MKRN1 Regulates RANKL-Induced Differentiation Via Akt and AMPK Pathways

Osteoclasts are bone-resorbing cells, and understanding the pathways involved in osteoclast differentiation and activation is essential for developing treatments for bone diseases such as osteoporosis. Since osteoclast differentiation is regulated by signaling pathways such as Akt, MAPK, and NF-κB, it is important to identify their regulators. Makorin ring finger protein 1 (MKRN1), a conserved member of the ring finger protein family, is closely linked to the regulation of Akt and AMP-activated protein kinase (AMPK), important signaling pathways in osteoclastogenesis. However, its specific role in osteoclasts has not been revealed. Mkrn1 was overexpressed using a retrovirus or silenced using small interfering RNA (siRNA) in bone marrow-derived macrophages (BMMs). Subsequently, osteoclast differentiation was induced using receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL) and assessed via tartrate-resistant acid phosphatase (TRAP) staining. Additionally, the role of Mkrn1 in osteoclastogenesis was confirmed using Mkrn1-deficient cells. Western blot analysis was employed to evaluate Mkrn1-associated Akt and AMPK activation. The bone phenotype of Mkrn1-deficient mice was investigated through bone analysis. This study revealed that Mkrn1 significantly influences RANKL-induced osteoclastogenesis. Overexpression of Mkrn1 in BMMs enhanced osteoclast differentiation by promoting Akt phosphorylation and inhibiting AMPK phosphorylation upon RANKL stimulation. Conversely, siRNA-mediated downregulation of MKRN1 reduced Akt phosphorylation and increased AMPK phosphorylation, impairing osteoclastogenesis. Furthermore, in vivo studies using Mkrn1 knockout mice revealed a phenotype with increased bone volume. Our findings establish Mkrn1 as a positive regulator of osteoclast differentiation, highlighting its potential as a therapeutic target for bone diseases characterized by excessive bone resorption.