The M4 gene of gammaHV68 encodes a secreted glycoprotein and is required for the efficient establishment of splenic latency.

Sequence analysis of the murine gamma-herpesvirus 68 (gammaHV68) genome previously identified several open reading frames (ORFs) located at the left end of the viral genome that do not share homology with other known herpesvirus or cellular genes. Here, we show that one of these ORFs, M4, encodes a secreted glycoprotein that influences the establishment of splenic latency at early times post-infection. We generated a mutant virus containing a premature translation termination codon in the M4 ORF (M4.STOP), and demonstrated that this mutant virus replicates in vitro equivalent to wild type and marker rescue (M4.MR) viruses. M4.STOP was also capable of high-titer lytic replication in vivo, but at 16 days post-infection the establishment of latency in the spleen was significantly impaired. The defect in the establishment of splenic latency was apparent following either intranasal or intraperitoneal inoculation. In contrast, the M4.STOP mutant did not exhibit a defect in the establishment of latency in peritoneal cells. These results suggest that M4 mediates an extracellular host-pathogen interaction that impacts the establishment of latent infection in the spleen, but not the peritoneum.