A Peptide Inhibitor of the Human Cytomegalovirus Core Nuclear Egress Complex

Abstract

The replication of human cytomegalovirus (HCMV) involves a process termed nuclear egress, which enables translocation of newly formed viral capsids from the nucleus into the cytoplasm. The HCMV core nuclear egress complex (core NEC), a heterodimer of viral proteins pUL50 and pUL53, is therefore considered a promising target for new antiviral drugs. We have recently shown that a 29-mer peptide presenting an N-terminal alpha-helical hook-like segment of pUL53, through which pUL53 interacts with pUL50, binds to pUL50 with high affinity, and inhibits the pUL50–pUL53 interaction in vitro. Here, we show that this peptide is also able to interfere with HCMV infection of cells, as well as with core NEC formation in HCMV-infected cells. As the target of the peptide, i.e., the pUL50–pUL53 interaction, is localized at the inner nuclear membrane of the cell, the peptide had to be equipped with translocation moieties that facilitate peptide uptake into the cell and the nucleus, respectively. For the resulting fusion peptide (NLS-CPP-Hook), specific cellular and nuclear uptake into HFF cells, as well as inhibition of infection with HCMV, could be demonstrated, further substantiating the HCMV core NEC as a potential antiviral target.

Assessment of Covalently Binding Warhead Compounds in the Validation of the Cytomegalovirus Nuclear Egress Complex as an Antiviral Target

“…The herpesviral nuclear egress complex (NEC) represents a functionally highly important determinant of viral replication efficiency. Recent studies on HCMV indicated that the disruption of the hook-into-groove interaction of pUL50 and pUL53 inhibits the late phase of viral replication [ 24 , 34 , 40 , 46 ]. Considering the features of core NEC sequences and structures, we addressed the question of whether distinct cysteine residues might be accessible to the attack of inhibitory warhead compounds.…”

“…As a methodological trait, however, it has to be emphasized that the speckled appearance of nuclear rim, i.e., the drug-mediated disruption of the typical NEC nuclear rim localization, does not allow the direct conclusion of a dissociation of the pUL50–pUL53 core NEC heterodimer/oligomer. Previous studies of our group demonstrated that the speckled appearance of the HCMV core NEC can be induced by both direct inhibitors of core NEC proteins and indirect inhibitors of NEC-associated proteins [ 24 , 25 , 34 , 40 ]. As an important finding of the present study, nonetheless, this novel group of warhead small molecules likewise exerts a pronounced phenotypic effect on the viral NEC nuclear rim localization.…”

“…Recent achievements of our group and other researchers have illustrated the functional relevance of herpesviral core NECs, together with the associated proteins of the higher-order multicomponent NECs. Consequently, these proteins are considered as upcoming antiviral targets for next-generation antiherpesviral drugs [ 18 , 21 , 26 , 34 , 40 , 46 ]. Here, we focused on the use of covalently binding warheads and the discussion about their usefulness in the generation of NEC-directed antiviral small molecules.…”

“…Concerning novel targeting strategies of anti-HCMV drugs, our group and many other researchers have studied a variety of different options of direct-acting and host-directed antivirals [ 27 , 32 , 33 ]. Very recently, we focused on the NEC as a highly promising target, which is composed of viral core components (core NEC) and, additionally, NEC-associated host and virus proteins (multicomponent NEC) [ 8 , 24 , 34 , 35 , 36 , 37 , 38 , 39 , 40 ]. A crucial step towards an improved level of discovery of NEC-directed small molecules was our experimental resolution of crystallization-based 3D structures of α-, β- and γ-herpesviral core NECs [ 41 , 42 , 43 ].…”

Investigating In Situ Expression of c-MYC and Candidate Ubiquitin-Specific Proteases in DLBCL and Assessment for Peptidyl Disruptor Molecule against c-MYC-USP37 Complex

“…Since the interacting region of c-MYC for USP37 is structurally disordered, it may not be a suitable target for small molecule inhibitor and/or disruptor [ 55 ]; rather, a peptidyl disruptor could be used to block the interaction between USP37 and c-MYC. Peptidyl disruptors has shown effective inhibition for the protein complex formation such as human ACE2 and the receptor-binding domain (RBD) of the Spike protein of SARS-CoV2 [ 56 ] YAP/TAZ-TEAD Transcriptional Complex [ 57 ] and Human Cytomegalovirus Core Nuclear Egress Complex [ 58 ]. Nearly 80% of the residues of c-MYC that interacts with USP37 have been found interacting with the designed peptide, suggesting that the peptide is specific to c-MYC-USP37 complex and could be used for targeted therapy against the ABC subtype of DLBCL.…”