Structural characterization of the Forkhead box O4 (FOXO4):p53 complex
P04.101
R. MandalI, O. PetrvalskáII, K. KohoutováI, V. ObšilováII, T. ObšilI
IDepartment of Physical and Macromolecular Chemistry, Faculty of Science, Charles University, Prague 2, Czech Republic, IIDepartment of Structural Biology of Signaling Proteins, Division BIOCEV, Institute of Physiology of the Czech Academy of Sciences, Vestec, Czech Republic
The forkhead box O (FOXO) transcription factor plays an important role in cell cycle regulation, metabolism, stress resistance and longevity of mammals. In human, FOXO protein family consists of four members: FOXO1, FOXO3, FOXO4 and FOXO6 which share a highly conserved DNAbinding domain called the forkhead box domain. They are expressed in all mammalian tissues and their signal is regulated by the interaction with other proteins and posttranslation modification including phosphorylation, acetylation and ubiquitination. In many human cancers, FOXO4 has been considered as a tumour suppressor and involved in the interaction with several proteins including p53 to regulate apoptosis, cancer and agerelated diseases. The tetrameric p53 protein is involved in the regulation of more than 500 target genes in the biological network. Recently, it has been reported that upon ionizing radiation (IR) damage, FOXO4 interacts with p53 which promotes cellular senescence over apoptosis and maintains the senescent cell viability by blocking p53 mediated apoptosis. However, the molecular details of FOXO4:p53 interaction remain unclear due to the absence of structural data. Therefore, the main aim of this project is to investigate the structural basis of the FOXO4:p53 complex. For that reason, sedimentation velocity analytical ultracentrifugation (SVAUC), 2D 1H15N HSQC NMR spectroscopy, chemical crosslinking coupled to mass spectroscopy and molecular docking experiments were conducted to obtain the molecular insight of the FOXO4:p53 complex with various constructs of FOXO4 and p53. Experimental data suggest that both proteins interact with the binding affinity in the micromolar (μM) range and that the Nterminal segments, as well as other regions of FOXO4 DNA binding domain are involved in the interaction with p53. This work was supported by the Charles University Grant Agency (project: 251203).