Characterising the role of the nsP3 macro domain in Chikungunya virus replication

Chikungunya virus (CHIKV) causes an acute fever with debilitating joint pain. Spread by the Aedes species of mosquito, recent increases in global temperature, and mutations in the viral glycoproteins have facilitated outbreaks worldwide with huge economic burden. Despite the recent resurgence of CHIKV, there are currently no vaccines or antiviral agents available.
CHIKV, an alphavirus, possesses a positive sense, single stranded RNA genome that encodes four non-structural proteins (nsPs). The CHIKV nsP3 possesses an N-terminal macro domain, a domain found in the proteins of all species, and are defined by their ability to bind ADP-ribose. It is unclear what role the nsP3 macro domain contributes to CHIKV replication.
Initially, a panel of cell lines was validated in terms of their physiological relevance and ability to support the replication of the CHIKV replicon and infectious virus. The phenotypes of a panel of mutants in the ADP-ribose binding pocket of the nsP3 macro domain were assessed in the context of a sub-genomic replicon and infectious virus in a range of relevant cell lines. Comparison of this data to the known biochemical properties of the nsP3 macro domain from the literature, indicated that ADP-ribose binding was crucial to CHIKV replication. In addition, this data suggested a role for the nsP3 macro domain in antagonising cellular innate immune pathways. ADP-ribose signalling has been implicated in the activation of the NFкB pathway therefore potential for the nsP3 macro domain to interfere with this cell signalling pathway was investigated. It was demonstrated that CHIKV did not activate the pathway and that expression of nsP3 actively was inhibitory. Furthermore, macro domain mutants with reduced ADP-ribose binding were unable to inhibit the pathway. It is therefore proposed that the CHIKV nsP3 macro domain is a virulence factor, able to suppress the host NFкB pathway to facilitate viral replication.