Identification and Characterization of a Positive Regulatory cis-element Within the Upstream Region of c-jun

Abstract

Jun oncoprotein, a component of transcription factor AP-1, is responsible for expression of multiple genes which trigger cell proliferation, differentiation and apoptosis. The diverse regulatory roles that c-Jun plays point towards its varied and complex transcriptional regulation under different physiological conditions. This could possibly involve an interplay of different trans-acting factors and multiple cis-acting elements present in the c-jun. It is, therefore, important to identify such regulatory elements that could be involved in its stringent up- or down-regulation. In the present study, transient transfection analysis with stepwise deletion variants of the upstream region (-563 to -273), linked to basal c-jun promoter in a reporter plasmid revealed the existence of multiple regulatory regions that modulated gene expression. Further delineation of the region, conferring positive regulation of transcription from the c-jun promoter, led to the identification of a cis-acting element spanning -538 to -514 region. The trans-acting factor(s) present in rat liver nuclear extract interacted specifically only in the phosphorylated form, and with a high affinity to the recognition sequence. UV cross-linking and South-western blotting, in conjunction with the analysis of affinity purified proteins interacting with this region, revealed that at least two protein factors of approximately 45 kDa and approximately 34 kDa bind specifically to this region.