? The objective of this study is to determine if Idl, Id2_ and Id3 proteins play an important role in BMP6-induced bone formation. BMPs belong to the TGFI3 superfamily and play an important role in skeletal development. We have conducted a comprehensive analysis of the in vitro and in vivo osteogenic activity of 14 BMPs and found that BMP6 is one of the most potent osteogenic BMPs. By determining the expression profiles of approximately 12,000 genes regulated by osteogenic BMPs (including BMP6), we have found that Idl, Id2, and Id3 helix-loop-helix (HLH) proteins are among the most significantly up-regulated genes by osteogenic BMPs. Id proteins function as a dominant-negative regulator of basic HLH (bHLH) transcription factors. While the dual functions of Id proteins (i.e., inhibiting differentiation and stimulating proliferation) are implicated in many developmental processes (e.g., myogenesis and bone morphogenesis), the actual role of Id proteins in BMP-induced osteogenesis remains obscure. We hypothesize that Idl, Id2, and Id3 genes are important early and direct targets of osteogenic BMP signaling, and that either depletion or constitutive expression of the three Id genes inhibits BMP-induced osteogenesis. BMP6 is one of the most potent osteogenic BMPs, yet it is one of the least characterized BMPs. The mechanism behind BMP6-mediated osteogenesis is poorly understood. In this application, we propose to elucidate the functional roles of Idl, Id2, and Id3 proteins in BMP6-induced bone formation by focusing on three specific aims.
The first aim i s to determine whether the Idl, Id2, and Id3 genes are early and direct targets of BMP6 signaling by characterizing Id and BMP6 expression patterns and delineating potential BMP6-responsive elements in the promoter regions of Idl, Id2, and Id3 genes.
The second aim i s to determine whether BMP6-induced in vitro osteogenic activity is inhibited when Idl, Id2, and Id3 genes are overexpressed or depleted in pre-osteoblast progenitor cells.
The third aim i s to determine whether an overexpression or depletion of Id 1, Id2, and Id3 genes inhibits BMP6-induced orthotopic bone formation in an athymic mouse model or in the Id 1/, Id2 _, and Id3 """"""""t""""""""knockout mice. At the completion of this application, we expect that findings from the proposed studies should fill in a major gap in our knowledge about whether Id 1, Id2, and Id3 play an important role in BMP6-induced osteogenesis. Furthermore, the findings from our proposed studies about BMP6 could be applicable to other osteogenic BMPs, such as BMP2. Successful execution of the proposed studies should expand our current understanding about BMP6-mediated bone formation, which could be beneficial to a possible use of BMP6 as an effective bone regeneration agent. ? ?
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