The ubiquitin ligase itch and ubiquitination regulate BFRF1-mediated nuclear envelope modification for Epstein-Barr Virus maturation

摘要

[[abstract]]The cellular endosomal sorting complex required for transport (ESCRT) was recently found to mediate important morphogenesis processes at the nuclear envelope (NE). We previously showed that the Epstein-Barr virus (EBV) BFRF1 protein recruits the ESCRT-associated protein Alix to modulate NE structure and promote EBV nuclear egress. Here, we uncover new cellular factors and mechanisms involved in this process. BFRF1-induced NE vesicles are similar to those observed following EBV reactivation. BFRF1 is ubiquitinated and elimination of possible ubiquitination with either lysine mutations or fusion of a de-ubiquitinase hampers NE-derived vesicle formation and virus maturation. While interacting with multiple Nedd4-like ubiquitin ligases, BFRF1 binds Itch ligase preferably. We show that Itch associates with Alix and BFRF1, and is required for BFRF1-induced NE vesicle formation. Our data demonstrate that Itch, ubiquitin and Alix control the BFRF1-mediated modulation of the NE and EBV maturation, uncovering novel regulatory mechanisms of nuclear egress of viral nucleocapsids. IMPORTANCE: The nuclear envelope (NE) of eukaryotic cells not only serves as a transverse scaffold for cellular processes, but also a natural barrier for most DNA viruses that assemble their nucleocapsids in the nucleus. Previously, we showed the cellular endosomal sorting complex required for transport (ESCRT) machinery is required for the nuclear egress of EBV. Here, we further report the molecular interplays among viral BFRF1, ESCRT-adaptor Alix and ubiquitin ligase Itch. We found that BFRF1-induced NE vesicles are similar to those observed following EBV reactivation. The lysine residues and the ubiquitination of BFRF1 regulate the formation of BFRF1-induced NE-derived vesicles and EBV maturation. During the process, an ubiquitin ligase Itch associates preferably with BFRF1 and is required for BFRF1-induced NE vesicle formation. Therefore, our data indicate that Itch, ubiquitin and Alix control the BFRF1-mediated modulation of the NE, suggesting novel regulatory mechanisms for ESCRT-mediated NE modulation.