The importance of the cytoskeleton and integrin signaling in osteoclast (OC) function has been established by ourselves and others. Loss of proteins that connect integrin-dependent attachment structures (podosomes) and the cytoskeleton, such as Src and Pyk2, results in impaired bone resorption. The potent inhibition of OCs by calcitonin (CT) is also well known. CT induces a rapid cytoskeletal change, resulting in OC retraction and loss of motility. We previously showed that CT affects the activities and phosphorylation state of several proteins that associate with the cytoskeleton or cell attachment structures or are involved in integrin signaling. Furthermore, dephosphorylation of Pyk2 by CT and ionomycin suggest that CT may be activating tyrosine phosphatase-lB via calpain, implicating two proteins that play key roles in regulating cell motility. Finally, yeast two-hybrid screening with the C-terminal domain of the CTR also yielded cytoskeletal proteins, reinforcing the idea that regulation of cytoskeletal function may be an important function of the CTR. This proposal presents a specific innovative perspective on elucidating how CT inactivates bone resorption, asking if CT indeed affects integrin signaling and cytoskeletal integrity, thereby leading to alterations in OC adhesion and motility.
The Specific Aims of this application are: (1) Characterize the effects of CT on podosome proteins (the Pyk2/Src/Cbl complex, paxiilin, Cas family members) in osteoclasts and the mechanisms involved in cross-talk between integrin signaling and CTR signaling. (2) Determine if the CTR-induced activation of calpain contributes to the modulation of cytoskeletal organization and cell adhesion and motility, identify the mechanisms by which it acts, and explore the functional role of some calpain-specific substrates, i.e., PTPIB and filamin; and (3) Characterize the interaction of the CTR with filamin, and determine how that interaction affects the CTR regulation of OC attachment and motility. Thus, this proposal can be expected to reveal as yet unappreciated features of the molecular mechanisms of osteoclast attachment and motility and bone resorption.

National Institute of Health (NIH)
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Research Project (R01)
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Orthopedics and Musculoskeletal Study Section (ORTH)
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Sharrock, William J
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Yale University
Schools of Medicine
New Haven
United States
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Coury, Fabienne; Zenger, Serhan; Stewart, Andrew K et al. (2013) SLC4A2-mediated Cl-/HCO3- exchange activity is essential for calpain-dependent regulation of the actin cytoskeleton in osteoclasts. Proc Natl Acad Sci U S A 110:2163-8
Nakajima, Arata; Sanjay, Archana; Chiusaroli, Riccardo et al. (2009) Loss of Cbl-b increases osteoclast bone-resorbing activity and induces osteopenia. J Bone Miner Res 24:1162-72
Marzia, Marilena; Chiusaroli, Riccardo; Neff, Lynn et al. (2006) Calpain is required for normal osteoclast function and is down-regulated by calcitonin. J Biol Chem 281:9745-54
Seck, Thomas; Pellegrini, Maria; Florea, Ana Maria et al. (2005) The delta e13 isoform of the calcitonin receptor forms a six-transmembrane domain receptor with dominant-negative effects on receptor surface expression and signaling. Mol Endocrinol 19:2132-44
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