Mouse Schlafen Family Proteins Control MHV68 Replication in Fibroblasts

Schlafen proteins are interferon-responsive regulators of cellular and antiviral pathways that can profoundly influence viral replication and host defense. Diverse viral infections are known to induce expression of Schlafen family members in human cells, while infection studies in mice have been primarily limited to mSlfn2, a group I Schlafen protein without a clear ortholog in humans. RNA-seq analysis of three distinct infection models reveals that mouse Schlafens are induced in response to murine gammaherpesvirus 68 (MHV68) in fibroblasts, B cells, and primary peritoneal macrophages. Notably, mSlfn8 expression is induced under multistep low-MOI infection conditions, whereas infections initiated with higher viral inoculum fail to induce mSlfn8. To assess whether mSlfn8 or related Slfns impact MHV68 replication, we performed siRNA-mediated knockdown and CRISPR/Cas9 knockout of mSlfn2, mSlfn8, and mSlfn9 in NIH 3T3 cells. Two independent clonal mSlfn2 and mSlfn8 knockout cell lines supported higher viral titer, indicating that these mSlfns are restriction factors for MHV68 replication in murine fibroblasts. Collectively, these results demonstrate that mSlfn2 and mSlfn8 function as antiviral effectors during MHV68 infection and support a model in which MHV68 counteracts interferon-induced mouse Schlafen expression to promote efficient replication.