MECHANISMS OF POLARIZED SECRETION BY BONE CELLS

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DESCRIPTION (provided by applicant): Maintenance of bone mass requires the integrated activity of osteoclasts (OCs) and osteoblasts (OBs). While they are functionally distinct, both are polarized and execute their activities by regulated secretion of bone-degrading and -synthesizing molecules, respectively. Bone resorption necessitates extracellular proton transport as a result of insertion of the vacuolar H+ATPase into the bone-apposed OC plasma membrane and targeted secretion of the collagenolytic enzyme cathepsin K. Similarly, skeletal synthesis involves secretion of specific matrix proteins such as type 1 collagen onto existing bone surfaces. Thus, the exocytic capacity of bone cells is fundamental to skeletal homeostasis. Regulated secretion, also called exocytosis, requires cell polarity in which intracellular, cargo-containing vesicles are delivered, by organization of the cytoskeleton, to an exocytic plasma membrane domain, which in OCs and OBs is that apposed to bone. Upon arrival at the site of exocytosis, the cargo-containing vesicles fuse with the plasma membrane via a number of linked steps, including high affinity binding of a synaptotagmin, a family of vesicle-associated adaptors, with several plasma membrane-residing proteins. We show data that synaptotagmin VII regulates exocytosis by OCs and OBs in a cell-autonomous manner. In consequence, mice lacking synaptotagmin VII have decreased remodeling and are a model for type II or low turnover osteoporosis. Cell polarity, a pre-requisite for exocytosis, is regulated by a conserved signaling pathway wherein activated cdc42 recruits an atypical protein kinase C, which phosphorylates essential downstream targets, including atypical PKCs such as PKC;. We find that RANKL-mediated organization of the OC cytoskeleton, and the cell's capacity to resorb bone, involve activation of the cdc42/PKC; pathway. Providing additional support for this posture, mice whose OCs express increased levels of active cdc42 are osteoporotic and those lacking the small GTPase have osteopetrosis. Similarly, cell-specific deletion of PKC; generates OCs that fail to polarize or resorb bone. These observations suggest a model in which 1) PKC;, activated by cdc42, promotes OC polarization and 2) following polarization, synaptotagmin VII controls exocytosis of bone- regulating molecules by OCs. Additionally Syt VII regulates OB exocytosis. Hence, our specific aims are to determine the mechanisms by which 1) PKC; signaling promotes OC polarization and 2) synaptotagmin VII controls exocytosis by OCs and OBs. PUBLIC HEALTH RELEVANCE: Maintenance of bone mass requires optimal and coordinated function of the cells regulating this parameter. We have identified novel proteins that individually control bone turnover. We propose to identify the mechanisms by which these two proteins control bone mass, with the view of generating novel drugs to treat diseases such as osteoporosis.