Canonical EDS1/PAD4 small-molecule binding sites are required for LRR-RP-mediated pattern-triggered immunity.

Pattern-triggered immunity (PTI) is a key plant defense mechanism initiated by the recognition of pathogen-associated molecular patterns (PAMPs) by cell-surface receptors, including leucine-rich repeat receptor proteins (LRR-RPs). Upon PAMP perception, LRR-RPs associate with SOBIR1 and recruit SERK coreceptors, such as BAK1, into the receptor complex to initiate PTI. The lipase-like protein EDS1 and PAD4 are central components of effector-triggered immunity (ETI). Small molecule (SM) phosphoribosyl-AMP/ADP, generated by TIR-domain NADase activity, promotes the recruitment of helper NLRs of the ADR1 family into the EDS1-PAD4 complex to execute ETI. This ETI signaling branch, known as the EDS1-PAD4-ADR1 node, is associated with SOBIR1 and is essential for LRR-RP-mediated PTI. To elucidate the molecular mechanism by which EDS1-PAD4 contributes to PTI, we show that PTI initiation mediated by BAK1 recruitment to LRR-RP complexes is unaffected in eds1-2 or pad4-1 mutants but is abolished in the sobir1-12 mutant, highlighting the essential role of SOBIR1 in surface immune receptor complex formation. Furthermore, we demonstrate that SM binding to EDS1 and PAD4, in addition to its role in ETI, is also critical for LRR-RP-mediated PTI, as SM-binding-deficient mutants fail to restore PTI responses. Moreover, de novo SM synthesis is required for late PTI responses, including PR1 expression and bacterial resistance, but not for early responses such as reactive oxygen species and ethylene production. Finally, the TNL SADR1 functions specifically in late PTI responses. Together, these findings indicate that a preformed EDS1-PAD4-ADR1 complex is essential for PTI and reveal a two-tiered role for TIR-domain proteins in PTI signaling.