Extracellular signal-regulated kinases 1/2 are serum-stimulated "Bim(EL) kinases" that bind to the BH3-only protein Bim(EL) causing its phosphorylation and turnover.
Bim, a "BH3-only" protein, is expressed de novo following withdrawal of serum survival factors and promotes cell death. We have shown previously that activation of the ERK1/2 pathway promotes phosphorylation of Bim(EL), targeting it for degradation via the proteasome. However, the nature of the kinase responsible for Bim(EL) phosphorylation remained unclear. We now show that Bim(EL) is phosphorylated on at least three sites in response to activation of the ERK1/2 pathway. By using the peptidylprolyl isomerase, Pin1, as a probe for proline-directed phosphorylation, we show that ERK1/2-dependent phosphorylation of Bim(EL) occurs at (S/T)P motifs. ERK1/2 phosphorylates Bim(EL), but not Bim(S) or Bim(L), in vitro, and mutation of Ser(65) to alanine blocks the phosphorylation of Bim(EL) by ERK1/2 in vitro and in vivo and prevents the degradation of the protein following activation of the ERK1/2 pathway. We also find that ERK1/2, but not JNK, can physically associate with GST-Bim(EL), but not GST-Bim(L) or GST-Bim(S), in vitro. ERK1/2 also binds to full-length Bim(EL) in vivo, and we have localized a potential ERK1/2 "docking domain" lying within a 27-amino acid stretch of the Bim(EL) protein. Our findings provide new insights into the post-translational regulation of Bim(EL) and the role of the ERK1/2 pathway in cell survival signaling.
Ley
Rebecca
R
Laboratory of Molecular Signalling, Signalling Programme, The Babraham Institute, Cambridge CB2 4AT, United Kingdom. becky.ley@bbsrc.ac.uk
Ewings
Katherine E
KE
Hadfield
Kathryn
K
Howes
Elizabeth
E
Balmanno
Kathryn
K
Cook
Simon J
SJ
D017209:Apoptosis / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D004789:Enzyme Activation; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D015398:Signal Transduction
D051017:Apoptosis Regulatory Proteins; C000606775:BCL2L11 protein, human; D000072224:Bcl-2-Like Protein 11; D002352:Carrier Proteins; D008565:Membrane Proteins; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases