The -148 to -124 region of c-jun interacts with a positive regulatory factor in rat liver and enhances transcription

The c-jun gene encodes the protein Jun, a component of the essential transcription factor, AP1. Jun/AP-1 occupies a central position in signal transduction pathways as it is responsible for the induction of a number of genes in response to growth promoters. However, the exact mechanisms leading to an enhanced expression of the c-jun gene itself during proliferation, differentiation, cell growth and development are not fully understood. Cell culture studies have given some insight in the mechanisms involved in the up-regulation of c-jun expression by UV irradiation and phorbol esters. However, it is well known that transformed cells do not accurately reflect the biology of a normal cell. We now report the identification of a positive regulatory factor from normal rat liver that activates transcription from the c-jun promoter by binding to the -148 to -124 region of c-jun. Preincubation of fractionated rat liver nuclear extract with an oligonucleotide encompassing this region of the gene significantly reduced transcription from cloned c-jun promoter. In vitro transfection studies using green fluorescent protein as a reporter gene under the control of the c-jun promoter with (-148 to +53) and without (-123 to +53) this region further confirmed its role in transcription. A DNA-binding protein factor, interacting with this region of c-jun was identified from rat liver by using electrophoretic mobility shift assays. This factor binds to its recognition sequence only in the phosphorylated form and exhibits high affinity and specificity. UV cross-linking studies, South-Western analysis and affinity purification collectively indicated the factor to be ≈40 kDa and to bind to its recognition sequence as a dimer.