Abstract TIR domains are signalling domains present in plant nucleotide-binding leucine-rich repeat receptors (NLRs), with key roles in plant innate immunity. They are required for the induction of a hypersensitive response (HR) in effector-triggered immunity, but the mechanism by which this occurs is not yet fully understood. It has been recently shown that the TIR domains from several plant NLRs possess NADase activity. The oligomeric structure of TIR-containing NLRs ROQ1 and RPP1 reveals how the TIR domains arrange into an active conformation, but low resolution around the NAD + binding sites leaves questions unanswered about the molecular mechanisms linking self-association and NADase activity. In this study, a number of crystal structures of the TIR domain from the grapevine NLR RUN1 reveal how self-association and enzymatic activity may be linked. Structural features previously proposed to play roles involve the “AE interface” (mediated by helices A and E), the “BB-loop” (connecting β-strand B and helix B in the structure), and the “BE interface” (mediated by the BB-loop from one TIR and the “DE surface” of another). We demonstrate that self-association through the AE interface induces conformational changes in the NAD + -binding site, shifting the BB-loop away from the catalytic site and allowing NAD + to access the active site. We propose that an intact “DE surface” is necessary for production of the signalling product (variant cyclic ADPR), as it constitutes part of the active site. Addition of NAD + or NADP + is not sufficient to induce self-association, suggesting that NAD + binding occurs after TIR self-association. Our study identifies a mechanistic link between TIR self-association and NADase activity.

Self-association configures the NAD+-binding site of plant NLR TIR domains