TRANSCRIPTIONAL REGULATION OF C-FOS

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The transcriptional regulation of c-fos will be studied as a method towards understanding the signal transduction pathways of the cell. Transcription of the c-fos is induced rapidly by growth factors and other agents that affect cell growth and differentiation. Growth factors bind to their receptors on the cell surface, yet most of he subsequent biochemical steps that lead to increased c-fos expression and ultimately cell division are not understood. Incorrect c-fos expression can cause the cancerous transformation of cells and aberrant control of the signal transduction pathways may also lead to oncogenesis. Thus, a detailed analysis of these pathways is crucial to the understanding and treatment of cancer. The key element in the c-fos promoter for serum and epidermal growth factor regulation is the serum response element (SRE). the regulation is likely to be mediated by the serum response factor (SRF) which binds specifically to SRE. Three approaches will be used analyze regulation through SRF. 1) Phosphorylation of SRF as a regulatory modification will be investigated. Since c-fos expression is induced in the presence of protein synthesis inhibitors, post-translational modifications must be involved in the regulation. SRF is phosphorylated in vivo and this modification is required for the DNA binding activity in vitro. 2) In vitro transcription systems with crude and purified components will be utilized to assay for changes n SRF's transcriptional activity. This activity appears to be regulated separately from its DNA binding activity. 3) SRF may be regulated positively or negatively by proteins which complex with it. Such proteins which stably bind to SRF will be sought by their copurification or co-immunoprecipitation with SRF. Changes in cellular calcium levels may play an important part in growth regulation. Calcium regulation of c-fos is independent of SRE and appears to utilize sequence elements downstream of the cap site. The calcium response element(s) will be mapped, nuclear factor(s) which bind to it will be identified, and the biochemical mechanism of regulation through these sequence elements and factors will be investigated.