KSHV Terminal Repeat Regulates Inducible Lytic Gene Promoters

Summary The Kaposi’s sarcoma-associated herpesvirus (KSHV) genome consists of an approximately 140 kb unique coding region flanked by multiple copies of 0.8 kb terminal repeat (TR) sequence. While TR’s function in plasmid maintenance is well-established, TR’s transcription regulatory roles have not been fully explored. Here, we show KSHV TR is a large transcription regulatory domain. A series of Cleavage Under Targets & Release Using Nuclease demonstrated that TR fragments are occupied by histone modifying enzymes that are known to interact with LANA in naturally infected cells, and the TR possessed characteristic enhancer histone modifications. The H3K4me3 and H3K27Ac modifications were conserved in unique region of the KSHV genome among naturally infected cells, and the KSHV Origin of lytic replication (Ori-Lyt) showed similar protein and histone modification occupancies with TR’s. In the Ori-Lyt region, the LANA complex colocalizes with H3K27Ac-modified nucleosome along with paused RNA polymerase II, and two K-Rta recruitment sites frank the nucleosome. The isolated reporter assays demonstrated that neighboring TR fragments enhanced viral lytic gene promoter activity independent of orientation in KSHV-infected and non-infected 293FT cells. K-Rta transactivation function was drastically enhanced with TR, while LANA acquired promoter repression function with TR. KSHV TR is, therefore a regulatory domain for KSHV inducible genes. However, in contrast to cellular enhancers that are bound by multiple transcription factors, perhaps the KSHV enhancer is predominantly regulated by the LANA nuclear body with TR. KSHV evolved a clever mechanism to tightly control the latency-lytic switch with the TR/LANA complex. Importance Enhancers are a crucial regulator of differential gene expression programs. Enhancer is the cis-regulatory sequences that determine target genes’ spatiotemporal and quantitative expression. Here, we show that KSHV terminal repeats fulfill the enhancer definition for KSHV inducible gene promoters. KSHV enhancer is occupied by LANA and its interacting proteins, such as CHD4, and CHD4 is known to restrict enhancers to access promoters for activation. This study thus proposes a new latency-lytic switch model in which TR accessibility to the KSHV gene promoters regulates lytic gene transcription.