ATM regulates NF-κB-dependent immediate-early genes via RelA Ser 276 phosphorylation coupled to CDK9 promoter recruitment

Abstract

Ataxia-telangiectasia mutated (ATM), a member of the phosphatidylinositol 3 kinase-like kinase family, is a master regulator of the double strand DNA break-repair pathway after genotoxic stress. Here, we found ATM serves as an essential regulator of TNF-induced NF-kB pathway. We observed that TNF exposure of cells rapidly induced DNA double strand breaks and activates ATM. TNF-induced ROS promote nuclear IKKγ association with ubiquitin and its complex formation with ATM for nuclear export. Activated cytoplasmic ATM is involved in the selective recruitment of the E3-ubiquitin ligase β-TrCP to phospho-IκBα proteosomal degradation. Importantly, ATM binds and activates the catalytic subunit of protein kinase A (PKAc), ribosmal S6 kinase that controls RelA Ser 276 phosphorylation. In ATM knockdown cells, TNF-induced RelA Ser 276 phosphorylation is significantly decreased. We further observed decreased binding and recruitment of the transcriptional elongation complex containing cyclin dependent kinase-9 (CDK9; a kinase necessary for triggering transcriptional elongation) to promoters of NF-κB-dependent immediate-early cytokine genes, in ATM knockdown cells. We conclude that ATM is a nuclear damage-response signal modulator of TNF-induced NF-κB activation that plays a key scaffolding role in IκBα degradation and RelA Ser 276 phosphorylation. Our study provides a mechanistic explanation of decreased innate immune response associated with A-T mutation.

Ataxia Telangiectasia Mutated Kinase Mediates NF-κB Serine 276 Phosphorylation and Interferon Expression via the IRF7-RIG-I Amplification Loop in Paramyxovirus Infection

“…Whole-cell extracts (WCE) were prepared in radioimmunoprecipitation (RIPA) buffer as previously described (43). Nuclear and cytoplasmic extracts were fractionated as previously described (39). For Western blotting (WB), equal amounts of extracts were fractionated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membranes.…”

“…Our earlier studies documented that the nuclear ROS and DNA damage sensor, ataxia telangiectasia mutated (ATM), plays a central role in cytokine-induced RelA activation (39). Here, we investigated whether ATM plays an essential role in the antiviral IIR (Fig.…”

“…In both the DSB-induced DNA damage response and TNFsignaling pathways, ATM activation is coupled with IKK␥/ NEMO-dependent nuclear-to-cytoplasmic export (39,51). We next investigated whether RSV infection also induces IKK␥/ NEMO-dependent ATM nuclear export.…”

“…We have recently shown that intracellular ROS generated by activated tumor necrosis factor RI (TNFRI) is sensed by the nuclear phosphatidylinositol 3 (PI3) kinase ataxia-telangiectasia mutated (ATM) kinase (39). In response to TNF-induced oxidative stress, nuclear ATM is activated and translocates into the cytosol, where it forms a scaffold to engage the RS6K PKAc, promoting RelA Ser 276 phosphorylation and immediate early gene activation (39).…”

“…In response to TNF-induced oxidative stress, nuclear ATM is activated and translocates into the cytosol, where it forms a scaffold to engage the RS6K PKAc, promoting RelA Ser 276 phosphorylation and immediate early gene activation (39). In this study, we sought to determine whether the ATM pathway participates in the IIR activated by paramyxovirus virus replication (Fig.…”