title: Quantitative analysis of the TNF-α-induced phosphoproteome reveals AEG-1/MTDH/LYRIC as an IKKβ substrate.
abstract: The inhibitor of the nuclear factor-κB (IκB) kinase (IKK) complex is a key regulator of the canonical NF-κB signalling cascade and is crucial for fundamental cellular functions, including stress and immune responses. The majority of IKK complex functions are attributed to NF-κB activation; however, there is increasing evidence for NF-κB pathway-independent signalling. Here we combine quantitative mass spectrometry with random forest bioinformatics to dissect the TNF-α-IKKβ-induced phosphoproteome in MCF-7 breast cancer cells. In total, we identify over 20,000 phosphorylation sites, of which ∼1% are regulated up on TNF-α stimulation. We identify various potential novel IKKβ substrates including kinases and regulators of cellular trafficking. Moreover, we show that one of the candidates, AEG-1/MTDH/LYRIC, is directly phosphorylated by IKKβ on serine 298. We provide evidence that IKKβ-mediated AEG-1 phosphorylation is essential for IκBα degradation as well as NF-κB-dependent gene expression and cell proliferation, which correlate with cancer patient survival in vivo.
authors: Department of Pharmacology, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Ramesh K
authors: RK
authors: Krishnan
authors: Biomolecular Mass Spectrometry, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Hendrik
authors: H
authors: Nolte
authors: Department of Pharmacology, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Tianliang
authors: T
authors: Sun
authors: Department of Pharmacology, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Harmandeep
authors: H
authors: Kaur
authors: Department of Cardiac Development and Remodelling, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Krishnamoorthy
authors: K
authors: Sreenivasan
authors: Biomolecular Mass Spectrometry, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Mario
authors: M
authors: Looso
authors: 1] Department of Pharmacology, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany [2] Medical Faculty, University of Frankfurt, Theodor-Stern-Kai 7, 60596 Frankfurt am Main, Germany.
authors: Stefan
authors: S
authors: Offermanns
authors: Biomolecular Mass Spectrometry, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Marcus
authors: M
authors: Krüger
authors: Department of Pharmacology, Max-Planck-Institute for Heart and Lung Research, Ludwigstrasse 43, 61231 Bad Nauheim, Germany.
authors: Jakub M
authors: JM
authors: Swiercz
mesh_terms: D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000187:drug effects* / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D002853:Chromatography, Liquid; D015972:Gene Expression Regulation, Neoplastic; D057809:HEK293 Cells; D006801:Humans; D051550:I-kappa B Kinase / Q000187:drug effects* / Q000378:metabolism; D020672:I-kappa B Proteins; D047468:Immunoprecipitation; D061986:MCF-7 Cells; D013058:Mass Spectrometry; D008565:Membrane Proteins; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B; D010750:Phosphoproteins; D010766:Phosphorylation / Q000187:drug effects*; D016601:RNA-Binding Proteins; D012694:Serine; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D014410:Tumor Stem Cell Assay; D015854:Up-Regulation
chemical_list: D015815:Cell Adhesion Molecules; D020672:I-kappa B Proteins; C483039:MTDH protein, human; D008565:Membrane Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D012694:Serine; D051550:I-kappa B Kinase; C496560:IKBKB protein, human