Abstract
The ubiquitin-proteasome system is a highly conserved machinery that plays a crucial role in plant defense against viruses. However, the number of E3 ligases targeting viral proteins remains limited. Although RING-between-RING (RBR)-type E3 ligases are evolutionarily conserved across organisms, their functions in plant responses to biotic stress remain largely unknown. Herein, it is found that the triple gene block 1 (TGB1) protein of the barley stripe mosaic virus (BSMV) undergoes ubiquitination during viral infection. Immunoprecipitation combined with mass spectrometry identified an RBR-type E3 ligase that interacted with TGB1 in vivo and in vitro. The overexpression of Ariadne-like protein 8 (ARI8) inhibits, whereas its knockout enhances, the local and systemic spread of BSMV. ARI8 mediated the ubiquitination of TGB1, and its Cys311 residue is required for the ARI8-mediated degradation of TGB1 and inhibition of BSMV infection. In addition to BSMV, ARI8 negatively regulates infection by other TGB-containing viruses, including potato virus X and beet necrotic yellow vein virus. Collectively, the findings identified a new E3 ligase that targets a plant viral protein and reveals a previously uncharacterized role for RBR-type E3 ligases in plant responses to biotic stress, providing a potential molecular target for the development of antiviral strategies in plants.