Aberrant epigenetic regulators control expansion of human CD34+ hematopoietic stem/progenitor cells

Abstract

Transcription is a tightly regulated process ensuring the proper expression of numerous genes regulating all aspects of cellular behavior. Transcription factors regulate multiple genes including other transcription factors that together control a highly complex gene network. The transcriptional machinery can be "hijacked" by oncogenic transcription factors, thereby leading to malignant cell transformation. Oncogenic transcription factors manipulate a variety of epigenetic control mechanisms to fulfill gene regulatory and cell transforming functions. These factors assemble epigenetic regulators at target gene promoter sequences, thereby disturbing physiological gene expression patterns. Retroviral vector technology and the availability of "healthy" human hematopoietic CD34+ progenitor cells enable the generation of pre-leukemic cell models for the analysis of aberrant human hematopoietic progenitor cell expansion mediated by leukemogenic transcription factors. This review summarizes recent findings regarding the mechanism by which leukemogenic gene products control human hematopoietic CD34+ progenitor cell expansion by disrupting the normal epigenetic program.

Keywords: HSPC; RUNX1/ETO; epigenetics; leukemia; stem/progenitor cell expansion.

Figure 1

Purification and genetic manipulation of human hematopoietic CD34+ progenitor cells. (A) Isolation of human CD34+ progenitor cells using a Ficoll gradient procedure and MACS column purification following labeling with anti-CD34 microbeads. (B) FACS analysis after MACS showing purified CD34+ cells. (C) Oncogene-mediated selection of RUNX1/ETO-expressing human progenitor cells in ex vivo culture after retroviral transduction with the indicated vectors co-expressing eGFP. RUNX1/ETO-L148D, DNA-binding defective mutant. (D) FACS analysis of CD34 surface marker expression of mock- and RUNX1/ETO-transduced human progenitor cells at week 4. (E) Cytomorphological analysis of mock- vs. RUNX1/ETO-expressing cells following the ex vivo selection process. (F) Experimental design of CD34+ cell long-term expansion analyses. PBMC, peripheral blood mononuclear cell; MACS, magnetic cell separation; MSCV, murine stem cell virus; i, IRES, internal ribosome entry side element; eGFP, enhanced green fluorescent protein; RN, retronectin; CFU, colony forming unit; LTC-ICs, long-term culture-initiating cells; td, transduction.

Figure 2

RUNX1/ETO assembles multiple epigenetic regulators. Open chromatin (left side) and chromatin modification following RUNX1/ETO DNA-binding and high molecular weight complex formation. RUNX1/ETO recruits several proteins, including N-CoR, mSIN3A, HDAC, MeCP2 and DNMT1, into a high molecular weight complex that triggers chromatin condensation (right side), thereby repressing gene expression. Locus-dependent, RUNX1/ETO recruits the co-activator p300 and mediates acetylation of RUNX1 at the two conserved lysine residues and nearby histones, thereby promoting gene transcription. Ac, acetylation; Met, methylation; CpG, CpG site; TF, basal transcription factor.