Sandbox 9CC8 AA

From Proteopedia

Introduction

NLRs, also known as nucleotide-binding leucine-rich repeat receptors, are responsible for plant innate immunity. NRC4, a ‘helper’ NLR required for cell death, assembles into a hexameric resistosome, a wheel-like oligomeric complex that forms on the plasma membrane. The formation of the complex is driven by conserved motifs in the coiled-coil (CC) domain. This assembly triggers the plant’s immune response by opening the Ca²⁺ channel. Various plant-specific factors, together with the activators, regulate calcium influx. This structural finding provides new insights into the mechanisms underlying defense signaling in plants.

Structural Highlights and Biological Insights

Structural Highlights

The cryo-EM structure of the resistosome reveals that six identical NRC4 protomers assemble into a symmetric hexamer of ~180Å in diameter at the plasma membrane. Each protomer contains a leucine-rich repeat (LRR) domain and a nucleotide-binding (NB-ARC) domain, which is itself subdivided into the Nucleotide-Binding domain (NBD), Helical Domain 1 (HD1), and a Winged-Helix Domain (WHD). The coiled-coil (CC) domains of each protomer face inward, enabling formation of a central pore, which is crucial for calcium (Ca²⁺) influx. Stability between adjacent CC domains is achieved when the α2-helix from one protomer engages with the α3-helix of its neighboring protomer. This pore is lined by the N-terminal α1-helix from each of the six protomers and is rich in key acidic residues, creating the Ca²⁺ channel. While ATP binding initiates assembly, the final resolved cryo-EM structure (9CC8) represents the post-hydrolysis state, with ADP and Mg²⁺ bound, and inter-subunit contacts stabilizing the overall complex. Structural analyses uncovered two states of the complex: the active hexamer (9CC8) and an alternative inactive (9CC9) dodecameric 'dumbbell' form where the CC domains of one ring are shielded by the LRR domains of the other, and the pore is closed.

Biological insights

The hexameric structure enables NRC4 to act directly as a Ca²⁺ channel, thus creating a direct mechanistic link between pathogen detection and defense execution through cell death. The discovery of a second, inhibited state highlights a structural mechanism for tight regulation of immune activation. These findings advance understanding of how plant immune receptors regulate robust cellular responses by modulating their own assembly and membrane channel activity. The unique topology also sets NRC4 apart from pentameric or tetrameric resistosomes and indicates diverse evolutionary strategies for NLR family immune receptors. Additionally, understanding of activation and channel formation of the resistosomes may help to design crops with enhanced resistance to pathogens.

References

• Liu, F., Yang, Z., Wang, C., You, Z., Martin, R., Qiao, W., Huang, J., Jacob, P., Dangl, J.L., Carette, J.E., Luan, S., Nogales, E., Staskawicz, B.J. "Activation of the helper NRC4 immune receptor forms a hexameric resistosome." Cell (2024) 187: 4877-4889.e15. [1](https://www.sciencedirect.com/science/article/pii/S0092867424007736)
• RCSB PDB 9CC8: Hexameric state of the NRC4 resistosome. [2](https://www.rcsb.org/structure/9CC8)
• RCSB PDB 9CC9: Dodecameric (inactive) state of the NRC4 resistosome. [3](https://www.rcsb.org/structure/9CC9)