Site-specific association with host and viral chromatin by Kaposi's sarcoma-associated herpesvirus LANA and its reversal during lytic reactivation.
Latency-associated nuclear antigen (LANA), a multifunctional protein expressed by the Kaposi sarcoma-associated herpesvirus (KSHV) in latently infected cells, is required for stable maintenance of the viral episome. This is mediated by two interactions: LANA binds to specific sequences (LBS1 and LBS2) on viral DNA and also engages host histones, tethering the viral genome to host chromosomes in mitosis. LANA has also been suggested to affect host gene expression, but both the mechanism(s) and role of this dysregulation in KSHV biology remain unclear. Here, we have examined LANA interactions with host chromatin on a genome-wide scale using chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) and show that LANA predominantly targets human genes near their transcriptional start sites (TSSs). These host LANA-binding sites are generally found within transcriptionally active promoters and display striking overrepresentation of a consensus DNA sequence virtually identical to the LANA-binding site 1 (LBS1) motif in KSHV DNA. Comparison of the ChIP-seq profile with whole-transcriptome (high-throughput sequencing of RNA transcripts [RNA-seq]) data reveals that few of the genes that are differentially regulated in latent infection are occupied by LANA at their promoters. This suggests that direct LANA binding to promoters is not the prime determinant of altered host transcription in KSHV-infected cells. Most surprisingly, the association of LANA to both host and viral DNA is strongly disrupted during the lytic cycle of KSHV. This disruption can be prevented by the inhibition of viral DNA synthesis, suggesting the existence of novel and potent regulatory mechanisms linked to either viral DNA replication or late gene expression.


OBJECTIVE
Here, we employ complementary genome-wide analyses to evaluate the distribution of the highly abundant latency-associated nuclear antigen, LANA, on the host genome and its impact on host gene expression during KSHV latent infection. Combined, ChIP-seq and RNA-seq reveal that LANA accumulates at active gene promoters that harbor specific short DNA sequences that are highly reminiscent of its cognate binding sites in the virus genome. Unexpectedly, we found that such association does not lead to remodeling of global host transcription during latency. We also report for the first time that LANA's ability to bind host and viral chromatin is highly dynamic and is disrupted in cells undergoing an extensive lytic reactivation. This therefore suggests that the association of LANA to chromatin during a productive infection cycle is controlled by a new regulatory mechanism.
Novartis Institutes for BioMedical Research, Department of Infectious Diseases, Emeryville, California, USA alexandre.mercier@novartis.com.
Alexandre
A
Mercier
Novartis Institutes for BioMedical Research, Department of Infectious Diseases, Emeryville, California, USA.
Carolina
C
Arias
Novartis Institutes for BioMedical Research, Department of Infectious Diseases, Emeryville, California, USA.
Alexis S
AS
Madrid
Novartis Institutes for BioMedical Research, Department of Infectious Diseases, Emeryville, California, USA.
Meghan M
MM
Holdorf
Novartis Institutes for BioMedical Research, Department of Infectious Diseases, Emeryville, California, USA.
Don
D
Ganem
D000956:Antigens, Viral / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D015967:Gene Expression Regulation, Viral; D055106:Genome-Wide Association Study; D019288:Herpesvirus 8, Human / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012514:Sarcoma, Kaposi / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D017735:Virus Latency
D000956:Antigens, Viral; D002843:Chromatin; D009687:Nuclear Proteins; D014764:Viral Proteins; C109540:latency-associated nuclear antigen