Chromatin remodeling of the bone-specific osteocalcin (OC) gene accompanies transcriptional activation during late stages of osteoblast differentiation. Defining the molecular, cellular, and physiological mechanisms that regulate osteoblast growth and differentiation, within the context of skeletal development and bone formation in vivo, is the long term goal of the NIH grant """"""""Control of Osteoblast Growth and Differentiation (R0l AR39588),"""""""" that serves as the parent grant for this FIRCA proposal. The studies proposed in this renewal will be done primarily in Chile. During the initial funding period of our collaborative program, we established mechanisms mediating nucleosome positioning and specific protein-DNA interactions on the osteocalcin gene promoter. The studies proposed in this renewal application will reveal molecular mechanisms operating in chromatin remodeling of the OC gene during osteoblast differentiation. We will be focused on establishing the components of the chromatin remodeling complexes that promote the changes in chromatin structure associated with basal and vitamin D-enhanced OC transcriptional activity. We hypothesize that BRG-1 containing complexes are responsible for alterations in OC gene chromatin structure. We will also examine the molecular mechanisms involved in targeting these chromatin-remodeling activities to the OC promoter. We hypothesize that sequence-specific transcription factors such as Cbfa1 play a major role in targeting chromatin-remodeling activities to the OC promoter We also propose that the translational positioning of nucleosomes in the OC promoter result from the activity of these targeted chromatin remodeling complexes. To test these hypotheses we will carry out studies using intact osteoblastic cell cultures as well as cell-free systems.
