PHOSPHORYLATION OF THE C-FOS AND C-JUN HOB1 MOTIF STIMULATES ITS ACTIVATION CAPACITY

The c-Fos and c-Jun proteins bind an AP1 site and activate transcripti on synergistically. These two proteins have a common activation domain which has two co-operating motifs, HOB1 and HOB2. The HOB1 motif of c -Jun includes S-73 which is required for Ha-Ras-induced super-activati on and phosphorylation by MAP kinase-like enzymes. Since c-Fos HOB1 ha s a conserved Thr residue (T-232) analogous to c-Jun S-73 we have prop osed that c-Fos HOB1 will be regulated in the same way as c-Jun HOB1. Here we show that the HOB1-containing activation domain of c-Fos is st imulated by Ha-Ras in vivo and phosphorylated by a MAP kinase family m ember in vitro and that mutating T-232 to Ala abolishes both functions . Collectively these results suggest that phosphorylation of the HOB1 motif increases its activation capacity. To provide direct evidence fo r this we change the context of c-Fos T-232 to a PKA recognition site, and show that HOB1 activity is now stimulated by the catalytic subuni t of PKA. This 'PKA specificity' experiment represents a novel and pow erful way to analyse phosphorylation events involved in a variety of b iological functions.