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1154 Part IX: Lymphocytes and Plasma Cells Chapter 74: Lymphopoiesis 1155

in this chapter focuses on genes that regulate the earliest commitment in any one of these genes have severely abnormal B-cell development;
decisions in the production of lymphoid progenitors; regulation of later however, of these genes, only EBF and Pax5 are B-cell specific within the
differentiation stages in each lineage is discussed in Chaps. 75 to 77, hematopoietic system.
respectively. Pax5 Pax5 is expressed specifically in B-lineage–committed pro-
The complex interplay between groups of genes involved in genitors and is required for normal expression of the B-lineage genes
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hematopoietic differentiation has been likened to a multidimensional CD19 and CD79a. Pax5–/– mice are blocked at the pro-B cell stage,
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network whose “regulatory space” is formed by a dynamic balance but express most early B-cell–related genes. Although Pax5 can
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between certain transcriptional regulators. Expression analysis of activate a small subset of B-lineage genes, its main function in B-cell
multiple genes in defined progenitor populations demonstrates levels of differentiation appears to be the suppression of T-cell and myeloid tran-
promiscuity at early stages of hematopoiesis, that preclude assignment scriptional programs at the murine pro-B–cell stage, thus enforcing
of any unique gene expression pattern to each stage. 106–108 As differen- commitment to the B lineage. 121,129,130 Consistent with this role, PU.1,
tiation proceeds, a more specific “genetic fingerprint” for each lineage E2A, and EBF function earlier than Pax5 in B lymphopoiesis, and forced
develops. expression of Pax5 does not rescue the B-cell defect seen in EBF–/–
mice or PU.1–/– mice. 121
Regulation of Early Lymphoid Commitment Ebf EBF (encoded by EBF1) is a helix-loop-helix zinc finger pro-
Ikaros Although no single gene has been identified as a lymphoid- tein that activates a B-lineage transcriptional program, and induces
specific master regulator, several transcription factors have been shown B lymphoid in preference to myeloid development, in part by antag-
to be essential for the early stages of lymphopoiesis. The gene Ikaros, onizing the expression of genes encoding alternative lineages such as
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which encodes a family of DNA-binding zinc finger proteins, was iden- C/EBPα (CCAAT/enhancer binding protein), Id2, and PU.1, and,
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tified in murine knockout studies as essential for all fetal lymphopoie- in part, by inducing Pax5 expression. Ebf1–/– lymphoid progenitor
sis. 109,110 However, in the postnatal setting, the role of Ikaros is more populations from mice lack the ability to generate B cells but retain the
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complex and less specific. Adult Ikaros mice completely lack B cells, ability to generate T, NK, and myeloid cells. Overexpression of EBF in
null
and although T cells are produced, their differentiation is abnormal. multipotent progenitors promotes B-cell production at the expense of
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A murine study has suggested that Ikaros is not required for the initial myeloid differentiation. EBF and E2A function cooperatively in early
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lymphomyeloid versus myeloerythroid commitment decision, and that B lymphopoiesis ; however, overexpression of EBF can rescue B-cell
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not only lymphoid differentiation, but also certain fate choices in the differentiation in E2A-deficient mice, including activation of Pax5.
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myeloerythroid pathway are affected by Ikaros. As the expression of Pax5 overexpression however cannot rescue the B-cell defect in EBF–/–
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two key lymphoid cytokine receptors, FLT3 and IL-7Rα, is dependent mice, demonstrating a critical, Pax5-independent role of EBF in early
on Ikaros, and as these markers are used to isolate murine LMPP and B-cell fate decisions.
CLP, respectively, it is still not completely clear at which exact lymphoid
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progenitor stage Ikaros exerts its effects. In addition to lymphoid Regulation of T-Cell Commitment
progenitors, Ikaros isoforms are also expressed in HSCs, and myeloid Notch Upon arrival into the thymus, multipotent progenitors from
lineages in mice 112–116 and humans. 116,117 Although Ikaros may act as a the marrow become rapidly committed to the T- and NK-cell path-
typical transcription factor in some settings, Ikaros also affects gene ways. The most important environmental cue for T-cell commitment
expression through its role in chromatin formation. 118 is delivered by the thymic epithelium in the form of the Notch ligands,
Pu.1 The transcription factor PU.1 is essential for normal B- and Delta-like 1 (DLL1) and Delta-like 4 (DLL4). Binding of one of these
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T-lymphocyte development, but its effects are highly dose dependent. ligands to the Notch 1 receptor expressed on the surface of thymo-
At high levels of PU.1, key myeloid regulatory genes are upregulated cyte precursors causes activation of intracellular Notch and a series of
and macrophage differentiation is induced preferentially over lymphoid transcriptional programs turn on to switch lineage fate toward the T
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differentiation. Low-level expression of PU.1, however, is essential for lineage at the expense of B-cell development. In mice, Notch is abso-
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lymphopoiesis. 120,121 Mice in which PU.1 is completely absent lack B cells lutely required for T-cell differentiation and proliferation, including β
and have abnormal fetal thymopoiesis. However, studies with mice in selection. Analogous to control of early B cell differentiation by E2A,
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which PU.1 is deleted specifically in B-lineage cells show that PU.1 is not Notch signaling activates a transcriptional network which includes fac-
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essential for B-cell differentiation beyond the pre-B stage. It is likely tors critical for lineage specification (GATA-3, TCF-1), and commit-
that the critical role for PU.1 in murine lymphopoiesis lies in its upregu- ment (BCL11b). However, although Notch signaling is necessary for
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lation of expression of the receptor for IL-7, which as mentioned above, murine thymopoiesis it is not sufficient for activation of the full com-
is a key cytokine in both B and T lymphopoiesis in mice. 120 plement of T-cell genes. The ability of hematopoietic progenitors to
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E2A E2A (encoded by TCF3) generates two basic helix-loop-helix respond to Notch signaling and commit to T-lineage fate depends on
proteins, E12 and E47, through differential splicing. Murine studies a balance between positive and negative regulators. Combinations of
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suggest that E2A is necessary for lymphoid priming of multipotent pro- at least four other transcription factors are required to initiate T-cell
genitors and that the E2A proteins prime expression of a number of development: PU.1, Ikaros, Runx family factors, and E2A. 124,133 In addi-
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lymphoid-associated genes. There is a dose-dependent requirement tion, leukemia-lymphoma–related factor (LRF/Pokemon, encoded by
for E2A expression in the development of LMPP and CLP. Both B- Zbtb7a) must be downregulated to allow Notch signaling to induce
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and T-lineage commitment are severely reduced in the absence of E2A, T-cell fate decisions. Notch signaling also plays important roles at
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but Ikaros and PU.1 expression are normal. 124–126 E2A affects B lympho- later stages of thymocyte differentiation. 133
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poiesis in part through upregulation of early B-cell factor (EBF) and T The effects of Notch signaling have been extensively studied in
lymphopoiesis through upregulation of expression and function of the mice, but the exact stages and processes regulated by Notch appear to
key T-cell specification factor Notch 1. 128 differ between mice and humans. For example, using in vitro studies of
human T-cell development, it appears that while Notch is essential for
Regulation of B-Cell Commitment early thymocyte proliferation, it is not required for β selection or T-cell
The transcription factors Ikaros, PU.1, E2A, EBF, and Pax5 are essential receptor αβ differentiation. 138,139 As with so much of the information
for normal B-cell differentiation. Mice that have functional deletions described in this chapter, the most important challenge that lies ahead is
Kaushansky_chapter 74_p1149-p1158.indd 1155 9/18/15 2:26 PM