ERM transactivation is up-regulated by the repression of DNA binding after the PKA phosphorylation of a consensus site at the edge of the ETS domain.
The final step of the transduction pathway is the activation of gene transcription, which is driven by kinase cascades leading to changes in the activity of many transcription factors. Among these latter, PEA3/E1AF, ER81/ETV1, and ERM, members of the well conserved PEA3 group from the Ets family are involved in these processes. We show here that protein kinase A (PKA) increases the transcriptional activity of human ERM and human ETV1, through a Ser residue situated at the edge of the ETS DNA-binding domain. PKA phosphorylation does not directly affect the ERM transactivation domains but does affect DNA binding activity. Unphosphorylated wild-type ERM bound DNA avidly, whereas after PKA phosphorylation it did so very weakly. Interestingly, S367A mutation significantly reduced the ERM-mediated transcription in the presence of the kinase, and the DNA binding of this mutant, although similar to that of unphosphorylated wild-type protein, was insensitive to PKA treatment. Mutations, which may mimic a phosphorylated serine, converted ERM from an efficient DNA-binding protein to a poor DNA binding one, with inefficiency of PKA phosphorylation. The present data clearly demonstrate a close correlation between the capacity of PKA to increase the transactivation of ERM and the drastic down-regulation of the binding of the ETS domain to the targeted DNA. What we thus demonstrate here is a relatively rare transcription activation mechanism through a decrease in DNA binding, probably by the shift of a non-active form of an Ets protein to a PKA-phosphorylated active one, which should be in a conformation permitting a transactivation domain to be active.
Baert
Jean-Luc
JL
UMR 8526 CNRS/Institut Pasteur de Lille, Institut de Biologie de Lille, BP 447, 1 rue Calmette, 59021 Lille Cedex, France.
Beaudoin
Claude
C
Coutte
Laurent
L
de Launoit
Yvan
Y
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D017364:Collagenases / Q000235:genetics / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D015398:Signal Transduction / Q000502:physiology; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation; D015854:Up-Regulation
D004268:DNA-Binding Proteins; C095239:ETV1 protein, human; C086680:ETV5 protein, human; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C065788:transcription factor PEA3; D017868:Cyclic AMP-Dependent Protein Kinases; D017364:Collagenases; C417249:collagenase 1