Structural study of the interaction between CBP TAZ1 domain and FOXO4 conserved region

Abstract

FOXO4 (Forkhead box O 4), a transcription factor, plays a pivotal role in various cellular processes such as metabolism, cell cycle progression, DNA damage response, and apoptosis. CBP (cAMP-response element binding protein (CREB) binding protein) is a coactivator recruited by several transcription factors, including FOXO4 at the transcription site. The interaction between FOXO4 and CBP is crucial in modulating the expression of target genes, such as p27kip. The transcription level of p27kip is regulated by the acetylation of CBP. In previous studies, it is known that the C-terminal region of FOXO4 interacts with CBP TAZ1 (transcriptional adapter zinc binding domain 1). However, detailed structural information about the FOXO4-CBP TAZ1 interaction remains unknown. To investigate this interaction and its structural details, we expressed and purified CBP TAZ1 (340-439), FOXO4 CR2C (341-378), and FOXO4 CR3 (469-505) as binding candidates. Using ITC experiments, we confirmed the binding of CBP TAZ1 to the FOXO4 CR3 and determined their binding affinities. We investigated the binding interface of CBP TAZ1 and FOXO4 CR3, by analyzing the chemical shift perturbations (CSPs) through NMR titration. Our results revealed that the binding site is the face formed by helices 1 and 3 of CBP TAZ1. Furthermore, circular dichroism (CD) experiments demonstrated that FOXO4 CR3 partially folds to α helix on binding to CBP TAZ1. To elucidate the part which experiences conformational change to α-helix, we conducted 3D NMR experiments and calculated the Chemical Shift Index (CSI) of α carbons. Our findings confirm that FOXO4 CR3 interacts with CBP TAZ1, folding to α helix put on helices 1 and 3 directions of TAZ1 based on hydrophobic and electrostatic interactions. Collectively, we provide structural and biophysical information at the molecular level about the interaction between CBP TAZ1 and FOXO4.