell machinery. One significant consequence is the induction of cell cycle arrest (Black et al., 1992; Di Luca et al., 1990) at the G1/S and perhaps also the G2/M- phase (De Bolle et al., 2004; Øster et al., 2005; Dietrich et al., 2004), but the molecular mechanisms behind this phenomenon are not yet understood. The tumour suppressor protein p53 is a transcription factor involved in the control of G1/S and G2/M transitions of the cell cycle (Krause et al., 2000). As a master regulator, p53 upregulates or * Corresponding author. Tel.: +45 8942 1772; fax: +45 8619 6128. E-mail address: ph@microbiology.au.dk (P. H¨ ollsberg). represses the transcription of more than 100 different genes by sequence-specific binding to DNA response elements (Resnick et al., 2005). Normally p53 is kept at low levels, but when cells are stressed or damaged, p53 is activated and induces cell cycle arrest to allow repair of the genome or, in severe cases, provokes apoptosis of the cell. Similar to infections by HCMV and HSV-1, HHV-6A/B induces accumulation of p53 in the host cell. We have shown previously that the accumulated p53 protein during HHV-6B infection is capable of binding DNA, but unable to induce a corresponding upregulation in mRNA of p21, a p53 target gene involved in the execution of cell cycle arrest (Øster et al., 2005). De Bolle et al. (2004) have previously noted that the HHV-6A-induced G2 arrest in cord blood mononuclear cells occurs in the presence of p53 accumulation, but without induction of p21. However, this group only observed a block in G2/M and not in G1/S, and this proliferation block was much more pronounced in HHV-6A- than in HHV-6B-infected cells. To further address the mechanisms of HHV-6B-induced cell cycle arrest, we here examine the role of p53. 1590-8658/ $ – see front matter © 2006 Elsevier B.V. All rights reserved.