Co-opted CHIP E3 Ub-ligase provides protein quality control by regulating molecular properties of condensates formed by a positive-strand RNA virus

Abstract

Positive-strand RNA viruses are important pathogens that built viral replication organelles (VROs) via exploiting co-opted host proteins and intracellular membranes in infected cells. Tomato bushy stunt virus (TBSV) membranous VROs are associated with dynamic virus-induced replication (vir)–condensate substructures driven by TBSV p33 replication protein. We targeted micropeptide “killswitch” to VROs to enhance p33 self-association and remodel vir-condensates. Recruitment of killswitch induced p33 aggregations, reduced mobility of co-opted glycolytic enzymes in vir-condensate, limited ATP production within VROs, and inhibited TBSV replication. We discovered that TBSV co-opts CHIP/STUB1 E3 ubiquitin ligase involved in protein quality control to limit detrimental effect of protein aggregations in VROs. Expression of CHIP reduced accumulation of aggregated TBSV p33. Knockdown of CHIP resulted in inhibition of TBSV replication, enhanced accumulation of aggregation-prone p33 mutants, and inhibited mobility of co-opted client host proteins in vir-condensate. Subverted CHIP plays critical role in regulating protein interactions, stability, and maintenance of dynamic vir-condensate. Hijacking protein quality control mechanism is a previously unknown phenomenon during (+)RNA virus replication.