The Bax N terminus is required for negative regulation by the mitogen-activated protein kinase kinase and Akt signaling pathways in T cells.
The Bcl-2 family proapoptotic protein, Bax, redistributes to the mitochondrion in response to varied stimuli, triggering loss of mitochondrial integrity and apoptosis. Suppression of MAPK kinase (MEK1) by the reagent UO126 in activated T cells maintained in the cytokine IL-2 disrupts cytoplasmic localization of Bax and cell survival. UO126 triggers mitochondrial translocation of ectopically expressed Bax-GFP, and both UO126 and dominant negative MEK-1 (DN-MEK1) trigger increased apoptosis in Bax-GFP-expressing T cell lines. Because inhibition of PI3K or its target Akt also triggers mitochondrial translocation of Bax in T cells and apoptosis in Bax-transfected cell lines, we generated Bax deletion mutants to identify the region(s) that confers sensitivity to regulation by MEK1 and Akt. A deletion mutant (Bax(1-171)) without the C terminus mitochondrial targeting sequence or an Akt target site (Ser(184)) localizes to the cytoplasm and triggers low level apoptosis that is enhanced by DN-Akt or DN-MEK1. A construct that lacks the first 29 aa (Bax-delta29) largely localizes to mitochondria, is highly apoptogenic, and is not inhibited by Akt or MEK1. Furthermore, Bax-delta29 overcomes IL-2-dependent survival in a T cell line, whereas Bax triggers comparatively low levels of apoptosis in these cells. Cytoplasmic localization and regulation by MEK1 and Akt are restored in a mutant deleted of the first 13 aa (Bax-delta13). Taken together, our results identify a region in the Bax N terminus that determines cellular localization regulated by MEK- and Akt-dependent signaling in T cells.
Parikh
Neha
N
National Center for Biological Sciences, New Bellary Road, Bangalore 560065, Karnataka, India.
Sade
Hadassah
H
Kurian
Leo
L
Sarin
Apurva
A
D000818:Animals; D017209:Apoptosis / Q000235:genetics / Q000276:immunology; D002470:Cell Survival / Q000235:genetics / Q000276:immunology; D003593:Cytoplasm / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017353:Gene Deletion; D006801:Humans; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000235:genetics; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors / Q000502:physiology*; D051379:Mice; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics / Q000276:immunology; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016176:T-Lymphocyte Subsets / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D051028:bcl-2-Associated X Protein
D004791:Enzyme Inhibitors; D048369:MAP Kinase Kinase 1; D010446:Peptide Fragments; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C494826:BAX protein, human; C494827:Bax protein, mouse; C494918:AKT1 protein, human