Ku suppresses RNA-mediated innate immune responses in human cells to accommodate primate-specific Alu expansion.

Zhu, Y.; LI, A.; Maji, S.; Lee, B. J.; Korn, S. M.; Gertie, J. A.; Dorrity, T. J.; Wang, J.; Wang, K. J.; Pelletier, A.; Moakley, D. F.; Kelly, R. D.; Holmes, A. B.; Rabadan, R.; Edgell, D. R.; Schild-Poulter, C.; Modesti, M.; Steckelberg, A.-L.; Hendrickson, E. A.; Chung, H.; Zhang, C.; Zha, S.

doi:10.1101/2025.01.31.633084

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bioRxiv · 10.1101/2025.01.31.633084

Ku suppresses RNA-mediated innate immune responses in human cells to accommodate primate-specific Alu expansion.

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Abstract

Ku70 and Ku80 form Ku, a ring-shaped protein that initiates the non-homologous end-joining (NHEJ) DNA repair pathway.1 Specifically, Ku binds to double-stranded DNA (dsDNA) ends and recruits other NHEJ factors (e.g., DNA-PKcs and LIG4). While Ku binds to double-stranded RNA (dsRNA)2 and traps mutated-DNA-PKcs on ribosomal RNA in vivo,3,4 the physiological significance of Ku-dsRNA interactions in otherwise wild-type cells remains elusive. Intriguingly, while dispensable for murine development,5,6 Ku is essential in human cells.7 Despite similar genome sizes, human cells express [~]100-fold more Ku than mouse cells, implying functions beyond NHEJ, possibly through a dose-sensitive interaction with dsRNA, which is [~]100 times weaker than with dsDNA.2,8 While investigating the essentiality of Ku in human cells, we found that depletion of Ku - unlike LIG4 - induces profound interferon (IFN) and NF-kB responses reliant on the dsRNA-sensor MDA5/RIG-I and adaptor MAVS. Prolonged Ku-degradation also activates other dsRNA-sensors, e.g. PKR that suppresses protein translation, and OAS/RNaseL that cleaves rRNAs and eventually induces growth arrest and cell death. MAVS, RIG-I, or MDA5 knockouts suppressed IFN signaling and, together with PKR knockouts, partially rescued Ku-depleted human cells. Ku-irCLIP analyses revealed that Ku binds to diverse dsRNA, predominantly stem-loops in primate-specific Alu elements9 at anti-sense orientation in introns and 3-UTRs. Ku expression rose sharply in higher primates tightly correlating with Alu-expansion (r = 0.94/0.95). Together, our study identified a vital role of Ku in accommodating Alu-expansion in primates by mitigating a dsRNA-induced innate immune response, explaining the rise of Ku levels and its essentiality in human cells.

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Zhu, Y., LI, A., Maji, S., Lee, B. J., Korn, S. M., Gertie, J. A., Dorrity, T. J., Wang, J., Wang, K. J., Pelletier, A., Moakley, D. F., Kelly, R. D., Holmes, A. B., Rabadan, R., Edgell, D. R., Schild-Poulter, C., Modesti, M., Steckelberg, A.-L., Hendrickson, E. A., Chung, H., Zhang, C., Zha, S.. 2025-02-01. Ku suppresses RNA-mediated innate immune responses in human cells to accommodate primate-specific Alu expansion.. https://doi.org/10.1101/2025.01.31.633084

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