Enhanced osetoclast (OC) formation is a key mechanism by which estrogen (E2) deficiency induces bone loss. E2 deficiency stimulates OC formation by altering the phenotypic characteristics of mature stromal cells (CS). As a result, mature SC from ovariectomized (ovx) mice produce increased levels of macrophage colony-stimulating factor (M-CSF), a cytokine essential for the proliferation and differentiation of OC precursors. SC from ovx mice produce high M-CSF levels because of decreased binding of the transcriptional initiator Sp-1 to an overlapping Egr/Sp site in the M-CSF promoter. We have recently discovered that the transcription factor Egr-1 inhibits Sp-1 induced M-SCF gene expression without itself binding to DNA. We have also found that Egr-1 interacts directly with Sp-1 forming a novel association of Egr-1 with Sp-1, and increased availability of unbound Sp-1, capable of transactivating the M-CSF gene. Thus, E2 deficiency, by increasing Egr-1 phosphoryl ati on, decreases association between Egr-1 and Sp-1. Mass spectrometry is being developed to follow the extent and site of phosphorylation in this protein.