Activation of the ERK1/2 signaling pathway promotes phosphorylation and proteasome-dependent degradation of the BH3-only protein, Bim.
Both the ERK and phosphatidylinositol 3'-kinase (PI3K) signaling pathways can protect cells from apoptosis following withdrawal of survival factors. We have previously shown that the ERK1/2 pathway acts independently of PI3K to block expression of the BH3-only protein, BimEL, and prevent serum withdrawal-induced cell death, although the precise mechanism by which ERK reduced BimEL levels was unclear. By comparing Bim mRNA and Bim protein, expression we now show that the rapid expression of BimEL following serum withdrawal cannot be accounted for simply by increases in mRNA following inhibition of PI3K. In cells maintained in serum BimEL is a phosphoprotein. We show that activation of the ERK1/2 pathway is both necessary and sufficient to promote BimEL phosphorylation and that this leads to a substantial increase in turnover of the BimEL protein. ERK1/2-dependent degradation of BimEL proceeds via the proteasome pathway because it is blocked by proteasome inhibitors and is defective at the restrictive temperature in cells with a temperature-sensitive mutation in the E1 component of the ubiquitin-conjugating system. Finally, co-transfection of BimEL and FLAG-ubiquitin causes the accumulation of polyubiquitinated forms of Bim, and this requires the ERK1/2 pathway. Our findings provide new insights into the regulation of Bim and the role of the ERK pathway in cell survival.
Ley
Rebecca
R
Inositide Laboratory, Signalling Programme, The Babraham Institute, Cambridge CB2 4AT, United Kingdom.
Balmanno
Kathryn
K
Hadfield
Kathryn
K
Weston
Claire
C
Cook
Simon J
SJ
D000818:Animals; D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; D001769:Blood; D016466:CHO Cells; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006224:Cricetinae; D016895:Culture Media, Serum-Free; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006358:Hot Temperature; D006801:Humans; D008565:Membrane Proteins; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009154:Mutation; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction; D025801:Ubiquitin / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; C000606775:BCL2L11 protein, human; D000072224:Bcl-2-Like Protein 11; D002352:Carrier Proteins; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D009097:Multienzyme Complexes; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011480:Protease Inhibitors; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D025801:Ubiquitin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex