KLF2 interacts with AP-1 to negatively affect osteoclast differentiation and activity

Osteoclasts function as the primary effectors of bone resorption and are essential for bone homeostasis. Krüppel-like factor 2 (KLF2) has been implicated in osteoclast differentiation, but its precise mechanism remains poorly understood. In this study, we aimed to investigate the role of KLF2 in osteoclast function by characterising osteoclastogenesis, maturation and activity and to explore the underlying biomechanism. We showed that KLF2 negatively regulates osteoclastogenesis by characterising the osteoclast number, size and protein markers; that it negatively regulates osteoclast maturation by characterising the formation of intact actin rings and punctured podosome clusters; and that it negatively regulates osteoclast activity by characterising intracellular acidification through KLF2 overexpression by lentiviral transfection and KLF2 knockdown by small interfering RNA (siRNA) transfection. The results of chromatin immunoprecipitation (ChIP) and co-immunoprecipitation (Co-IP) assays revealed that KLF2 directly interacted with c-Fos and c-Jun, which together constitute the activator protein-1 (AP-1) complex. By overexpressing KLF2 and knockdown with si-KLF2, we revealed that KLF2 mediated osteoclast function via negative regulation of the AP-1 complex. The inhibition of AP-1 activity confirmed its importance in KLF2-mediated osteoclast differentiation; Moreover, the inhibition of JNK signalling decreased the AP-1 activation induced by KLF2 knockdown, indicating the importance of the RANKL-MAPK-AP-1-NFATc1 axis in KLF2-regulated osteoclast differentiation. This study reveals a typical type of negative regulation of osteoclasts by KLF2 and provides information for potential therapeutic targets in osteolytic diseases, including osteoporosis.