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 transcription 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 S73 which is required for Ha-Ras-induced super-activation and phosphorylation by MAP kinase-like enzymes. Since c-Fos HOB1 has a conserved Thr residue (T232) analogous to c-Jun S73 we have proposed 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 stimulated by Ha-Ras in vivo and phosphorylated by a MAP kinase family member in vitro and that mutating T232 to Ala abolishes both functions. Collectively these results suggest that phosphorylation of the HOB1 motif increases its activation capacity. To provide direct evidence for this we change the context of c-Fos T232 to a PKA recognition site, and show that HOB1 activity is now stimulated by the catalytic subunit of PKA. This 'PKA specificity' experiment represents a novel and powerful way to analyse phosphorylation events involved in a variety of biological functions.
Bannister
A J
AJ
Wellcome/CRC Institute, Cambridge, UK.
Brown
H J
HJ
Sutherland
J A
JA
Kouzarides
T
T
D000595:Amino Acid Sequence; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002460:Cell Line; D016384:Consensus Sequence; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017354:Point Mutation / Q000502:physiology; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics / Q000378:metabolism*; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013912:Threonine / Q000502:physiology; D015533:Transcriptional Activation / Q000502:physiology*; D018631:ras Proteins / Q000502:physiology
D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011993:Recombinant Fusion Proteins; D013912:Threonine; D017868:Cyclic AMP-Dependent Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D018631:ras Proteins