The anti-viral protein Shiftless blocks p-body formation during KSHV infection.

Processing bodies (P-bodies/PBs) are non-membranous foci involved in coordinating RNA fate by regulating translation and mRNA decay. In this study, we characterize the anti-viral factor Shiftless (SHFL) as a potent disruptor of PB dynamics. We show SHFL expression restricts PB accumulation even in the context of oxidative stress, suggesting that SHFL expression impedes PB formation. Mutational approaches revealed that SHFL RNA-binding activity is not required to restrict PB formation. However, we have identified a new region of SHFL, a bridge between two distant SHFL domains, as necessary for SHFL-mediated PB disruption. Furthermore, we show that SHFL's ability to disrupt PB formation also impacts its anti-viral activity during infection by the gammaherpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV). While WT SHFL efficiently restricts KSHV lytic reactivation, SHFL mutants defective in PB disruption no longer restrict KSHV reactivation. SHFL-mediated PB disruption also leads to increased expression of several anti-viral cytokines, further emphasizing the connection among SHFL, PB dynamics and the SHFL-dependent anti-viral state. Taken together, our observations suggest a role of SHFL in inhibiting PB formation to restrict KSHV lytic replication, reinforcing the importance of crosstalk between RNA fate and the innate immune response to viral infection.