Surface modifications of Ti-6Al-4V discs modulate macrophage inflammatory response

Background: Transcription factors (TFs) regulate gene expression and coordinate key cellular processes, including proliferation, differentiation, and immune responses. NFATc1 is a central regulator of immune signaling, while c-Jun mediates stress and oncogenic pathways. Their cooperative DNA binding is critical for controlling complex transcriptional programs, yet existing approaches inadequately capture the real-time kinetics underlying these interactions. This study addresses the lack of dynamic characterization of cooperative NFATc1 and c-Jun DNA binding using surface plasmon resonance (SPR).
Results: Using SPR, we quantified the individual and cooperative DNA-binding kinetics of NFATc1 and c-Jun. NFATc1 binds DNA with a dissociation constant (KD) of (4.11 ±0.07) ×10− 7 M, while c-Jun shows a slightly stronger affinity KD =(1.95 ±0.03) ×10− 7 M. Not surprisingly, when forming a heterodimeric complex, the NFATc1–c-Jun binding affinity further lowers the KD =(1.63 ±0.17) ×10− 7 M, indicating cooperative interaction. More important, kinetic analysis revealed that the association rate (ka) increased more than threefold, from (2.44 ±0.10) ×105 M− 1 s− 1 to (8.29 ±0.19) ×105 M− 1 s− 1, while dissociation kinetics remained dynamic. These results demonstrate that NFATc1 facilitates c-Jun recruitment, enhancing cooperative DNA engagement. Together, the findings highlight the unique ability of SPR to resolve cooperative TF-DNA interactions with high temporal precision, providing insights not attainable through conventional techniques.
Significance: This study reveals a kinetic mechanism underlying NFATc1–c-Jun synergistic gene regulation and demonstrates the power of SPR to resolve cooperative TF–DNA interactions in real time, bridging static structural data with dynamic transcriptional regulation.