PDTC inhibits picornavirus polyprotein processing and RNA replication by transporting zinc ions into cells

Abstract

Previously, it was shown that pyrrolidine dithiocarbamate (PDTC) inhibits proteolytic polyprotein processing and replication of human rhinovirus by transporting metal ions into cells. Here, it is shown that PDTC also inhibits replication of two other picornaviruses: coxsackievirus B3 (CVB3), a closely related virus that belongs to the genus Enterovirus, and mengovirus, an encephalomyocarditis virus strain that belongs to the genus Cardiovirus, and that this inhibition is due to the dithiocarbamate moiety of the compound. Making use of subgenomic replicons, evidence is provided that PDTC inhibits replication of these two viruses by disturbing viral RNA synthesis. Furthermore, it is shown that PDTC transports zinc ions into cells and that these zinc ions play an important role in the antiviral activity mediated by PDTC. Finally, it is shown that PDTC interferes with proteolytic processing of the polyproteins of both CVB3 and mengovirus, but that the underlying mechanism between these two viruses differs. In CVB3-infected cells, PDTC interferes strongly with the proteolytic activity of 3CD pro , as shown by the impaired production of the mature capsid proteins as well as the autocleavage of 3CD pro into 3C pro and 3D pol . In mengovirus-infected cells, however, PDTC had no effect on the proteolytic production of capsid proteins or the autocleavage of 3CD pro . Instead, PDTC caused the accumulation of a high-molecular-mass precursor protein, due to an impairment in the primary 'break' that normally occurs at the 2A-2B junction. Thus, PDTC disturbs polyprotein processing and replication of two groups of picornaviruses, enteroviruses and cardioviruses, but the underlying mechanism is different.

Antiviral Activity of the Zinc Ionophores Pyrithione and Hinokitiol against Picornavirus Infections

“…Five hours p.i., cells were starved half an hour prior to 1 h of labeling with [ 35 S]Met/Cys. After labeling, cells were lysed, and cellular extracts were subjected to immunoprecipitation using either antibodies specific for 3D or 3C (kindly provided by C. Cameron, Pennsylvania State University) as described previously (22). Then, the protein pattern was analyzed by SDS-PAGE and autoradiography.…”

“…In particular, we have demonstrated that PDTC has negative effects on picornavirus replication by influencing the processing of the viral polyprotein (21,22).…”

“…We have previously described the commonly used NF-B inhibitor and metal ion chelator pyrrolidine-dithiocarbamate (PDTC) to significantly inhibit the replication of several picornaviruses such as human rhinovirus (HRV), poliovirus, coxsackievirus, and mengovirus (9,22). These examples suggest that a common step in the life cycle of these picornaviruses is the target for the antiviral drug.In particular, we have demonstrated that PDTC has negative effects on picornavirus replication by influencing the processing of the viral polyprotein (21,22).The antiviral activity of PDTC is not restricted to the family Picornaviridae, since PDTC was shown to prevent the multiplication of human influenza virus, a member of the Orthomyxoviridae (33, 34). However, due to strong differences in the life cycle and host-cell interaction between human influenza virus and picornaviruses, it is likely that entirely different mechanisms might be relevant for the antiviral action of PDTC against these viruses.…”

“…We have previously described the commonly used NF-B inhibitor and metal ion chelator pyrrolidine-dithiocarbamate (PDTC) to significantly inhibit the replication of several picornaviruses such as human rhinovirus (HRV), poliovirus, coxsackievirus, and mengovirus (9,22). These examples suggest that a common step in the life cycle of these picornaviruses is the target for the antiviral drug.…”

Cellular kinase inhibitors that suppress enterovirus replication have a conserved target in viral protein 3A similar to that of enviroxime

“…Calpain inhibitors inhibit the replication of echovirus 1, possibly by targeting the activities of calpain 1 and 2 (Upla et al, 2008). Pyrrolidine dithiocarbamate (PDTC) inhibits polyprotein processing of entero-and cardioviruses by transporting zinc ions into the cells with different mechanisms depending on the virus species (Krenn et al, 2005;Lanke et al, 2007). For PDTC, a mechanism via the ubiquitin-proteasome pathway is also proposed for the inhibitory effect on coxsackievirus B3 (CVB3) infection (Si et al, 2005).…”