This project is a continuation of ongoing studies of the mechanism of transformation by oncogenes that act through the MAP kinase signaling pathway. During the past year we have focused our studies on DRM (Down- Regulated by Mos)/Gremlin, a gene which we initially identified and whose expression was down-regulated in cell transformed by a variety of oncogenes. We have theorized that this loss of expression may be important for the initiation or progression of specific tumors. Our current efforts have focused on characterizing the synthesis and properties of DRM and analyzing its possible function. We have demonstrated that in-vitro synthesized DRM can be cleaved and glycosylated in a microsomal membrane system, generating processed forms of the protein with mobilities identical to the forms generated in DRM producing cells in culture. Mutation of the predicted N-linked glycosylation site blocked glycosylation both in vivo and in vitro but did not affect processing or protein trafficking through the ER/Golgi. Our analysis of cells expressing either endogenous DRM, or exogenous DRM encoded by transfected expression constructs, revealed that while the bulk of the protein was cell associated, a fraction was released into the media in a soluble form. Both cell associated and soluble forms were capable of interfering with BMP-4 signaling which induced osteogenic differentiation of C2C12 cells in culture. Direct immunoprecipitation and protein crosslinking studies also demonstrated that DRM bound BMP in a relatively stable complex, suggesting a possible mechanism of DRM/Gremlin function as a BMP antagonist. Mutant non-glycosylated DRM was functional in both assays, indicating that glycosylation was not required for BMP binding. Ongoing work is focusing on the role of soluble and cell associated DRM and the ability of DRM to alter the growth properties of gliomas and glioblastoma cell lines. - mos, Oncogene, retrovirus, Tumor Suppressor, Bone Morphogenetic Proteins, MAPK, - Human Tissues, Fluids, Cells, etc.
