Serotonin Receptor Agonist 5-Nonyloxytryptamine Alters the Kinetics of Reovirus Cell Entry

Abstract

Mammalian orthoreoviruses (reoviruses) are nonenveloped double-stranded RNA viruses that infect most mammalian species, including humans. Reovirus binds to cell surface glycans, junctional adhesion molecule A (JAM-A), and the Nogo-1 receptor (depending on the cell type) and enters cells by receptor-mediated endocytosis. Within the endocytic compartment, reovirus undergoes stepwise disassembly, which is followed by release of the transcriptionally active viral core into the cytoplasm. In a small-molecule screen to identify host mediators of reovirus infection, we found that treatment of cells with 5-nonyloxytryptamine (5-NT), a prototype serotonin receptor agonist, diminished reovirus cytotoxicity. 5-NT also blocked reovirus infection. In contrast, treatment of cells with methiothepin mesylate, a serotonin antagonist, enhanced infection by reovirus. 5-NT did not alter cell surface expression of JAM-A or attachment of reovirus to cells. However, 5-NT altered the distribution of early endosomes with a concomitant impairment of reovirus transit to late endosomes and a delay in reovirus disassembly. Consistent with an inhibition of viral disassembly, 5-NT treatment did not alter infection by in vitro-generated infectious subvirion particles, which bind to JAM-A but bypass a requirement for proteolytic uncoating in endosomes to infect cells. We also found that treatment of cells with 5-NT decreased the infectivity of alphavirus chikungunya virus and coronavirus mouse hepatitis virus. These data suggest that serotonin receptor signaling influences cellular activities that regulate entry of diverse virus families and provides a new, potentially broad-spectrum target for antiviral drug development. IMPORTANCEIdentification of well-characterized small molecules that modulate viral infection can accelerate development of antiviral therapeutics while also providing new tools to increase our understanding of the cellular processes that underlie virus-mediated cell injury. We conducted a small-molecule screen to identify compounds capable of inhibiting cytotoxicity caused by reovirus, a prototype double-stranded RNA virus. We found that 5-nonyloxytryptamine (5-NT) impairs reovirus infection by altering viral transport during cell entry. Remarkably, 5-NT also inhibits infection by an alphavirus and a coronavirus. The antiviral properties of 5-NT suggest that serotonin receptor signaling is an important regulator of infection by diverse virus families and illuminate a potential new drug target. A s obligate intracellular pathogens, viruses exploit broadly conserved cellular processes to internalize, replicate, assemble, and release to infect adjacent cells and tissues. To counteract viral infection, hosts have evolved a variety of innate and adaptive immune mechanisms that antagonize these viral replication steps. Understanding host components that regulate the outcome of viral infection can uncover previously overlooked cellular pathways that might serve as new targets for antiviral therapeutics and enhance design of v...

Serotonergic drugs inhibit CHIKV infection at different stages of the cell entry pathway

“…The above data shows that 5-NT does not interfere with the initial stages of CHIKV cell entry which is in contrast to what has been described for reovirus (22). In this work the authors also used methiothepin mesylate (MM), which is a 5-HT receptor antagonist blocking 5-HT1/6/7 receptors (35,36) and showed that MM enhanced reovirus infectivity.…”

“…The effect of 5-NT on CHIKV infectivity was analyzed in human bone osteosarcoma epithelial U-2 OS cells as epithelial cells are natural targets during human CHIKV infection (29,30). Also, these cells were used by Mainou and co-workers who previously identified 5-NT as an inhibitor of CHIKV and reovirus infection (22). First, the mRNA expression levels of 10 distinct 5-HT receptor subtypes were determined in U-2 OS cells.…”

“…The 5-HT2 receptor family has been described to facilitate cell entry of JC polyomavirus (20) and 5-HT1 is suggested to be involved in HIV-1 replication (21). Furthermore, the infectivity of multiple RNA viruses were found to be controlled by 5-HT receptor agonists and antagonists (9,(22)(23)(24). Indeed, reovirus and CHIKV infectivity was found reduced in the presence of the 5-HT receptor agonist 5-nonyloxytryptamine (5-NT), which is described as a specific 5-HT1B/5-HT1D receptor agonist though it has also low level affinity to other 5-HT receptor families (22,25).…”

“…Furthermore, the infectivity of multiple RNA viruses were found to be controlled by 5-HT receptor agonists and antagonists (9,(22)(23)(24). Indeed, reovirus and CHIKV infectivity was found reduced in the presence of the 5-HT receptor agonist 5-nonyloxytryptamine (5-NT), which is described as a specific 5-HT1B/5-HT1D receptor agonist though it has also low level affinity to other 5-HT receptor families (22,25). 5-NT was shown to interfere with reovirus intracellular transport and disassembly kinetics during cell entry (22).…”

“…Indeed, reovirus and CHIKV infectivity was found reduced in the presence of the 5-HT receptor agonist 5-nonyloxytryptamine (5-NT), which is described as a specific 5-HT1B/5-HT1D receptor agonist though it has also low level affinity to other 5-HT receptor families (22,25). 5-NT was shown to interfere with reovirus intracellular transport and disassembly kinetics during cell entry (22). Opposed to the agonist, the 5-HT receptor antagonist methiothepin mesylate (MM) increased reovirus infectivity.…”