ABSTRACT
Chronic myeloid leukemia (CML) is a myeloproliferative disorder characterized by the t(9;22) translocation coding for the chimeric protein p210 BCR-ABL. The tumor suppressor phosphatase and tensin homolog (PTEN) has recently been shown to have a critical role in the pathogenesis of CML. Nuclear localization and proper nuclear-cytoplasmic shuttling are crucial for PTEN's tumor suppressive function. In this study, we show that BCR-ABL enhances HAUSP-induced de-ubiquitination of PTEN in turn favoring its nuclear exclusion. We further demonstrate that BCR-ABL physically interacts with and phosphorylates HAUSP on tyrosine residues to trigger its activity. Importantly, we also find that PTEN delocalization induced by BCR-ABL does not occur in the leukemic stem cell compartment due to high levels of PML, a potent inhibitor of HAUSP activity toward PTEN. We therefore identify a new proto-oncogenic mechanism whereby BCR-ABL antagonizes the nuclear function of the PTEN tumor suppressor, with important therapeutic implications for the eradication of CML minimal residual disease.
Subject(s)
Cell Nucleus/metabolism , Cytoplasm/metabolism , Fusion Proteins, bcr-abl/metabolism , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/metabolism , Nuclear Proteins/metabolism , PTEN Phosphohydrolase/metabolism , Transcription Factors/metabolism , Tumor Suppressor Proteins/metabolism , Ubiquitin Thiolesterase/metabolism , Animals , Blotting, Western , Cells, Cultured , Flow Cytometry , Fluorescent Antibody Technique , Humans , Immunoenzyme Techniques , Immunoprecipitation , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Mice , Phosphorylation , Promyelocytic Leukemia Protein , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Ubiquitin-Specific Peptidase 7 , UbiquitinationABSTRACT
New, potentially tumor-specific antigens have been described in Bcr/Abl positive leukemias. Besides the main BCR/ABL hybrid fusion transcripts, a small number of transcripts derived from alternative splicing between BCR exons 1, 13, and 14 with ABL exons 4 and 5 have been identified. These variants are expressed in chronic myelogenous leukemia and acute lymphocytic leukemia patients. The transcriptional products were characterized at their C-terminus by a large amino acid portion derived from out of frame (OOF) reading of the ABL gene. This OOF peptide is expressed only in leukemic cells and has no homology with known human proteins. In order to study an in vivo model, three 39-amino acid peptides, each corresponding to a third of the whole human OOF peptide sequence, were tested for their capacity to elicit specific immune responses in HLA A2.1 transgenic mice. Peptides A and B, but not C, induced the production of specific antisera, while A and C induced the generation of specific cytotoxic T lymphocytes.
Subject(s)
Alternative Splicing , Cancer Vaccines/immunology , Frameshift Mutation/immunology , Fusion Proteins, bcr-abl/immunology , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Amino Acid Sequence , Animals , Antigens, Neoplasm/genetics , Antigens, Neoplasm/immunology , Cell Line, Tumor , Enzyme-Linked Immunosorbent Assay , Frameshift Mutation/genetics , Fusion Proteins, bcr-abl/genetics , HLA-A2 Antigen/genetics , Humans , Interferon-gamma/biosynthesis , Interferon-gamma/immunology , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/immunology , Mice , Mice, Inbred C57BL , Mice, Transgenic , Molecular Sequence Data , Peptides/genetics , Peptides/immunology , Proto-Oncogene Proteins c-bcr/genetics , Proto-Oncogene Proteins c-bcr/immunology , T-Lymphocytes, Cytotoxic/immunologyABSTRACT
Mutations in nucleophosmin (NPM) exon 12 and the resulting delocalization of NPM into the cytoplasm are the most specific and frequent cellular events in acute myeloid leukemia patients (AML) with normal karyotype. Cytoplasmatic NPM (NPMc+) is associated with responsiveness to chemotherapy and better prognosis. The activation of nuclear factor-kappaB (NF-kappaB) has been demonstrated to occur in a subset of AML patients and is thought to induce resistance to many chemotherapeutical agents. In this study, we demonstrate the increased in vitro sensitivity of NPMc+ cells to chemotherapeutical agents and their reduced NF-kappaB activity. Furthermore, we provide evidence of the interaction between NPMc+ and NF-kappaB in the cytoplasm, resulting in the sequestration and inactivation of NF-kappaB. The cytosolic localization and consequent inactivation of NF-kappaB justifies the reduced NF-kappaB DNA-binding activity observed in NPMc+ patients. These data, taken together, may provide a possible explanation for the increased rate of chemosensitivity observed among the NPMc+ patients.
