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DNA-PK: gatekeeper for IKKγ/NEMO nucleocytoplasmic shuttling in genotoxic stress-induced NF-kappaB activation.

Cellular and molecular life sciences : CMLS (2020-01-15)
Senad Medunjanin, Maximilian Putzier, Till Nöthen, Sönke Weinert, Thilo Kähne, Blerim Luani, Werner Zuschratter, Ruediger C Braun-Dullaeus
ABSTRACT

The transcription factors of the nuclear factor κB (NF-κB) family play a pivotal role in the cellular response to DNA damage. Genotoxic stress-induced activation of NF-κB differs from the classical canonical pathway by shuttling of the NF-κB Essential Modifier (IKKγ/NEMO) subunit through the nucleus. Here, we show that DNA-dependent protein kinase (DNA-PK), an enzyme involved in DNA double-strand break (DSB) repair, triggers the phosphorylation of NEMO by genotoxic stress, thereby enabling shuttling of NEMO through the nucleus with subsequent NF-κB activation. We identified serine 43 of NEMO as a DNA-PK phosphorylation site and point mutation of this serine to alanine led to a complete block of NF-κB activation by ionizing radiation (IR). Blockade of DNA-PK by a specific shRNA or by DNA-PKcs-deficient cells abrogated NEMO entry into the nucleus, as well. Accordingly, SUMOylation of NEMO, a prerequisite of nuclear NEMO, was abolished. Based on these observations, we propose a model in which NEMO phosphorylation by DNA-PK provides the first step in the nucleocytoplasmic trafficking of NEMO.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
NU7026, ≥98% (HPLC), solid
Sigma-Aldrich
Monoclonal Anti-IKBKG antibody produced in mouse, clone 4G9, purified immunoglobulin, buffered aqueous solution
Sigma-Aldrich
Anti-IKKγ/NEMO (C-Terminal) antibody produced in rabbit, affinity isolated antibody, buffered aqueous solution