An E3 ubiquitin ligase fromNicotiana benthamianatargets the replicase ofBamboo mosaic virusand restricts its replication

Abstract

AbstractOne upregulated host gene identified previously was found involved in the infection process ofBamboo mosaic virus(BaMV). The full-length cDNA of this gene was cloned by 5′- and 3′-rapid amplification of cDNA ends and found to encode a polypeptide containing a conserved RING-domain and a transmembrane domain. The gene might function as an E3 ubiquitin ligase. We designated this protein inNicotiana benthamianaas ubiquitin E3 ligase containing RING-domain 1 (NbUbE3R1). Further characterization by usingTobacco rattle virus-based virus-induced gene silencing revealed an increased BaMV accumulation in both knockdown plants and protoplasts. To further inspect the functional role of NbUbE3R1 in BaMV accumulation, NbUbE3R1 was expressed inN. benthamianaplants. The wild-type NbUbE3R1-orange fluorescent protein (NbUbE3R1-OFP), NbUbE3R1/△TM-OFP (removal of the transmembrane domain) and NbUbE3R1/mRING-OFP (mutation at the RING domain, the E2 interaction site) were transiently expressed in plants. NbUbE3R1 and its derivatives all functioned in restricting BaMV accumulation. The common feature of these constructs was the intact substrate-interacting domain. Yeast two-hybrid and co-immunoprecipitation experiments used to determine the possible viral-encoded substrate of NbUbE3R1 revealed the replicase of BaMV as the possible substrate. In conclusion, we identified an upregulated gene, NbUbE3R1, that plays a role in BaMV replication.