Abstract
All herpesviruses share a remarkable propensity to establish latent infection. Human Kaposi's sarcoma-associated herpesvirus (KSHV) effectively enters latency after de novo infection, suggesting that KSHV has evolved with strategies to facilitate latent infection. NF-κB activation is imperative for latent infection of gammaherpesviruses. However, how NF-κB is activated during de novo herpesvirus infection is not fully understood. Here, we report that KSHV infection activates the inhibitor of κB kinase β (IKKβ) and the IKK-related kinase epsilon (IKKε) to enable host NF-κB activation and KSHV latent infection. Specifically, KSHV infection activated IKKβ and IKKε that were crucial for latent infection. Knockdown of IKKβ and IKKε caused aberrant lytic gene expression and impaired KSHV latent infection. Biochemical and genetic experiments identified RelA as a key player downstream of IKKβ and IKKε. Remarkably, IKKβ and IKKε were essential for phosphorylation of S(536) and S(468) of RelA, respectively. Phosphorylation of RelA S(536) was required for phosphorylation of S(468), which activated NF-κB and promoted KSHV latent infection. Expression of the phosphorylation-resistant RelA S(536)A increased KSHV lytic gene expression and impaired latent infection. Our findings uncover a scheme wherein NF-κB activation is coordinated by IKKβ and IKKε, which sequentially phosphorylate RelA in a site-specific manner to enable latent infection after KSHV de novo infection.