Runx1 regulation of Pu.1 corepressor/coactivator exchange identifies specific molecular targets for leukemia differentiation therapy.
Gene activation requires cooperative assembly of multiprotein transcription factor-coregulator complexes. Disruption to cooperative assemblage could underlie repression of tumor suppressor genes in leukemia cells. Mechanisms of cooperation and its disruption were therefore examined for PU.1 and RUNX1, transcription factors that cooperate to activate hematopoietic differentiation genes. PU.1 is highly expressed in leukemia cells, whereas RUNX1 is frequently inactivated by mutation or translocation. Thus, coregulator interactions of Pu.1 were examined by immunoprecipitation coupled with tandem mass spectrometry/Western blot in wild-type and Runx1-deficient hematopoietic cells. In wild-type cells, the NuAT and Baf families of coactivators coimmunoprecipitated with Pu.1. Runx1 deficiency produced a striking switch to Pu.1 interaction with the Dnmt1, Sin3A, Nurd, CoRest, and B-Wich corepressor families. Corepressors of the Polycomb family, which are frequently inactivated by mutation or deletion in myeloid leukemia, did not interact with Pu.1. The most significant gene ontology association of Runx1-Pu.1 co-bound genes was with macrophages, therefore, functional consequences of altered corepressor/coactivator exchange were examined at Mcsfr, a key macrophage differentiation gene. In chromatin immunoprecipitation analyses, high level Pu.1 binding to the Mcsfr promoter was not decreased by Runx1 deficiency. However, the Pu.1-driven shift from histone repression to activation marks at this locus, and terminal macrophage differentiation, were substantially diminished. DNMT1 inhibition, but not Polycomb inhibition, in RUNX1-translocated leukemia cells induced terminal differentiation. Thus, RUNX1 and PU.1 cooperate to exchange corepressors for coactivators, and the specific corepressors recruited to PU.1 as a consequence of RUNX1 deficiency could be rational targets for leukemia differentiation therapy.
From the Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, and.
Xiaorong
X
Gu
From the Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, and.
Zhenbo
Z
Hu
From the Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, and.
Quteba
Q
Ebrahem
Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, Ohio 44195 and.
John S
JS
Crabb
From the Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, and.
Reda Z
RZ
Mahfouz
the Department of Epidemiology and Biostatistics, Case Western Reserve University, Cleveland, Ohio 44106.
Tomas
T
Radivoyevitch
Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, Ohio 44195 and.
John W
JW
Crabb
From the Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, and saunthy@ccf.org.
Yogen
Y
Saunthararajah
D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000235:genetics*; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D056970:Co-Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D050676:Core Binding Factor Alpha 2 Subunit / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D057809:HEK293 Cells; D006801:Humans; D007938:Leukemia / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000099232:Proto-Oncogene Protein Spi-1
D056970:Co-Repressor Proteins; D050676:Core Binding Factor Alpha 2 Subunit; D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; D000099232:Proto-Oncogene Protein Spi-1
Chromatin Modifying Enzymes; Enhanceosome; Enzyme Inhibitors; Epigenetics; Leukemogenesis; Protein Complexes; Protein Interactome; Transcription Coactivators; Transcription Regulation; Transcription Repressor