Inflammatory osteolysis is governed by exacerbated osteoclastogenesis. Ample evidence points to central role of NF-κB in such pathologic responses, yet the precise mechanisms underpinning specificity of these responses remain unclear. We propose that motifs of the scaffold protein IKKγ/NEMO partly facilitate such functions. As proof-of-principle, we used site-specific mutagenesis to examine the role of NEMO in mediating RANKL-induced signaling in mouse bone marrow macrophages, known as osteoclast precursors. We identified lysine (K)270 as a target regulating RANKL signaling as K270A substitution results in exuberant osteoclastogenesis in vitro and murine inflammatory osteolysis in vivo. Mechanistically, we discovered that K270A mutation disrupts autophagy, stabilizes NEMO, and elevates inflammatory burden. Specifically, K270A directly or indirectly hinders binding of NEMO to ISG15, a ubiquitin-like protein, which we show targets the modified proteins to autophagy-mediated lysosomal degradation. Taken together, our findings suggest that NEMO serves as a toolkit to fine-tune specific signals in physiologic and pathologic conditions.
Keywords: ISG15; NEMO; RANKL; human biology; medicine; mouse; nk-kb; osteoclast; osteolysis.
The human skeleton contains over 200 bones that together act as an internal framework for the body. Over our lifetime, the body constantly removes older bone tissue from the skeleton and replaces it with new bone tissue. This “bone remodeling” also controls how bones are repaired after being damaged by injuries, disease or normal wear and tear. Cells known as osteoclasts are responsible for breaking down old bone tissue and participate in repairing damaged bone. A cellular pathway known as NF-kB signaling stimulates other cells called “bone marrow macrophages” to become osteoclasts. A certain level of NF-kB signaling is required to maintain a healthy skeleton. However, under certain inflammatory conditions, the level of NF-kB signaling becomes too high causing hyperactive osteoclasts to accumulate and inflict severe bone breakdown. This abnormal osteoclast activity leads to eroded and fragile bones and joints, as is the case in diseases such as rheumatoid arthritis and osteoporosis. Previous studies have shown that a protein called NEMO is a core component of the NF-kB signal pathway, but the precise role of NEMO in the diseased response remained unclear. Adapala, Swarnkar, Arra et al. have now used site-directed mutagenesis approach to study the role of NEMO in bone marrow macrophages in mice. The experiments showed that one specific site within the NEMO protein, referred to as lysine 270, is crucial for its role in controlling osteoclasts and the breakdown of bone tissue. Mutating NEMO at lysine 270 led to uncontrolled NF-kB signaling in the bone marrow macrophages. Further experiments showed that lysine 270 served as a sensor to allow NEMO to bind another protein called ISG15, which in turn helped to decrease NF-kB signaling and slow down the erosion of the bone. These findings suggest that site-specific targeting of NEMO, rather than inhibiting the whole NF-kB pathway, may help to reduce the symptoms of bone disease while maintaining the beneficial roles of this essential pathway. However, additional research is required to identify NEMO sites responsible for controlling the inflammatory component.
© 2020, Adapala et al.