The anti-apoptotic effect of Notch-1 requires p56lck-dependent, Akt/PKB-mediated signaling in T cells.
The Notch family of transmembrane receptors have been implicated in a variety of cellular decisions in different cell types. Here we investigate the mechanism underlying Notch-1-mediated anti-apoptotic function in T cells using model cell lines as the experimental system. Ectopic expression of the intracellular domain of Notch-1/activated Notch (AcN1) increases expression of anti-apoptotic proteins of the inhibitors of apoptosis (IAP) family, the Bcl-2 family, and the FLICE-like inhibitor protein (FLIP) and inhibits death triggered by multiple stimuli that activate intrinsic or extrinsic pathways of apoptosis in human and murine T cell lines. Numb inhibited the AcN1-dependent induction of anti-apoptotic proteins and anti-apoptotic function. Using pharmacological inhibitors and dominant-negative approaches, we describe a functional role for phosphatidylinositol 3-kinase (PI3K)-dependent activation of the serine-threonine kinase Akt/PKB in the regulation of AcN1-mediated anti-apoptotic function and the expression of FLIP and IAP family proteins. Using a cell line deficient for the T cell-specific, Src family protein, the tyrosine kinase p56(lck) and by reconstitution approaches we demonstrate that p56(lck) is required for the Notch-1-mediated activation of Akt/PKB function. Furthermore, the Src tyrosine kinase inhibitor, PP2, abrogated ectopically expressed AcN1-mediated anti-apoptotic function and phosphorylation of p56(lck). We present evidence that endogenous Notch-1 associates with p56(lck) and PI3K but that Akt/PKB does not co-immunoprecipitate with the Notch1.p56(lck).PI3K complex. Finally, we demonstrate that the Notch1.p56(lck).PI3K complex is present in primary T cells that have been activated in vitro and sustained in culture with the cytokine interleukin-2.
Sade
Hadassah
H
National Centre for Biological Sciences, University of Agricultural Sciences-Gandhi Krishi Vignan Kendra Campus, New Bellary Road, Bangalore 560065, Karnataka, India.
Krishna
Sudhir
S
Sarin
Apurva
A
D017209:Apoptosis; D006801:Humans; D019169:Jurkat Cells; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D051881:Receptor, Notch1; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000378:metabolism*; D014157:Transcription Factors
C497761:NOTCH1 protein, human; D011518:Proto-Oncogene Proteins; D051881:Receptor, Notch1; D011956:Receptors, Cell Surface; D014157:Transcription Factors; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt