Global analysis of ubiquitome in PRRSV-infected pulmonary alveolar macrophages

Abstract

Protein lysine ubiquitination is a dynamic reversible post-translational modification that plays key roles in modulating different cellular processes. Porcine reproductive and respiratory syndrome virus (PRRSV) is a notorious pathogen, causing tremendous economic losses for the global swine industry. The possible involvement of ubiquitination in PRRSV infection is unclear. So anti-ubiquitination-based enrichment and LC-MS were performed to investigate the global ubiquitination events triggered by PRRSV infection in pulmonary alveolar macrophages. We totally identified 4044 lysine ubiquitination sites on 1580 cellular proteins, of which 983 sites on 717 proteins were significantly altered at 36 h postinfection. A systematic, intensive bioinformatic analysis of the ubiquitome data suggested that PRRSV suppresses the host immune responses by manipulating the ubiquitination of important adaptors and effectors, including TRAF6, JAK1, STAT1, and ISGs. Ubiquitination was also observed on 15 PRRSV proteins, including important virus proteases and structural proteins that function in virus infectivity and neutralizing antibody elicitation. The efficient replication of PRRSV requires an intact ubiquitin–proteasome system. Our study is the first to analyze the global ubiquitination events in pulmonary alveolar macrophages during PRRSV infection. It provides insight into the molecular mechanisms of PRRSV pathogenesis, promoting the development of antiviral drugs. Biological significancePRRSV is a notorious pathogen which has been resulting in huge economic losses in the swine industry since the first outbreak. Therefore, more in-depth knowledge of the PRRSV immunoregulatory mechanisms and valid control methods to combat the virus are urgently needed. Ubiquitination is an important post-translational modification regulating various cellular processes. However, information about the possible involvement of ubiquitination responses to PRRSV infection is limited. In this study, a quantitative proteomic approach was first used to analyze ubiquitination level alteration in PRRSV-infected PAMs. We demonstrate that PRRSV can suppresses the host immune responses by manipulating the ubiquitination of important effectors that include TRAF6, JAK1, STAT1, and ISGs. Furthermore, 15 PRRSV proteins undergo ubiquitination and efficient replication of PRRSV requires an intact ubiquitin–proteasome system. Our study will significantly expand our knowledge about the molecular mechanisms of PRRSV pathogenesis and provides novel insights into the development of antiviral drugs.