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346 Part IV Disorders of Hematopoietic Cell Development

factors in human leukemia. In addition, acquired mutations in of T-cell acute lymphoblastic leukemia. SCL forms obligate heterodi-
RUNX1 have been identified in significant number of patients mers with ubiquitously expressed E proteins (such as E12 and E47),
with myelodysplastic syndrome, particularly those that progress which bind to E-box motifs (sequence CANNTG). It participates in
to AML. Homozygous knockout of either RUNX1 or CBF-β in multiprotein complexes that include E2A, GATA1, LMO2, LDB1,
−/−
mice is embryonic-lethal because of a complete failure of definitive and the repressor ETO-2. SCL mice die during embryogenesis as a
hematopoiesis. This is thought to arise from a defect in the ontogeny result of failure of all hematopoiesis and defective vasculogenesis.
of HSCs in the AGM region. Conditional knockout of RUNX1 However, conditional SCL knock-out models show a specific role for
in adult hematopoiesis demonstrate a specific role of RUNX1 in SCL in late stages of megakaryopoiesis and stress thrombopoiesis
megakaryocytopoiesis. BM deletion of RUNX1 in adult mice results during adult hematopoiesis. SCL-null megakaryocytes have disorga-
in up to ≈80% reduction in peripheral blood platelet numbers, nized DMSs and a reduced number of platelet granules. SCL modu-
although no bleeding diathesis. BM megakaryocytes are small, lack lates thrombopoiesis, in part, by direct transcriptional activation of
lobulated nuclei, have poorly developed demarcation membranes, NF-E2 p45.
and reduced polyploidization. These findings are reminiscent of the
abnormal “micromegakaryocytes” seen in humans with myelodysplas-
tic syndromes and myelofibrosis. Paradoxically, there is an increase Gfi-1b
in in vitro megakaryocyte colony plating efficiency, suggesting an
expansion of early MkPs. These effects are cell-autonomous. No Gfi-1b (Gfi standing for “growth factor independent”) is a member of
defects are seen in the erythroid lineage. Reduced dosage of RUNX1’s a family of hematopoietic expressed zinc finger transcription factors
essential cofactor CBF-β also perturbs megakaryocytopoiesis in vivo. that contain a unique amino terminal transcriptional repressor Snail/
Taken together, these findings indicate a specific role of RUNX1/ Gfi-1 (SNAG) domain. It represses target genes by recruitment of
CBF-β in megakaryocyte terminal maturation. histone lysine methyltransferases. Knockout of Gfi-1b in mice results
in embryonic lethality because of severe anemia. The fetal liver of
mutant mice contains erythroid and megakaryocytic progenitors that
NF-E2 p45 are blocked in their maturation. Culture of these cells in the presence
of TPO, in contrast to those from wild-type animals, generates only
NF-E2 is a heterodimeric transcription factor composed of two basic small colonies and the cells are acetylcholinesterase negative (a marker
region-leucine zipper (bZip) subunits: a hematopoietic-specific of maturing megakaryocytes in mice). They contain markedly reduced
45 kDa protein (p45) and a widely expressed 18 kDa subunit (p18). mRNA transcript levels of vWF, NF-E2 p45, c-Mpl, and GPIIb,
NF-E2 p45 is expressed in erythroid, megakaryocytic and mast cell compared with wild-type, suggesting a requirement for Gfi-1b in at
lineages. In vitro studies implicated NF-E2 p45 as a critical factor for least relatively early stages of megakaryocytopoiesis. Germline domi-
−/−
β-globin expression. Unexpectedly, NF-E2 p45 mice were found nant negative Gfi1b gene mutations have recently been reported as a
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to have only mild perturbations of the erythroid lineage. However, cause of familial gray platelet syndrome. The patients have macro-
these mice fail to produce platelets secondary to a maturational arrest thrombocytopenia, platelet dysfunction, and red blood cell anisopoi-
in the megakaryocyte lineage, and succumb to hemorrhage in the kilocytosis. Megakaryocytes from the patients are dysplastic-appearing
neonatal period. Since the initial studies, NF-E2 p45 has been rec- with some having hypolobulated nuclei and others with multiple sepa-
ognized as being a major regulator of terminal megakaryocyte matu- rated nuclei. All of the mutations described to date involve the carboxyl
−/−
ration and platelet release. Notably, although NF-E2 p45 mice are terminal zinc finger domain and result in truncated protein products.
severely thrombocytopenic, they have normal serum levels of TPO.
In addition, megakaryocytes from these animals proliferate in vivo in
response to TPO administration. These findings suggest that NF-E2 HOX-Related Genes
p45 regulates target genes independent of the action of TPO.
Several important target genes of NF-E2 p45 have been identified, Homeobox containing transcription factors (HOX factors) play
including β-1 tubulin, 3β-hydroxysteroid dehydrogenase (3β-HSD), central role in embryonic patterning and development. Several
thromboxane synthase, caspase 12, and Rab27b. β-1 tubulin is a of these factors have specific functions in megakaryopoiesis. The
megakaryocyte-restricted isoform of β-tubulin that plays a key role best evidence is for MEIS1, a homeodomain protein belonging to
in the marginal band structure of platelets and is essential for their the Transcription activator-like effector (TALE) subfamily. MEIS1
discoid shape. Deficiency of β-1 tubulin leads to spherocytic platelets. knock-out mice fail to produce megakaryocytes. Genome-wide
Heterozygosity for a polymorphism (Q43P, because of the double chromatin occupancy studies show high enrichment for binding near
nucleotide substitution AG>CC) in the human β1-tubulin gene is megakaryocyte and platelet function-specific genes. Heterodimers
present in about 11% of individuals in a Caucasian Northern Euro- of MEIS1 and the homeobox protein PBX1 have been shown to
pean population and correlates with a reduced risk for cardiovascular functionally regulate the PF4 gene in megakaryopoiesis.
disease in humans. This may be caused by alterations in platelet Other HOX genes have also been implicated in megakaryopoiesis.
structure and function. 3β-HSD catalyzes autocrine biosynthesis of Mutations in the HOXA11 gene cause a rare syndrome of congenital
estradiol within megakaryocytes and plays an important role in thrombocytopenia with radio-ulnar synostosis (CTRUS; OMIM
proplatelet formation. #605432). The described mutations reduce DNA binding and lead to
p18 (also called mafK) is a member of a family of small maf proteins impaired in vitro megakaryocyte differentiation. There are also limited
(mafF, mafG, mafK) related to the chicken v-maf oncoprotein. Knock- data suggesting a role for HOXA10 in murine megakaryopoiesis.
out of mafK, and the related mafF, in mice has no discernable pheno-
types, whereas deficiency of mafG leads to mild thrombocytopenia.
Compound mafK::mafG null mice have profound thrombocytopenia, c-Myb
phenocopying NF-E2 p45 mice. This indicates functional redundancy
26a
of the small maf family members in megakaryocytopoiesis. Carpinelli et al. performed an N-ethyl-N-Nitrosourea (ENU)
−/−
mutagenesis screen in TPO receptor mice to identify factors that
might influence thrombocytopoiesis. They identified two indepen-
SCL (TAL1) dent loss-of-function alleles of the transcription factor c-Myb (sub-
stitution of valine for aspartic acid at residue 152 within the DNA
SCL (also known as TAL1) is a member of the basic helix-loop-helix binding domain and residue 384 within the leucine zipper domain).
−/−
member of transcription factors and is expressed predominantly in Both TPO receptor and wild-type mice containing these mutations
megakaryocytic and erythroid cells. Dysregulated expression of SCL have supraphysiologic production of platelets as a result of excessive
because of chromosomal translocation is associated with certain cases megakaryocytopoiesis, at the expense of erythroid and lymphocyte