K48-linked polyubiquitination of dengue virus NS1 protein inhibits its interaction with the viral partner NS4B

Abstract

Dengue virus (DENV) is a member of the Flaviviridae family, which is transmitted to mammalian species through arthropods, and causes dengue fever or severe dengue fever in humans. The DENV genome encodes for multiple nonstructural (NS) proteins including NS1. NS1 plays an essential role in replication by interacting with other viral proteins including NS4B, however how these interactions are regulated during virus infection is not known. By using bioinformatics, mass spectrometry analysis, and co-immunoprecipitation assays, here we show that DENV-NS1 is ubiquitinated on multiples lysine residues during DENV infection, including K189, a lysine residue previously shown to be important for efficient DENV replication. Data from in vitro and cell culture experiments indicate that dengue NS1 undergoes modification with K48-linked polyubiquitin chains, which usually target proteins to the proteasome for degradation. Furthermore, ubiquitinated NS1 was detected in lysates as well as in supernatants of human and mosquito infected cells. Ubiquitin deconjugation of NS1 using the deubiquitinase OTU resulted in increased interaction with the viral protein NS4B suggesting that ubiquitinated NS1 has reduced affinity for NS4B. In support of these data, a K189R mutation on NS1, which abrogates ubiquitination on amino acid residue 189 of NS1, also increased NS1-NS4B interactions. Our work describes a new mechanism of regulation of NS1-NS4B interactions and suggests that ubiquitination of NS1 may affect DENV replication.

Pleiotropic roles of the ubiquitin-proteasome system during viral propagation

“…To date, many viral proteins have been reported to be ubiquitinated, including influenza A virus-encoded NP, M2 and PB1 [25][26][27][28], Dengue Virus-encoded NP [29], and 54 viral proteins encoded by the wellcharacterised cowpox virus strain [30]. In ubiquitinated proteins, Ub is frequently found to be covalently conjugated to Lys residues, but non-Lys residues of substrates such as serine (Ser), threonine (Thr) and cysteine (Cys) are also modified in rare cases.…”

The E3 Ubiquitin Ligase TRIM21 Promotes HBV DNA Polymerase Degradation

“…In addition, most members of the virus family are also degraded by the ubiquitin proteasome pathway, such as the ubiquitination-mediated degradation of cowpox virus proteins, which provide the diverse roles of ubiquitin in the process of virus replication [50]. The NS1 protein encoded by dengue virus is degraded by polyubiquitination with a K48-linked polyubiquitin chain, which may affect virus replication [51]. The degradation of HBV core protein (HBc) is mediated by NIRF (Np95/ICBP90-like RING finger protein) via the ubiquitin proteasome pathway, which affects the release of virus particles [52].…”

Porcine RING Finger Protein 114 Inhibits Classical Swine Fever Virus Replication via K27-Linked Polyubiquitination of Viral NS4B

“…The E3 ubiquitin ligase MKRN1 induces degradation of WNV capsid protein in a proteasome-dependent manner (36). It has been reported that the K48-linked polyubiquitination of the DENV NS1 protein inhibits its interaction with the viral partner NS4B, suggesting that NS1 ubiquitination may affect virus replication (37). In addition, it has been shown that the CSFV N pro induces degradation of IRF3 via the proteasome pathway; possibly, N pro recruits E3 ubiquitin ligase complex to degrade IRF3 (38).…”