Accumulation of Heterochromatin Components on the Terminal Repeat Sequence of Kaposi's Sarcoma-Associated Herpesvirus Mediated by the Latency-Associated Nuclear Antigen

Abstract

In the latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), its 160-kb circularized episomal DNA is replicated and maintained in the host nucleus. KSHV latency-associated nuclear antigen (LANA) is a key factor for maintaining viral latency. LANA binds to the terminal repeat (TR) DNA of the viral genome, leading to its localization to specific dot structures in the nucleus. In such an infected cell, the expression of the viral genes is restricted by a mechanism that is still unclear. Here, we found that LANA interacts with SUV39H1 histone methyltransferase, a key component of heterochromatin formation, as determined by use of a DNA pull-down assay with a biotinylated DNA fragment that contained a LANA-specific binding sequence and a maltose-binding protein pull-down assay. The diffuse localization of LANA on the chromosomes of uninfected cells changed to a punctate one with the introduction of a bacterial artificial chromosome containing most of the TR region, and SUV39H1 clearly colocalized with the LANA-associated dots. Thus, the LANA foci in KSHV-infected cells seemed to include SUV39H1 as well as heterochromatin protein 1. Furthermore, a chromatin immunoprecipitation assay revealed that the TR and the open reading frame (ORF) K1 and ORF50/ RTA genes, but not the ORF73/LANA gene, lay within the heterochromatin during KSHV latency. Taken together, these observations indicate that LANA recruits heterochromatin components to the viral genome, which may lead to the establishment of viral latency and govern the transcription program.Kaposi's sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8, was discovered in Kaposi's sarcoma (KS) lesions (8) and is strongly associated with multicentric Castleman's disease and primary effusion lymphoma (PEL), which are found predominantly in AIDS patients (6). KSHV belongs to the gammaherpesviruses, which have two distinct replication systems, lytic and latent. For lytic replication, the immediate-early gene product RTA (31), which functions as a strong transactivator that upregulates many viral genes, must be expressed. In PEL cells, KSHV is mainly in the latent state, during which RTA is not expressed (18). Thus, it is likely that KSHV has regulatory machinery that enables it to maintain the latent phase as a default state.The latency-associated nuclear antigen (LANA) is the 1,162-amino-acid (aa) product of the KSHV open reading frame 73 (ORF73) gene, which is expressed during latency (20, 37). LANA has an acidic amino acid repeat region in the middle and a DNA-binding/dimerization domain at its C terminus (42). During latency, LANA binds a DNA sequence in the terminal repeat (TR) region of the viral genome, which consists of repeated sequences made up of 801-bp units and contains an origin of replication (OriP) (2, 3, 13). Most cell lines derived from PELs carry a viral genome with a long TR sequence of more than 20 kb (17). Given LANA's functional similarity to EBNA1, which is encoded by another gammaherpesvirus, Epstein-Barr virus, it is lik...

Histone Modification Pattern of the T-Cellular Herpesvirus Saimiri Genome in Latency

“…Tandem repeats have long been known to initiate heterochromatin formation also by involving the small interfering RNA machinery (20,34). Evidence for heterochromatinization of herpesviral terminal repeats has been provided by Sakakibara et al for the KSHV genome (48). These researchers found that KSHV LANA, when bound to the terminal repeats, interacts with HP1 and also with the histone H3 K9 specific methyltransferase SUV39H1.…”

Viral epigenomes in human tumorigenesis

“…LANA is unmethylated (Chen et al, 2001) and is known to be associated with histone modifications and subsequent gene expression alterations in host and viral genes that contribute to viral oncogenesis (Sakakibara et al, 2004;Lu et al, 2006;Stuber et al, 2007). It has also been shown that the recruitment of a de novo methyltransferase (DNMT3b) causes downregulation of genes such as cadherin 13 and TGF-b by promoter methylation (Shamay et al, 2006;Di Bartolo et al, 2008).…”

The Kaposi's Sarcoma-Associated Herpesvirus LANA Protein Stabilizes and Activates c-Myc

“…Targeting of LANA to DNA either through the use of Gal4-LANA fusion proteins (38,53) through binding of LANA to the KSHV terminal repeats (25) or through LANA recruitment to cell (57) or viral promoters (39,42) leads to transcriptional repression. LANA binds to histone deacetylase-associated corepressors (38) and is also capable of recruiting de novo DNA methyltransferases and the histone methyl transferase SUV39H1 to downregulate targeted cell promoters through CpG methylation (50,57).…”