The long term goal of this study is to develop safer and effective therapeutic approach for bone loss caused by
excessive osteoclast (OC) differentiation implicated in many metabolic bone diseases. The immediate goal of
this application is to understand how OC differentiation is regulated through negative signaling mediated by
transcription factors and epigenetic regulators, which may serve as novel therapeutic targets for bone diseases
such as osteoporosis. Current therapies for osteolytic diseases are hindered by lack of understanding of the
mechanisms underlying the negative regulation of OC function to prevent bone loss. Moreover, the
mechanism(s) underlying how transcriptional factors and epigenetic factors co-regulate, especially negatively
co-regulate, OC differentiation and function remain unclear. We have identified Cbx3/HP1¿. Our subsequent
immunoprecipitation analysis confirmed that C/EBPa interacts with Cbx3/HP1¿ in OCs. Notably, our preliminary
data showed that Cbx3/HP1¿ overexpression inhibited OC differentiation and activity, while Cbx3/HP1¿ silencing
enhanced OC lineage commitment and formation. Consistently, Cbx3/HP1¿-deficient mice were found to exhibit
osteoporosis-like phenotype due to enhanced OC formation and activity. Interestingly, through ChIP analysis, we
found several binding sites of Cbx3/HP1¿ on the promoters of C/EBPa regulated OC genes, NFATC1 and
C-FOS. Our RNA-seq analysis also revealed that Cbx3 deficiency in monocytes led to increased OC gene
expression. Collectively, the preliminary data indicated that Cbx3 can restrict C/EBPa-mediated OC
differentiation and activity. Based on our preliminary studies, we hypothesize that Cbx3 negatively regulates
osteoclast differentiation through interacting with C/EBPa and epigenetic factors as a result of epigenetic
modification and preventing osteoclast gene expression in bone homeostasis. We will test the hypothesis
through three specific aims. In Aim 1, we examine the function of Cbx3 in OC differentiation, skeletal
development and bone homeostasis under physiological and pathological conditions through characterization of
the phenotypes and pathomechanism in two Cbx3 CKO mouse models through loss-of-function studies. In Aim
2, we determine the role of Cbx3 in OC differentiation, skeletal development and bone homeostasis under
physiological and pathological conditions by characterizing the phenotypes and pathomechanism in Cbx3
conditional transgenic overexpression mice through gain-of-function approach. We define the molecular
mechanism underlying how Cbx3 negatively regulates OC differentiation through interacting with C/EBPa and
controlling epigenetic modification in Aim 3. The study will elucidate the mechanism(s) through which Cbx3
cooperates with transcriptional factors and other epigenetic factors to negatively regulate OC differentiation and
activity. Insights gained from this study will not only address the basic scientific question about epigenetic
regulation of gene expression in OC biology, but also will provide the foundation for the ultimate goal of
facilitating the design of safer and novel therapeutic approach for osteolytic diseases (e.g. osteoporosis).
