STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor

Bao-Cun Zhang; Alice Pedersen; Line S Reinert; Ying Li; Ryo Narita; Manja Idorn; Lili Hu; Morten K Skouboe; Sirui Li; Muyesier Maimaitili; Xiangning Ding; Yingying Cong; Jian Zhao; Marie-Louise Frémond; Kasper Mikkelsen; Zongliang Gao; Jin-Rong Huang; Emil A Thomsen; Jakob H Mikkelsen; Rajan Venkatraman; Martin K Thomsen; Marie B Iversen; Sonia Assil; Ran Zhang; Linda Henneman; Martin R Jakobsen; Claus Oxvig; Tina S Dalgaard; Peter Møller; Angela Fago; Tobias Wang; Christian B F Andersen; Dominic De Nardo; Fulvio Reggiori; Jenny P-Y Ting; Jacob G Mikkelsen; Rasmus O Bak; Trine H Mogensen; Pingwei Li; Søren R Paludan

doi:10.1038/s41590-025-02283-8

STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor

Bao-Cun Zhang^*, Alice Pedersen, Line S Reinert, Ying Li, Ryo Narita, Manja Idorn, Lili Hu, Morten K Skouboe, Sirui Li, Muyesier Maimaitili, Xiangning Ding, Yingying Cong, Jian Zhao, Marie-Louise Frémond, Kasper Mikkelsen, Zongliang Gao, Jin-Rong Huang, Emil A Thomsen, Jakob H Mikkelsen, Rajan VenkatramanMartin K Thomsen, Marie B Iversen, Sonia Assil, Ran Zhang, Linda Henneman, Martin R Jakobsen, Claus Oxvig, Tina S Dalgaard, Peter Møller, Angela Fago, Tobias Wang, Christian B F Andersen, Dominic De Nardo, Fulvio Reggiori, Jenny P-Y Ting, Jacob G Mikkelsen, Rasmus O Bak, Trine H Mogensen, Pingwei Li, Søren R Paludan

^*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

Abstract

NF-κB is central for activation of immune responses. Cytosolic DNA activates the cGAS-STING pathway to induce type I interferons (IFNs) and signaling through NF-κB, thus instigating host defenses and pathological inflammation. However, the mechanism underlying STING-induced NF-κB activation is unknown. Here we report that STING activates NF-κB in a delayed manner, following exit from the Golgi to endolysosomal compartments. Activation of NF-κB is dependent on the IFN-inducing transcription factor IRF3 but is independent of type I IFN signaling. This activation pattern is evolutionarily conserved in tetrapods. Mechanistically, the monomer IRF3 is recruited to STING pS358, with delayed kinetics relative to IRF3 recruitment to STING pS366, which promotes type I IFN responses. IRF3 engagement with STING pS358 induces trafficking to late endolysosomal compartments, supporting recruitment of TRAF6 and activation of NF-κB. We identify a TRAF6 binding motif in IRF3 that facilitates recruitment of TRAF6. This work defines a signaling surface on STING and a function for IRF3 as an adaptor in immune signaling. These findings indicate that STING signaling to NF-κB is enabled only within a short time window between exit from the Golgi and lysosomal degradation, possibly limiting inflammation under homeostatic and danger-sensing conditions.

Original language	English
Journal	Nature Immunology
ISSN	1529-2908
DOIs	https://doi.org/10.1038/s41590-025-02283-8
Publication status	E-pub / Early view - 19 Sept 2025

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Zhang, B.-C., Pedersen, A., Reinert, L. S., Li, Y., Narita, R., Idorn, M., Hu, L., Skouboe, M. K., Li, S., Maimaitili, M., Ding, X., Cong, Y., Zhao, J., Frémond, M.-L., Mikkelsen, K., Gao, Z., Huang, J.-R., Thomsen, E. A., Mikkelsen, J. H., ... Paludan, S. R. (2025). STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor. Nature Immunology. Advance online publication. https://doi.org/10.1038/s41590-025-02283-8

@article{574eeff76c394797bc041e19f5f78617,

title = "STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor",

abstract = "NF-κB is central for activation of immune responses. Cytosolic DNA activates the cGAS-STING pathway to induce type I interferons (IFNs) and signaling through NF-κB, thus instigating host defenses and pathological inflammation. However, the mechanism underlying STING-induced NF-κB activation is unknown. Here we report that STING activates NF-κB in a delayed manner, following exit from the Golgi to endolysosomal compartments. Activation of NF-κB is dependent on the IFN-inducing transcription factor IRF3 but is independent of type I IFN signaling. This activation pattern is evolutionarily conserved in tetrapods. Mechanistically, the monomer IRF3 is recruited to STING pS358, with delayed kinetics relative to IRF3 recruitment to STING pS366, which promotes type I IFN responses. IRF3 engagement with STING pS358 induces trafficking to late endolysosomal compartments, supporting recruitment of TRAF6 and activation of NF-κB. We identify a TRAF6 binding motif in IRF3 that facilitates recruitment of TRAF6. This work defines a signaling surface on STING and a function for IRF3 as an adaptor in immune signaling. These findings indicate that STING signaling to NF-κB is enabled only within a short time window between exit from the Golgi and lysosomal degradation, possibly limiting inflammation under homeostatic and danger-sensing conditions.",

author = "Bao-Cun Zhang and Alice Pedersen and Reinert, \{Line S\} and Ying Li and Ryo Narita and Manja Idorn and Lili Hu and Skouboe, \{Morten K\} and Sirui Li and Muyesier Maimaitili and Xiangning Ding and Yingying Cong and Jian Zhao and Marie-Louise Fr{\'e}mond and Kasper Mikkelsen and Zongliang Gao and Jin-Rong Huang and Thomsen, \{Emil A\} and Mikkelsen, \{Jakob H\} and Rajan Venkatraman and Thomsen, \{Martin K\} and Iversen, \{Marie B\} and Sonia Assil and Ran Zhang and Linda Henneman and Jakobsen, \{Martin R\} and Claus Oxvig and Dalgaard, \{Tina S\} and Peter M{\o}ller and Angela Fago and Tobias Wang and Andersen, \{Christian B F\} and \{De Nardo\}, Dominic and Fulvio Reggiori and Ting, \{Jenny P-Y\} and Mikkelsen, \{Jacob G\} and Bak, \{Rasmus O\} and Mogensen, \{Trine H\} and Pingwei Li and Paludan, \{S{\o}ren R\}",

year = "2025",

month = sep,

day = "19",

doi = "10.1038/s41590-025-02283-8",

language = "English",

journal = "Nature Immunology",

issn = "1529-2908",

publisher = "Nature Research",

}

Zhang, B-C , Pedersen, A , Reinert, LS, Li, Y, Narita, R, Idorn, M, Hu, L, Skouboe, MK, Li, S, Maimaitili, M , Ding, X , Cong, Y , Zhao, J, Frémond, M-L, Mikkelsen, K, Gao, Z, Huang, J-R, Thomsen, EA , Mikkelsen, JH, Venkatraman, R, Thomsen, MK , Iversen, MB, Assil, S, Zhang, R, Henneman, L, Jakobsen, MR , Oxvig, C , Dalgaard, TS, Møller, P, Fago, A , Wang, T , Andersen, CBF, De Nardo, D, Reggiori, F, Ting, JP-Y, Mikkelsen, JG , Bak, RO , Mogensen, TH, Li, P & Paludan, SR 2025, 'STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor', Nature Immunology. https://doi.org/10.1038/s41590-025-02283-8

TY - JOUR

T1 - STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor

AU - Zhang, Bao-Cun

AU - Pedersen, Alice

AU - Reinert, Line S

AU - Li, Ying

AU - Narita, Ryo

AU - Idorn, Manja

AU - Hu, Lili

AU - Skouboe, Morten K

AU - Li, Sirui

AU - Maimaitili, Muyesier

AU - Ding, Xiangning

AU - Cong, Yingying

AU - Zhao, Jian

AU - Frémond, Marie-Louise

AU - Mikkelsen, Kasper

AU - Gao, Zongliang

AU - Huang, Jin-Rong

AU - Thomsen, Emil A

AU - Mikkelsen, Jakob H

AU - Venkatraman, Rajan

AU - Thomsen, Martin K

AU - Iversen, Marie B

AU - Assil, Sonia

AU - Zhang, Ran

AU - Henneman, Linda

AU - Jakobsen, Martin R

AU - Oxvig, Claus

AU - Dalgaard, Tina S

AU - Møller, Peter

AU - Fago, Angela

AU - Wang, Tobias

AU - Andersen, Christian B F

AU - De Nardo, Dominic

AU - Reggiori, Fulvio

AU - Ting, Jenny P-Y

AU - Mikkelsen, Jacob G

AU - Bak, Rasmus O

AU - Mogensen, Trine H

AU - Li, Pingwei

AU - Paludan, Søren R

PY - 2025/9/19

Y1 - 2025/9/19

N2 - NF-κB is central for activation of immune responses. Cytosolic DNA activates the cGAS-STING pathway to induce type I interferons (IFNs) and signaling through NF-κB, thus instigating host defenses and pathological inflammation. However, the mechanism underlying STING-induced NF-κB activation is unknown. Here we report that STING activates NF-κB in a delayed manner, following exit from the Golgi to endolysosomal compartments. Activation of NF-κB is dependent on the IFN-inducing transcription factor IRF3 but is independent of type I IFN signaling. This activation pattern is evolutionarily conserved in tetrapods. Mechanistically, the monomer IRF3 is recruited to STING pS358, with delayed kinetics relative to IRF3 recruitment to STING pS366, which promotes type I IFN responses. IRF3 engagement with STING pS358 induces trafficking to late endolysosomal compartments, supporting recruitment of TRAF6 and activation of NF-κB. We identify a TRAF6 binding motif in IRF3 that facilitates recruitment of TRAF6. This work defines a signaling surface on STING and a function for IRF3 as an adaptor in immune signaling. These findings indicate that STING signaling to NF-κB is enabled only within a short time window between exit from the Golgi and lysosomal degradation, possibly limiting inflammation under homeostatic and danger-sensing conditions.

AB - NF-κB is central for activation of immune responses. Cytosolic DNA activates the cGAS-STING pathway to induce type I interferons (IFNs) and signaling through NF-κB, thus instigating host defenses and pathological inflammation. However, the mechanism underlying STING-induced NF-κB activation is unknown. Here we report that STING activates NF-κB in a delayed manner, following exit from the Golgi to endolysosomal compartments. Activation of NF-κB is dependent on the IFN-inducing transcription factor IRF3 but is independent of type I IFN signaling. This activation pattern is evolutionarily conserved in tetrapods. Mechanistically, the monomer IRF3 is recruited to STING pS358, with delayed kinetics relative to IRF3 recruitment to STING pS366, which promotes type I IFN responses. IRF3 engagement with STING pS358 induces trafficking to late endolysosomal compartments, supporting recruitment of TRAF6 and activation of NF-κB. We identify a TRAF6 binding motif in IRF3 that facilitates recruitment of TRAF6. This work defines a signaling surface on STING and a function for IRF3 as an adaptor in immune signaling. These findings indicate that STING signaling to NF-κB is enabled only within a short time window between exit from the Golgi and lysosomal degradation, possibly limiting inflammation under homeostatic and danger-sensing conditions.

UR - https://www.scopus.com/pages/publications/105016716720

U2 - 10.1038/s41590-025-02283-8

DO - 10.1038/s41590-025-02283-8

M3 - Journal article

C2 - 40973797

SN - 1529-2908

JO - Nature Immunology

JF - Nature Immunology

ER -

STING signals to NF-κB from late endolysosomal compartments using IRF3 as an adaptor

Abstract

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