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262 Part IV: Molecular and Cellular Hematology Chapter 18: Hematopoietic Stem Cells, Progenitors, and Cytokines 263

cells derived from multiple organs. This observation also provides a All isoforms of Ikaros contain a highly conserved carboxyl-terminal
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powerful tool to identify such proteins. Such studies have also begun to activation domain and two zinc-finger domains that mediate their
identify novel genes expressed in HSCs, potentially allowing our better dimerization. However, only isoforms 1 to 3 of the six known alter-
understanding of their role in hematopoiesis. nately spliced forms contain more than three of the four N-terminal
zinc fingers required for DNA binding to the consensus DNA core motif
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TRANSCRIPTION FACTOR PROFILE GGGA. The PU.1 gene is 1 of approximately 30 members of the Ets
family of transcription factors that bind to the purine-rich sequence
An important goal of modern cell biology is to provide a molecular 5′-GGAA-3′. Genetic elimination of the Ikaros and PU.1 genes have
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explanation for the gene or sets of genes required to orchestrate specific established their critical role in commitment of HSCs to the lymphoid
developmental events. Fundamental to this process is an understand- lineage; fetal stem cells in Ikaros −/− mice fail to generate any definitive
ing of the proteins present in cells that regulate gene transcription in T- or B-lymphocyte precursors, and although thymocyte precursors
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a lineage-, ontogenic stage-, and developmental level-specific manner. can be identified postnatally, they undergo aberrant differentiation or
Unlike what is claimed for many organ-specific programs, no single fail to develop into the CD4, dendritic, and some γδT-cell subsets in
lineage-unique family of master regulators exerts executive control adult mice. Consequently, Ikaros is essential for all of lymphopoiesis
over hematopoiesis. Rather, an assemblage of specific and nonunique early during ontogeny, and for several subsets of lymphocytes later in
factors and signals converge to determine lineage and differentiation life. In a similar fashion, PU.1-deficient mice also lack any definitive
patterns. Several transcription factors have been identified in stem cell T- and B-cell precursors in their lymphoid organs at birth (and myeloid
populations or have been shown to affect stem cell differentiation into cells; see “HSC to CMP Commitment” below), and if knockout mice
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the lymphoid and myeloid lineages. In addition to transcription factors are maintained on antibiotics and survive the first 48 hours of life, they
that regulate HSC expansion, a number of epigenetic and microRNA begin to develop normal-appearing T cells 3 to 5 days later. In contrast,
changes have been identified that affect gene expression in these cells. mature B cells and macrophages remain undetectable in the older mice,
The BMI1 gene encodes a protein that forms part of a polychrome indicating absolute tissue dependence for this lineage.
group repressor complex, which represses a number of important
target genes including the cell-cycle regulator p16/INK4a, a pathway Hematopoietic Stem Cell to Common Myeloid
that regulates HSC function in normal and malignant hematopoie- Progenitor Commitment
sis. In addition, methylation can affect HSC gene expression, as the The SCL (stem cell leukemia) gene encodes one of the transcription fac-
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DNA methyltransferases DNMT3A and DNMT3B affect HSC self-re- tors responsible for the initial stages of myeloid development, a gene
newal. And microRNA (miRNA) species are regularly being identi- first identified at the site of chromosomal rearrangement in a patient
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fied that regulate transcription and translation of critical HSC genes. with SCL. SCL belongs to the helix-loop-helix family of transcription
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For example, 9 miRNA were overexpressed and 22 downregulated in factors, which form dimers and bind DNA at consensus E-box motifs
CD34+/CD38− HSCs compared with CD34+/CD38+ cells. Among the (CANNTG). Although initially identified as a gene rearranged in
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most upregulated miRNAs in the more primitive cells was miR-520h, T-cell acute lymphocytic leukemia, an essential role for SCL in hemato-
predicted to target ATP-binding cassette, subfamily G (ABCG2) gene, poietic development was established by gene ablation studies, which
known to be involved in stem cell maintenance. Transduction of miR- revealed a complete absence of primitive blood cells and lethality in
520h into CD34+ cells increased the numbers of several progenitor cell scl−/− embryos at day 9.5 postcoitum. Consistent with this panhe-
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types (colony-forming unit–erythroid [CFU-E], burst-forming unit– matopoietic phenotype, previous studies showed that SCL is down-
erythroid [BFU-E], and colony-forming unit–granulocyte-macrophage regulated in differentiating granulocytic and monocytic progenitor
[CFU-GM]) as well as the total number of CD34+ cells (reviewed in cells and that forced expression of the gene in hematopoietic cell lines
Ref. 114). inhibits cytokine-induced granulocytic and monocytic differentia-
tion. 129,130 Consistent with their respective roles in promoting stem cell
Hematopoietic Stem Cell Self-Renewal and Expansion and mature cell survival and proliferation, SCF sustains SCL expression
Members of the Hox family of transcription factors are important regu- in primary CD34+ cells, maintaining them in an undifferentiated state,
lators of hematopoietic cell decisions, at least at the level of self-renewal/ whereas granulocyte-monocyte colony-stimulating factor (GM-CSF)
expansion, based on (1) a similar role in multiple organ systems ; (2) downregulates SCL levels and favors granulocyte and monocyte dif-
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their lineage- and differentiation-stage-specific expression pattern ferentiation. 130,131 Together, these results suggest that SCL expression is
in hematopoietic cells ; (3) disruption of their usual level or pattern required for HSC and CMP maintenance, and that down-modulation of
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of expression that leads to hematologic expansion or malignancies ; the transcription factor is essential for myeloid differentiation.
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and (4) their elimination, or elimination of the gene(s) that regulate The GATA transcription factor family contains six members
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them, which leads to significant defects in hematopoiesis. In addi- possessing a highly related DNA-binding domain composed of two
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tion, members of the extradenticle family of homeodomain-contain- conserved zinc-finger motifs. GATA1 and GATA2 are present in
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ing proteins serve as cofactors for Hox proteins, altering their cellular hematopoietic cells, GATA2 is found in the same cells as SCL, with
localization, DNA-binding affinities, and specificities. Like Hox genes, GATA1 expression restricted to latter stages of erythroid/megakaryo-
genetic elimination of some of these cofactor proteins can lead to HSC cytic (MEP) differentiation. Because genetic elimination of GATA2 is
defects. For example, Pbx1 null mice display greatly reduced numbers lethal as a result of numerous nonhematopoietic defects, and because
of CMPs, and overexpression or altered expression of MEIS1 is asso- individual hematopoietic lineage-specific knockouts have not yet been
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ciated with hematologic malignancy. 121 engineered, the role of GATA2 in early hematopoiesis is uncertain.
However, like SCL, elimination of GATA2 expression is required for
Hematopoietic Stem Cell to Common Lymphoid hematopoietic cell maturation. 133
Progenitor Commitment As noted above, numerous lines of evidence indicate that HSCs express
The Ikaros gene encodes a family of lymphoid-restricted zinc-finger the TPO receptor, c-Mpl, as best exemplified by its expression on all AA4+/
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transcription factors related to the Drosophila hunchback gene. Sca+ cells that are capable of long-term hematopoietic repopulation.
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