Caspase-mediated cleavage of Murine Norovirus NS1/2 potentiates apoptosis and is required for persistent infection

Abstract

Human norovirus (HuNoV) is the leading cause of acute gastroenteritis and is spread by fecal shedding that can often persist for weeks to months after infection. Murine norovirus (MNV) is also shed persistently in the feces and provides a tractable model to study molecular mechanisms of enteric persistence. Previous studies have identified non-structural protein 1 (NS1) from the persistent MNV strain CR6 as critical for persistent infection in intestinal epithelial cells (IECs), but its mechanism of action remains unclear. We now find that the function of NS1 in promoting persistence is regulated by apoptosis. Following induction of apoptosis in infected cells, a minority of NS1 is cleaved from the precursor NS1/2 protein, and this cleavage is prevented by mutation of caspase target motifs. MNV strain CR6 with these mutations (CR6{triangleup}casp) is profoundly compromised in infection of IECs and persistence in the intestine. Conversely, replication in tissues outside of the intestine, or in a cultured macrophage cell line, is unchanged, indicating that the requirement of NS1/2 cleavage is intestine-specific. Intriguingly, we also find that cleavage of CR6 NS1/2 potentiates apoptosis, suggesting that regulation of cell death is a novel function of this viral protein. Together, these data indicate that the ability of NS1 to promote MNV persistence in IECs is regulated by host caspases, and suggest that potentiation of apoptosis plays a role in viral tropism in the intestine.\n\nAuthor SummaryHuman Norovirus infection is highly contagious and the most common cause of acute gastroenteritis. Norovirus can persist and be shed for months after infection, leading to continued outbreaks. There are many unanswered questions as to host and viral components of norovirus pathogenesis that can be addressed within the murine norovirus (MNV) model system. We previously identified a critical role for a viral protein, NS1, for intestinal persistence. Herein we describe how the regulation of NS1 is critical for persistent infection in intestinal epithelial cells, but is not required for acute infection of non-epithelial cells, or infection of tissues outside of the gut. Additionally, we demonstrate that NS1 is both regulated by the host cell death machinery, and also reciprocally regulates that machinery to promote cell death during MNV infection, and found that this is specific to persistent strain of MNV. Altogether these data identify a role for how NS1 in a new pathway involved in establishing a persistent norovirus infection in the intestine.