Lysosomal Ca2+ Signaling is Essential for Osteoclastogenesis and Bone Remodeling

Lysosomal Ca²⁺ emerges as a critical component of receptor-evoked Ca²⁺ signaling and plays a crucial role in many lysosomal and physiological functions. Lysosomal Ca²⁺ release is mediated by the transient receptor potential (TRP) family member TRPML1, mutations that cause the lysosomal storage disease mucolipidosis type 4. Lysosomes play a key role in osteoclast function. However, nothing is known about the role of lysosomal Ca²⁺ signaling in osteoclastogenesis and bone metabolism. In this study, we addressed this knowledge gap by studying the role of lysosomal Ca²⁺ signaling in osteoclastogenesis, osteoclast and osteoblast functions, and bone homeostasis in vivo. We manipulated lysosomal Ca²⁺ signaling by acute knockdown of TRPML1, deletion of TRPML1 in mice, pharmacological inhibition of lysosomal Ca²⁺ influx, and depletion of lysosomal Ca²⁺ storage using the TRPML agonist ML-SA1. We found that knockdown and deletion of TRPML1, although it did not have an apparent effect on osteoblast differentiation and bone formation, markedly attenuated osteoclast function, RANKL-induced cytosolic Ca²⁺ oscillations, inhibited activation of NFATc1 and osteoclastogenesis-controlling genes, suppressed the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells (MNCs), and markedly reduced the differentiation of bone marrow-derived macrophages into osteoclasts. Moreover, deletion of TRPML1 resulted in enlarged lysosomes, inhibition of lysosomal secretion, and attenuated the resorptive activity of mature osteoclasts. Notably, depletion of lysosomal Ca²⁺ with ML-SA1 similarly abrogated RANKL-induced Ca²⁺ oscillations and MNC formation. Deletion of TRPML1 in mice reduced the TRAP-positive bone surfaces and impaired bone remodeling, resulting in prominent osteopetrosis. These findings demonstrate the essential role of lysosomal Ca²⁺ signaling in osteoclast differentiation and mature osteoclast function, which play key roles in bone homeostasis.