Two homologous Alt a1-like fungal proteins possess dual activities in HIR-associated immune signaling and EDS1-dependent cell death

Necrotrophic fungi secrete numerous Cell Death-Inducing Proteins (CDIPs) that manipulate host immunity to promote disease, yet the signaling pathways underlying their phytotoxic activity remain poorly understood. Here, we identify the Botrytis cinerea Hypersensitive response-inducing protein 1 (Hip1) as a close homolog of the recently described Sclerotinia sclerotiorum effector Plant Early Immunosuppressive Effector 1 ( PEIE1) and investigate the molecular basis of its activity. HIP1 and PEIE1 share high sequence similarity and a conserved AlphaFold-predicted Alt a1-like fold, they interact with the Arabidopsis plasma membrane protein HIR4, and they induce strong necrosis in Nicotiana benthamiana . Despite their high structural similarity, Hip1 and PEIE1 differ in their reported roles during fungal infection. Unexpectedly, Hip1-induced cell death requires the central immune regulator ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) as well as the downstream helper NLR network comprising ADR1 and NRG1. Together, our findings establish Hip1 as a closely related homolog of PEIE1 and suggest that these closely related Alt a1-like proteins possess dual activities: modulation of HIR-associated immune signaling and activation of EDS1-dependent host cell death.