Abstract

The βC1 protein encoded by the Tomato yellow leaf curl China virus associated betasatellite functions as a pathogenicity determinant. To better understand the molecular basis whereby βC1 functions in pathogenicity, a yeast two-hybrid screen of a tobacco cDNA library was carried out using βC1 as bait. The screen revealed that βC1 interacts with a tobacco RING-finger protein designated NtRFP1, and the interaction was confirmed using a bimolecular fluorescence complementation (BiFC) and Co-immunoprecipitation (CoIP) assays inNicotiana benthamiana cells. NtRFP1 expression was induced by βC1, and in vitro ubiquitination assays showed that NtRFP1 is a functional E3 ubiquitin ligase that mediates βC1 ubiquitination. In addition, βC1 was shown to be ubiquitinated in vivo and degraded by the plant 26S proteasome. After viral infection, plants overexpressing NtRFP1 developed attenuated symptoms, and plants silenced for NtRFP1 expression had severe symptoms. Other lines of evidence showed that NtRFP1 attenuates βC1 symptoms by promoting βC1 degradation by the 26S proteasome. These results suggest that the tobacco RING E3 ligase NtRFP1 attenuates disease symptoms by interacting with βC1 to mediate its ubiquitination and degradation via the ubiquitin/26S proteasome system.