ATM regulates NF-κB-dependent immediate-early genes via RelA Ser 276 phosphorylation coupled to CDK9 promoter recruitment.
Bottom Line: In ATM knockdown cells, TNF-induced RelA Ser 276 phosphorylation is significantly decreased.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.
Affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), 301 University Blvd, Galveston, TX 77555 USA Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX 77555, USA.Show MeSH
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Mentions: In unstimulated cells, ATM resides in the nucleus as an inactive form. Following DNA double-strand breaks (DSBs), it undergoes autophosphorylation at Ser 1981 (21). ATM activation has been reported to be coupled with nuclear export in the DSB-induced DNA repair pathway (8). Therefore, we investigated whether a similar phenomenon happens upon TNF stimulation. Nuclear (NE) and cytoplasmic extract (CE) were prepared from cells stimulated with TNF and assayed by Western immunoblot. First, Lamin B and β-tubulin were detected to characterize subcellular enrichment. Here, we observed that although the nuclear fractions stained strongly with Lamin B, the cytosolic fractions did not, indicating that CEs were largely devoid of nuclear contamination (Figure 1A). We next examined the distribution of ATM using anti-ATM Ab. Under unstimulated conditions, an ∼350 kDa ATM band is primarily located in the nucleus and present in lower abundance in the cytosol. By contrast, after 0.25 h of TNF stimulation, ATM is detected in the cytoplasmic fraction where it continues to accumulate until 1 h of stimulation (Figure 1A). From this experiment, we concluded that TNF induces ATM nuclear-to-cytoplasmic transport.
Affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), 301 University Blvd, Galveston, TX 77555 USA Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX 77555, USA.