Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic gamma-herpesvirus and an AIDS-associated pathogen. KSHV is associated with Kaposi’s sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman’s disease. KSHV-associated disease occurs more commonly in immunocompromised individuals, indicating an intricate relationship between disease progression and the host immune system. It is well established that cell intrinsic immune sensors, and in particular the DNA sensors Toll-like receptor 9 (TLR9) and cyclic GMP-AMP synthase (cGAS), sense KSHV infection. However, it is less clear whether RNA sensors, such as the RIG-I like receptors RIG-I and MDA5, participate in host defense against KSHV. Data within this proposal demonstrate that RIG-I and MDA5 impose a significant restriction on KSHV lytic reactivation in PEL. Using high- throughput transcriptomics we have defined the RNAs that are recognized by RIG-I and MDA5 during KSHV lytic infection in PEL, and this proposal builds upon our observations. These studies will enable us to (Aim 1) determine the contribution of RLR-associated RNAs to KSHV restriction, (Aim 2) determine how KSHV-encoded proteins impact RIG-I and MDA5 sensing, and (Aim 3) identify the precise mechanism of MDA5 activation during KSHV infection. Completion of these studies will provide fundamental knowledge regarding how the cell intrinsic immune response is activated during KSHV infection.