The contribution of host shutoff towards the gammaherpesviral lifecycle is unknown, and efforts to delete muSOX from MHV68 yielded virus unable to replicate, presumably due to an essential role for the DNase activity [16] . Previous studies found the DNase and host shutoff functions of SOX and BGLF5 to be genetically separable [25, 26] , making it possible to evaluate the role of each function of the protein in isolation. We report herein the identification of a functionally similar MHV68 muSOX point mutant that renders the virus selectively defective for host shutoff, enabling the analysis of the role of this phenotype during infection of cultured cells and in vivo. The mutant virus exhibited little to no defect during multi-step growth curves in tissue culture or during acute replication in the mouse lung following intranasal inoculation. Unexpectedly, the virus accumulated to significantly reduced levels in the lymph nodes at 10 days post infection and was highly attenuated at the downstream stage of latency establishment. Once trafficking was bypassed via an intraperitoneal infection, the host shutoff defective virus still was attenuated in establishing latency in the spleen. Our results identify for the first time a key role for the lytic mRNA turnover activity in establishing viral latency, emphasizing the important interplay between these seemingly disparate stages of the viral lifecycle.