Identification of trans‐acting factors interacting with the positive regulatory element spanning −148 to −124 region of c‐jun.

c‐Jun is one of the major component of the transcription factor AP‐1 and involved in regulation of the expression of genes indispensable for cell proliferation and differentiation. Earlier reports from our laboratory have identified a positive regulatory element located between −148 to −124 (Jun25) region of the c‐jun that is differentially recognized in normal and proliferating rat liver. The factor interacting with this region is present in the cytosol and undergoes post‐translational modification and nuclear translocation upon growth stimulus. To identify the factors interacting with this region, EMSA using rat liver cytosolic extract (RCE) and nuclear extract (RNE) was performed. A slow‐migrating complex was obtained with RCE when compared with that obtained with RNE. DNA‐affinity purification of factors from the RCE and RNE, and their identification by MALDI were carried out. Two protein bands of ~ 42 and 70 kDa in the affinity purified RCE were identified to beγactin and HSP70, respectively. Western blot analysis using anti‐actin antibody confirmed the ~42 kDa band to be actin. A single band of ~40 kDa obtained in the affinity purified RNE was identified to be nuclear actin. Southwestern blot analysis of the affinity purified RCE showed that only ~42 kDa band was recognized with the Jun‐25 oligonucleotides. Thus, actin directly interacts with the positive regulatory element Jun25 of the c‐jun and is likely to be involved in its positive regulation.