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bioRxiv · 10.64898/2026.10.13.758695

Dual role of the RIG-I 2CARD as a signaling module and a brake on dsRNA translocation

Kondo, S.·Zhang, J.·Matsushima, K.·Kurihara, N.·Flechsig, H.·Umeda, K.·Kodera, N.·Kato, K.·Shibata, M.
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Abstract

RIG-I is an innate immune receptor for viral double-stranded RNA (dsRNA). Upon binding dsRNA, RIG-I nucleates MAVS filaments through its two caspase activation and recruitment domains (2CARD). However, how the 2CARD behaves during activation has remained unclear. Using high-speed atomic force microscopy, we visualized the 2CARD in full-length RIG-I across its apo, dsRNA-bound, translocating, multimeric, and polyubiquitin-bound states. Protrusions consistent with the 2CARD appeared only transiently in the apo state, whereas a mobile 2CARD emerged upon RNA binding. During ATP-driven translocation, wild-type RIG-I moved approximately twofold slower and showed more frequent pauses and multimers than RIG-I {Delta}2CARD. Within multimers, 2CARDs formed a complex that polyubiquitin stabilized into larger, dsRNA-deforming assemblies. These findings reveal the 2CARD as a brake on the RIG-I motor, beyond signaling.

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Kondo, S., Zhang, J., Matsushima, K., Kurihara, N., Flechsig, H., Umeda, K., Kodera, N., Kato, K., Shibata, M.. 2026-10-14. Dual role of the RIG-I 2CARD as a signaling module and a brake on dsRNA translocation. https://doi.org/10.64898/2026.10.13.758695

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