Involvement of DNA-PKcs in the type I IFN response to CpG-ODNs in conventional dendritic cells in TLR9-dependent or -independent manners.

Lucia Gabriele,Chi Ma,Ting Gong,Can Xin Jones,Narrissa P Spies,Wen-Ming Chu

doi:10.1371/journal.pone.0121371

Lucia Gabriele, Chi Ma + Show 4 more

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https://doi.org/10.1371/journal.pone.0121371

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Abstract

CpG-ODNs activate dendritic cells (DCs) to produce interferon alpha (IFNα) and beta (IFNβ). Previous studies demonstrated that Toll-like receptor 9 (TLR9) deficient DCs exhibited a residual IFNα response to CpG-A, indicating that yet-unidentified molecules are also involved in induction of IFNα by CpG-A. Here, we report that the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) but not Ku70 deficient BMDCs showed defective IFNα and IFNβ responses to CpG-A or CpG-B. Loss of both DNA-PKcs and TLR9 further reduced the IFNα response to CpG-A. These DNA-PKcs and TLR9 effects were mediated by their downstream Akt/mTORC1 pathway and downstream events IRAK1 and IKKα. Loss of DNA-PKcs, TLR9, MyD88 or IRAK4 impaired phosphorylation of Akt(S473), S6K, S6, IRAK1, or IKKα in BMDCs in response to CpG-ODNs. The residual IFNα and IFNβ in DNA-PKcs-deficient BMDCs were partially responsible for the induction of IL-6 and IL-12 by CpG-ODNs and their stimulatory effect was blocked by IFNAR1 neutralizing antibodies. Further analysis indicated that CpG-ODN associated with DNA-PKcs and Ku70, and induced DNA-PKcs’s interaction with TRAF3. Intriguingly, DNA-PKcs but not Ku70 expression level was reduced in TLR9-deficient BMDCs. Taken together, our data suggest that DNA-PKcs is an important mediator in the type I IFN response to CpG-ODNs in TLR9-dependent or -independent fashions.

Highlights

Bacterial and viral genomic DNA containing the CpG motif (CpG-DNA) and its analog oligodeoxynucleotides containing CpG motif (CpG-ODNs) are powerful activators of the innate immune system
We previously reported that DNA-PKcs regulated the IL-6 and IL-12 responses to CpG-ODNs in BMDCs in a time- and dose-dependent fashion[15]
We examined the IFNα and IFNβ responses to CpG-ODNs in Toll-like receptor 9 (TLR9)-deficient BMDCs

Summary

Introduction

Bacterial and viral genomic DNA containing the CpG motif (CpG-DNA) and its analog oligodeoxynucleotides containing CpG motif (CpG-ODNs) are powerful activators of the innate immune system. There are two major types of CpG-ODNs, CpG-A and CpG-B. CpG-B strongly activates DCs and macrophages to produce pro-inflammatory IL-6 and IL-12, which is critical for the Th1. Mechanisms of IFN Response to CpG-ODN in DCs response and subsequent anti-infectious and anti-tumor activities. CpG-A together with DOTAP (a lipid) triggers DCs to produce the type I IFN (IFNα and IFNβ), and CpG-B is able to induce IFNβ expression. It is known that CpG-ODNs activate TLR9, which in turn recruits the adaptor protein myeloid differentiation factor 88 (MyD88) and IL-1 receptor associated protein kinase 4 (IRAK4) leading to activation of IRAK1 and IKKα, which activate the IFN regulatory factor 7, resulting in expression of the type I IFN[2, 3]. Loss of TLR9 does not abolish the IFNα response to CpG-A, but abolishes the IFNβ response to CpG-B suggesting that an unidentified factor mediates the IFNα response to CpG-A

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