PROJECT SUMMARY Our long-term goal is to discover novel molecular-based therapies for regulating the size of bone as a means to address the need for non-surgical treatments of craniofacial malformations. Little is known regarding the precise molecular mechanisms controlling jaw size during growth and development. This lack of knowledge leaves distraction osteogenesis as one of the few available interventions to change jaw size, even though bones of the face are known to be much more difficult to manipulate with this technique than long bones. Procedures to treat jaw size disparities will greatly benefit from a broader knowledge base of molecular and cellular mechanisms that control jaw size. The objective of the current study is to build toward this goal by understanding how the cranial neural crest (CNC), which forms all the elements in the facial and jaw skeletons, regulates jaw size. To address this issue, we propose to manipulate in vivo the CNC, a highly accessible embryonic population. For example, we can transplant quail donor CNC into a duck host, which creates a chimeric quck; and we transplant duck donor CNC into the quail host, generating chimeric duail. Exploiting the divergent developmental programs of quail and duck provides a unique way to manipulate signaling between CNC and adjacent host tissues and allows discovery of CNC-dependent processes. Some groups have looked at bone deposition in controlling jaw size, but our data suggest that bone resorption by mesodermally-derived osteoclasts with participation of CNC- derived osteocytes control jaw size at later stages of development. We have previously shown that CNC controls jaw size, bone resorption is controlled by CNC and that bone resorption controls jaw size. How CNC accomplishes such a complex task, and what factors are sufficient to replicate this phenomenon, is unknown. Likely candidates may include members and targets of WNT5A/ROR2 noncanonical signaling, since they are known to play critical roles during bone resorption. Therefore, we hypothesize that by modulating WNT5A/ROR2 non-canonical signaling, CNC directs bone resorption to control jaw bone size. To test our hypothesis, we propose two complementary and highly translational Specific Aims. Specific Aim 1 will determine the mechanism by which CNC-derived osteocytes and mesodermally-derived osteoclasts act via WNT5A/ROR2 non-canonical signaling to resorb bone and regulate jaw bone size. Specific Aim 2 will determine the mechanism via which CNC acts by WNT5A/ROR2 noncanonical signaling to regulate the actions of osteoclast-mediated bone resorption to regulate jaw bone size. We will employ gain- and loss-of-function techniques to identify molecular mechanisms that endow CNC with the ability to control mandibular bone development. By determining the precise mechanism of control of bone resorption by osteoclasts and osteocytes, future studies will be able to develop accurate therapies to control lower jaw bone development.