FOXO3a Induces Differentiation of Bcr-Abl-transformed Cells through Transcriptional Down-regulation of Id1

Kim U Birkenkamp,Abdelkader Essafi,Kristan E Van Der Vos,Marco Da Costa,Rosaline C.-Y Hui,Frank Holstege,Leo Koenderman,Eric W.-F Lam,Paul J Coffer

doi:10.1074/jbc.m606669200

Kim U Birkenkamp, Abdelkader Essafi + Show 7 more

Open Access

https://doi.org/10.1074/jbc.m606669200

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Abstract

Leukemic transformation often requires activation of protein kinase B (PKB/c-Akt) and is characterized by increased proliferation, decreased apoptosis, and a differentiation block. PKB phosphorylates and inactivates members of the FOXO subfamily of Forkhead transcription factors. It has been suggested that hyperactivation of PKB maintains the leukemic phenotype through actively repressing FOXO-mediated regulation of specific genes. We have found expression of the transcriptional repressor Id1 (inhibitor of DNA binding 1) to be abrogated by FOXO3a activation. Inhibition of PKB activation or growth factor deprivation also resulted in strong down-regulation of Id1 promoter activity, Id1 mRNA, and protein expression. Id1 is highly expressed in Bcr-Abl-transformed K562 cells, correlating with high PKB activation and FOXO3a phosphorylation. Inhibition of Bcr-Abl by the chemical inhibitor STI571 resulted in activation of FOXO3a and down-regulation of Id1 expression. By performing chromatin immunoprecipitation assays and promoter-mutation analysis, we demonstrate that FOXO3a acts as a transcriptional repressor by directly binding to the Id1 promoter. STI571 treatment, or expression of constitutively active FOXO3a, resulted in erythroid differentiation of K562 cells, which was inhibited by ectopic expression of Id1. Taken together our data strongly suggest that high expression of Id1, through PKB-mediated inhibition of FOXO3a, is critical for maintenance of the leukemic phenotype.

Highlights

Homeostasis of the hematopoietic system requires tight control of proliferation, differentiation, and survival of progenitor cells [1,2,3,4,5]
Because the phosphatidylinositol 3-kinase (PI3K)/PKB pathway is constitutively activated in leukemic cells [23, 24], this suggests that hyperactivation of the PI3K/PKB pathway maintains the leukemic phenotype by regulating proliferation and apoptosis and by actively repressing a set of genes regulating hematopoiesis
Myeloid malignancies are often associated with mutations in upstream signaling components, such as for instance Bcr-Abl in chronic myeloid leukemia (CML), which result in constitutive activation of the PI3K/PKB pathway [7,8,9, 15]

Summary

Introduction

Homeostasis of the hematopoietic system requires tight control of proliferation, differentiation, and survival of progenitor cells [1,2,3,4,5]. Combinations of cytokines acting on a progenitor cell initiate a specific developmental program through activation of distinct downstream signal-transduction pathways [6] Interference with this highly regulated process can lead to the development of hematopoietic malignancies. Upon stimulation with growth factors or cytokines, PKB activity is induced, and it translocates to the nucleus and phosphorylates FOXOs, leading to inhibition of transcriptional activity and nuclear export [22]. Recently it has been shown that FOXO3a was constitutively phosphorylated and inactive in cell lines expressing Bcr-Abl [26, 27] This suggests that inhibition of FOXO3a transcriptional activity may be required to maintain the leukemic phenotype. Our data strongly suggest that the high expression of Id1, through a PI3K/PKB-mediated inhibition of FOXO3a, is critical for maintaining the leukemic phenotype

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