1. A Spatial and Functional Interaction of a Heterotetramer Survivin-DNA-PKcs Complex in DNA Damage Response.
- Author
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Güllülü Ö, Hehlgans S, Mayer BE, Gößner I, Petraki C, Hoffmann M, Dombrowsky MJ, Kunzmann P, Hamacher K, Strebhardt K, Fokas E, Rödel C, Münch C, and Rödel F
- Subjects
- Catalytic Domain genetics, Cell Line, Tumor, Colorectal Neoplasms pathology, DNA Breaks, Double-Stranded, DNA Repair genetics, DNA-Activated Protein Kinase genetics, HEK293 Cells, Humans, Hydrophobic and Hydrophilic Interactions, Molecular Docking Simulation, Molecular Dynamics Simulation, Multiprotein Complexes genetics, Phosphatidylinositol 3-Kinases chemistry, Phosphatidylinositol 3-Kinases genetics, Phosphatidylinositol 3-Kinases metabolism, Phosphorylation genetics, Substrate Specificity genetics, Survivin genetics, Transfection, Colorectal Neoplasms genetics, Colorectal Neoplasms metabolism, DNA Damage, DNA-Activated Protein Kinase metabolism, Multiprotein Complexes metabolism, Signal Transduction genetics, Survivin metabolism
- Abstract
Substantial evidence has shown that overexpression of the inhibitor of apoptosis protein (IAP) survivin in human tumors correlates significantly with treatment resistance and poor patient prognosis. Survivin serves as a radiation resistance factor that impacts the DNA damage response by interacting with DNA-dependent protein kinase (DNA-PKcs). However, the complexity, molecular determinants, and functional consequences of this interrelationship remain largely unknown. By applying coimmunoprecipitation and flow cytometry-based Förster resonance energy transfer assays, we demonstrated a direct involvement of the survivin baculovirus IAP repeat domain in the regulation of radiation survival and DNA repair. This survivin-mediated activity required an interaction of residues S20 and W67 with the phosphoinositide 3-kinase (PI3K) domain of DNA-PKcs. In silico molecular docking and dynamics simulation analyses, in vitro kinase assays, and large-scale mass spectrometry suggested a heterotetrameric survivin-DNA-PKcs complex that results in a conformational change within the DNA-PKcs PI3K domain. Overexpression of survivin resulted in enhanced PI3K enzymatic activity and detection of differentially abundant phosphopeptides and proteins implicated in the DNA damage response. The survivin-DNA-PKcs interaction altered the S/T-hydrophobic motif substrate specificity of DNA-PKcs with a predominant usage of S/T-P phosphorylation sites and an increase of DNA-PKcs substrates including Foxo3. These data demonstrate that survivin differentially regulates DNA-PKcs-dependent radiation survival and DNA double-strand break repair via formation of a survivin-DNA-PKcs heterotetrameric complex. SIGNIFICANCE: These findings provide insight into survivin-mediated regulation of DNA-PKcs kinase and broaden our knowledge of the impact of survivin in modulating the cellular radiation response. See related commentary by Iliakis, p. 2270 GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/9/2304/F1.large.jpg., (©2021 American Association for Cancer Research.)
- Published
- 2021
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