The Tomato Nucleotide-binding Leucine-rich Repeat Immune Receptor I-2 Couples DNA-binding to Nucleotide-binding Domain Nucleotide Exchange

Abstract

Plant nucleotide-binding leucine-rich repeat (NLR) proteins enable plants to recognize and respond to pathogen attack. Previously, we demonstrated that the Rx1 NLR of potato is able to bind and bend DNA in vitro. DNA binding in situ requires its genuine activation following pathogen perception. However, it is unknown whether other NLR proteins are also able to bind DNA. Nor is it known how DNA binding relates to the ATPase activity intrinsic to NLR switch function required to immune activation. Here we investigate these issues using a recombinant protein corresponding to the N-terminal coiled-coil and nucleotide-binding domain regions of the I-2 NLR of tomato. Wild type I-2 protein bound nucleic acids with a preference of ssDNA ≈ dsDNA > ssRNA, which is distinct from Rx1. I-2 induced bending and melting of DNA. Notably, ATP enhanced DNA binding relative to ADP in the wild type protein, the null P-loop mutant K207R, and the autoactive mutant S233F. DNA binding was found to activate the intrinsic ATPase activity of I-2. Because DNA binding by I-2 was decreased in the presence of ADP when compared with ATP, a cyclic mechanism emerges; activated ATP-associated I-2 binds to DNA, which enhances ATP hydrolysis, releasing ADP-bound I-2 from the DNA. Thus DNA binding is a general property of at least a subset of NLR proteins, and NLR activation is directly linked to its activity at DNA.

The Coiled-Coil and Nucleotide Binding Domains of BROWN PLANTHOPPER RESISTANCE14 Function in Signaling and Resistance against Planthopper in Rice

“…In addition, the NB domains of several disease resistance proteins exhibit nucleotide phosphatase activity consistent with the switch model described above (Fenyk et al, 2012). Interestingly, Rx1 was recently found to bend and locally melt double-stranded DNA in a process mediated by its NB domain (Fenyk et al, 2015(Fenyk et al, , 2016. The functional activity of this NLR protein may thus be linked to a conserved direct interaction with DNA.…”

Animal NLRs provide structural insights into plant NLR function

“…It remains to be seen whether the DNA-binding properties of certain NLRs prove to play a role in plant NLR signalling (Fenyk et al, 2016(Fenyk et al, , 2015.…”

The Conformation of a Plasma Membrane-Localized Somatic Embryogenesis Receptor Kinase Complex Is Altered by a Potato Aphid-Derived Effector

“…Similarly, the functional importance of Mi-1.2 nuclear localization for immune signaling remains to be investigated. Nevertheless, it is intriguing to speculate that Mi-1.2 directly interacts with TFs WRKY72a or WRKY72b to regulate immune responses or might act directly on the plant DNA like Rx and I-2, two solanaceous C proteins recently found to interact with DNA following their activation (Fenyk et al, 2015(Fenyk et al, , 2016.…”