BBTV Nuclear Shuttle Protein Mediates Banana Ubiquitination Pathway Dysregulation

Banana bunchy top virus (BBTV) is a devastating pathogen threatening global banana production. The plant ubiquitin-proteasome system (UPS) governs immune signaling and is frequently subverted by invading viruses, yet the molecular mechanism through which BBTV interferes with host UPS remains unclear. Here, we show that BBTV nuclear shuttle protein (NSP) serves as the core viral effector to disrupt banana ubiquitination homeostasis. RT-qPCR time-series assays confirmed that BBTV infection dynamically remodels the transcription of eight phylogenetically divergent RING-type E3 ubiquitin ligases: four subfamily I E3-SIS3 paralogs and E3-HIP1 are significantly upregulated at 14 dpi and 21 dpi, while E3-BOI and E3-RHA1B are suppressed at 21 dpi. Transient expression screening of all six BBTV-encoded proteins verified that only NSP reproduces the UPS perturbation signature triggered by viral infection. Cross-species sequence alignment identified an evolutionarily conserved FNGSF motif within NSP orthologs of all Nanoviridae members. Alanine substitution mutagenesis (NSP^AAAAA) completely abolished NSP's capacity to alter E3 ligase transcription. Western blot assays further validated that wild-type NSP induces massive accumulation of ubiquitinated host proteins, whereas the FNGSF-deficient mutant does not disrupt cellular ubiquitination. Phylogenetic analysis revealed that NSP-targeted E3 ligases share low overall sequence similarity but retain conserved catalytic RING domains, indicating that NSP exerts broad-spectrum regulatory effects on host UPS via the FNGSF motif. Collectively, this study reveals a novel pathogenic strategy whereby BBTV NSP recruits diverse host RING E3 ligases via its conserved FNGSF motif to dysregulate plant ubiquitination and elicit plant pathogenicity. Our findings provide two promising targets-the NSP FNGSF motif and defense-associated E3-SIS3 ligases-for developing antiviral agents and breeding BBTV-resistant banana germplasm.