Downloaded from www.microbiologyresearch.org by IP: 54.87.0.57 On: Mon, 12 Jun 2017 23:41:45 Kaposi’s sarcoma-associated herpesvirus K8 protein interacts with hSNF5 Seungmin Hwang, 1 Daeyoup Lee, 1 Yousang Gwack, 1 Hyesun Min 2 and Joonho Choe 1 Correspondence Joonho Choe jchoe@mail.kaist.ac.kr 1 Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea 2 Department of Food and Nutrition, Hannam University, Daejeon 306-791, Korea Received 9 July 2002 Accepted 13 November 2002 Kaposi’s sarcoma-associated herpesvirus (KSHV) is a human gammaherpesvirus related to Epstein–Barr virus (EBV) and herpesvirus saimiri. KSHV open reading frame K8 encodes a basic region-leucine zipper protein of 237 aa that homodimerizes. K8 shows significant similarity to the EBV immediate-early protein Zta, a key regulator of EBV reactivation and replication. In this study, a carboxyl-terminal deletion mutant of K8, K8(1–115), that had strong transactivating properties was found. Screening using transcriptionally inactive K8(1–75) showed that K8 interacts and co-localizes with hSNF5, a cellular chromatin-remodelling factor, both in vivo and in vitro. This interaction requires aa 48–183 of hSNF5 and 1–75 of K8. In a yeast expression system, the ability of K8 and K8(1–115) to activate transcription requires the presence of SNF5, the yeast homologue of hSNF5. These data suggest a mechanism by which the SWI–SNF complex is recruited to specific genes. They also suggest that K8 functions as a transcriptional activator under specific conditions and that its transactivation activity requires its interaction with the cellular chromatin remodelling factor hSNF5. INTRODUCTION Kaposi’s sarcoma-associated herpesvirus (KSHV), or human herpesvirus-8 (HHV-8), is a gammaherpesvirus related to Epstein–Barr virus (EBV) and herpesvirus saimiri (Boshoff & Weiss, 1998; Russo et al., 1996; Schulz et al., 1998). KSHV is implicated in the pathogenesis of Kaposi’s sarcoma (KS), primary effusion B cell lymphomas and multicentric Castleman’s disease. Like EBV, KSHV infection in tumour tissue or lymphoma-derived cell lines is predominantly latent. The KSHV genome is present in a latent state in B cell lines, such as body cavity-based lymphoma cells (BCBL-1). These cells have multiple copies of circularized KSHV DNA maintained in the nucleus as episomes whose expression is highly attenuated (Boshoff & Weiss, 1998; Cesarman et al., 1995; Soulier et al., 1995). The most important step in the KSHV life cycle may be the switch from latency to lytic replication; virus lytic replica- tion is critical for the development of KS (Boshoff et al., 1995; Miller et al., 1997; Schulz et al., 1998). In the latent phase, expression of a limited number of viral genes occurs and the genome is maintained in an episomal form. Lytic phase is characterized by a cascade of gene expression resulting in productive release of virions from infected cells. Lytic reactivation of KSHV can be induced artificially by phorbol esters such as 12-O-tetradecanoylphorbol-13- acetate (TPA) or n-butyrate, in a manner similar to EBV- infected B cell lines (Renne et al., 1996). Upon chemical induction, KSHV produces immediate-early viral trans- cripts within 4 h. These transcripts encode viral transcrip- tional activator proteins, such as open reading frame 50 (ORF 50). From its expression pattern and function, ORF 50 appears to be the principal driver of the lytic cascade and functions as a switch gene in the disruption of latency (Lukac et al., 1998; Sun et al., 1998; Zhu et al., 1999). The KSHV ORF K8 gene encodes an early lytic protein that is activated by, and expressed after, KSHV ORF 50 protein (Lin et al., 1999; Sun et al., 1999). Three forms of K8 protein result from alternative splicing and usage of different stop codons. The major form of K8 is a protein of 237 aa with a prototypic basic region-leucine zipper (bZIP) domain at the carboxyl terminus and it homodimerizes using this domain. An acidic domain between residues 6 and 47 (Zhu et al., 1999) suggests that K8 (K-bZIP) may be a member of the bZIP family of transcription factors. This hypothesis is supported further by amino acid sequence analysis showing significant similarity between K8 and the EBV immediate- early gene product Zta (BZLF1, EB1, Z), a transactivator responsible for EBV replication and reactivation from latency to the lytic life cycle (Gruffat et al., 1999; Lin et al., 1999; Zhu et al., 1999). Published ahead of print on 25 November 2002 as DOI 10.1099/ vir.0.18699-0. 0001-8699 G 2003 SGM Printed in Great Britain 665 Journal of General Virology (2003), 84, 665–676 DOI 10.1099/vir.0.18699-0