Ubiquitin E3 ligase SPOP is a host negative regulator of enterovirus 71-encoded 2A protease

The 2A protease (2A(pro)) encoded by enterovirus 71 (EV71) serves as an accessory protein with significant involvement in the regulation of EV71 infection and viral replication. EV71-2A(pro) exhibits the ability to suppress various host factors, thereby disrupting the cellular antiviral immune response. However, whether host factors downregulate EV71-2A(pro) remains largely unknown. Here, we discovered that the speckle-type POZ protein (SPOP), functioning as a host E3 ubiquitin ligase, triggers ubiquitination modifications and subsequent degradation of EV71-2A(pro). SPOP interacts with EV71-2A(pro) and exhibits a dose-dependent downregulation of EV71-2A(pro) levels. Conversely, knockdown of endogenous SPOP upregulated EV71-2A(pro) levels. Subsequent investigations revealed that the SPOP facilitates the lysosome-dependent degradation of EV71-2A(pro) by inducing K48-linked polyubiquitination of EV71-2A(pro), which ultimately restricts EV71 replication. These findings shed light on the ubiquitination and lysosome-dependent regulation of the crucial EV71-encoded protease, offering a potential therapeutic target for the treatment of EV71 infection. IMPORTANCE EV71 poses a significant health threat to children aged 5 and below. The process of EV71 infection and replication is predominantly influenced by ubiquitination modifications. Our previous findings indicate that EV71 prompts the activation of host deubiquitinating enzymes, thereby impeding the host interferon signaling pathway as a means of evading the immune response. Nevertheless, the precise mechanisms by which the host employs ubiquitination modifications to hinder EV71 infection remain unclear. The present study demonstrated that the nonstructural protein 2A(pro), which is encoded by EV71, exhibits ubiquitination and degradation mediated by the host E3 ubiquitin ligase SPOP. In addition, it is the first report, to our knowledge, that SPOP is involved in the host antiviral response.