Functional Characterization of Deubiquitinase UBP Family and Proteomic Analysis of Aaubp14-Mediated Pathogenicity Mechanism in Alternaria alternata

TheAlternaria alternatatangerine pathotype causes Alternaria brown spot, a devastating disease of susceptible tangerine varieties and their hybrids.Alternaria citritoxin (ACT) is the primary virulence factor, but the regulatory mechanisms governing ACT synthesis remain unclear. Deubiquitinating enzymes maintain ubiquitination homeostasis and regulate fungal pathogenicity, yet their role inA. alternataremains unexplored. We characterized 13 ubiquitin-specific protease (UBP) family members inA. alternatatangerine pathotype. Six UBP genes (Aaubp2,Aaubp3,Aaubp4,Aaubp6,Aaubp14, andAaubp15) regulated mycelial growth.Aaubp14deletion abolished sporulation, while mutations ofAaubp3,Aaubp4,Aaubp6,Aaubp8, andAaubp15altered conidial morphology. qRT-PCR demonstrated distinct host-induced expression patterns amongAaubpgenes. Pathogenicity tests showed that ΔAaubp6, ΔAaubp14, and ΔAaubp15mutants failed to produce lesions onCitrus reticulatacv. Hongjv leaves. Moreover,Aaubp14deletion significantly suppressed ACT biosynthesis gene expression and blocked ACT production. Comparative proteomics showed Aaubp14 regulates ACT biosynthesis by modulating protein ubiquitination in metabolic pathways and controls pathogenicity via a complex network. Our findings elucidateAaubpgene function in development and pathogenicity, particularly the Aaubp14-mediated regulation mechanism, providing insights into ubiquitination-mediated pathogenicity in phytopathogenic fungi.