An antiviral targeting strategy based on the inducible interference with cytomegalovirus nuclear egress complex

Abstract

The nucleocytoplasmic capsid egress of herpesviruses like the human cytomegalovirus (HCMV) is based on a uniquely regulated process. The core nuclear egress complex (NEC) of HCMV, represented by the pUL50-pUL53 heterodimer, is able to oligomerize and thus to build hexameric lattices. Recently, we and others validated the NEC as a novel target for antiviral strategies. So far, the experimental targeting approaches included the development of NEC-directed small molecules, cell-penetrating peptides and NEC-directed mutagenesis. Our postulate states that an interference with the hook-into-groove interaction of pUL50-pUL53 prevents NEC formation and strictly limits viral replication efficiency. Here, we provide an experimental proof-of-concept of the antiviral strategy: the inducible intracellular expression of a NLS-Hook-GFP construct exerted a pronounced level of antiviral activity. The data provide evidence for the following points: (i) generation of a primary fibroblast population with inducible NLS-Hook-GFP expression showed nuclear localization of the construct, (ii) interaction between NLS-Hook-GFP and the viral core NEC was found specific for cytomegaloviruses but not for other herpesviruses, (iii) construct overexpression exerted a strong antiviral activity against three strains of HCMV, (iv) confocal imaging demonstrated the interference with NEC nuclear rim formation in HCMV-infected cells, and (v) quantitative nuclear egress assay confirmed the block of viral nucleocytoplasmic transition and, consequently, an inhibitory effect onto viral cytoplasmic virion assembly complex (cVAC). Combined, data confirmed that the specific interference with protein-protein interaction of the HCMV core NEC represents an efficient antiviral targeting strategy.

Keywords: Antiviral proof-of-concept; Cytomegalovirus infection; Efficient interference with viral replication; Hook-into-groove principle; Inducible NLS-Hook-GFP expression; Major human pathogen; Novel drug targeting options; Nuclear egress-based antiviral strategy.