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Chapter 27 Granulocytopoiesis and Monocytopoiesis 329

Disruption of neutrophil transcriptional regulation is a recurring with CN tend to have milder symptoms that often manifest later
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theme in the pathogenesis of leukemia. Nearly half of patients with in life. In addition, CN responds well to lower doses of G-CSF
AML have pathognomonic translocations resulting in the fusion of and does not transform to MDS/AML. It has been hypothesized
a transcription factor with a tissue-specific gene. These transloca- that rather than being two separate disease entities, SCN and CN
tions have been shown to interfere with appropriate myeloid dif- represent two ends of the same disease spectrum.
ferentiation and emphasize the role of transcription factors in that Mutations in ELANE encoding neutrophil elastase (NE), a
process. neutrophil primary granule protein, are the most common causes
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As discussed previously, the same transcription factors that are of both SCN (35–60%) as well as CN (80–100%). To date, 73
implicated in the induction of neutrophil differentiation also direct distinct mutations of ELANE have been identified in patients with
the expression of genes encoding neutrophil-specific functional pro- SCN and CN. Although some studies have suggested a correlation
teins. The link between morphologic differentiation and synthesis between ELANE mutations and disease severity, a clear genotype–
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of neutrophil functional proteins is illustrated by the demonstration phenotype connection has not been established. While the role
that disruption of C/EBPε signaling results in SGD, associated with of ELANE mutations in contributing to CN remains unknown,
both morphologic abnormalities and increased infections attributable two schools of thought have emerged in the literature with regard
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to neutrophil functional defects. It is intriguing that C/EBPε mice to SCN. The first suggests that ELANE mutations in SCN disrupt
share these abnormalities while also demonstrating a predilection for NE function by altering its enzymatic activity. The second suggests
the development of myelodysplasia (i.e., myelodysplastic syndrome a structural rather than a functional perturbation of mutant ELANE,
[MDS]). Although the development of MDS or AML has not resulting in activation of a terminal UPR leading to a disruption of
been reported in patients with SGD, the deficiency is a rare disease neutrophil maturation and an increase in apoptosis. 12,13 Many of the
described in fewer than a dozen patients, so a tendency to develop ELANE mutations yield proteins that share a potential propensity to
MDS/AML could easily be missed. misfold, 12,14 leading to misfolded ELANE protein accumulation and
Other diseases have been linked to defects in functionally mislocalization in the cell, thus triggering the UPR with increased
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important neutrophil proteins. Abnormalities in integrin expression, apoptosis in the granulocyte compartment. It has been suggested
notably, loss of the common β-chain of the integrin receptors, result that the UPR is not activated to the same degree in CN as in SCN,
in leukocyte adhesion deficiency, and absence of any of the compo- accounting for the milder phenotype associated with CN. Typically,
nents of the reduced nicotinamide adenine dinucleotide phosphate the UPR is activated when a load of misfolded proteins entering
(NADPH) oxidase leads to chronic granulomatous disease. the ER exceeds the cell’s capacity to handle the protein load. The
Abnormalities in granule protein gene expression again underscore response to this cellular crisis involves (a) decreasing protein synthesis
the complexity of the granulocyte functional program. Congenital so as to reduce levels of the misfolded proteins; (b) increasing the
absence of many individual granule proteins, including myeloper- transcriptional activation of UPR target genes, including ones that
oxidase, LF, and transcobalamin, has been described. In the absence contribute to proper ER-protein folding such as chaperones (e.g.,
of the more global defects seen in SGD, which presumably reflect BiP); and (c) triggering ER-associated protein degradation (ERAD).
more complex abnormalities than simple protein deficiency, these If these measures are unsuccessful at reversing the ER stress, the cell is
defects tend to be incidental laboratory findings with minimal or no programmed to die. A moderate increase in levels of UPR-associated
associated pathology. proteins has been observed in both mouse and human models of
One prominent exception to that observation is the association SCN. 12,13 The UPR hypothesis however, is unable to explain why
between point mutations in the gene encoding NE (ELA2; neutrophil pharmacologic doses of G-CSF alleviate the neutropenia of both CN
elastase, ELANE) and SCN. The pathogenesis of SCN originally and SCN, although it may reflect a prominent antiapoptotic effect
was sought in studies of G-CSFR, supported by the observation of of high doses of cytokine. 15
a truncation mutation in G-CSFR in select patients with Kostmann
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syndrome (reviewed by Berliner ). It was later demonstrated that
these were acquired mutations that may predispose the patient to Role of MicroRNAs in Controlling Gene Expression
secondary AML but did not constitute the pathologic basis for the in Granulopoiesis
neutropenia itself.
MicroRNAs (miRNAs) are 18- to 24-nucleotide-long noncoding
Severe Congenital Neutropenia and the Unfolded RNAs that regulate eukaryotic gene expression in general, by binding
to specific sites in the 3′ UTR of target genes and altering expression
Protein Response (UPR) by destabilizing mRNA or blocking mRNA translation. miRNAs
are encoded in the genome and are initially transcribed by RNA
SCN is a rare BM failure syndrome characterized by maturation arrest polymerase II as long primary transcripts referred to as primary
at the promyelocyte/myelocyte stage during neutrophil maturation miRNAs (pri-miRNAs). These transcripts are recognized and pro-
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in the BM (reviewed by Coffman and Lichtinger ). Because of cessed by a ribonuclease called Drosha into 60- to 80-nucleotide
profound neutropenia, the disease was almost uniformly fatal as a intermediates called precursor miRNAs (pre-miRNAs), which are
result of bacterial infections before the advent of granulocyte colony then exported to the cytoplasm where a second ribonuclease termed
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stimulating factor (G-CSF) therapy (reviewed by Calkhoven et al ). Dicer cleaves pre-miRNAs to generate double-stranded 18- to
In addition to having an increased risk of infections, patients with 24-nucleotide-long miRNAs. The miRNAs are then incorporated
SCN treated with G-CSF also have an underlying predisposition for into the RNA-induced silencing complex or RISC, a large protein
the later development of myelodysplastic syndrome/acute myelog- complex that also contains the Argonaute or mRNA cleaving proteins.
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enous leukemia (MDS/AML) (see Lichtinger et al and references The miRNA guides the RISC complex to target complementary
therein). Although originally described as an autosomal recessive regions in the 3′ UTRs of mRNAs, leading to repression of transla-
disorder, SCN is genetically heterogeneous, with multiple modes tion or destabilization of the mRNA by deadenylation (reviewed by
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of inheritance including autosomal recessive (30% of patients), Manikandan et al ).
autosomal dominant (60% of patients), X-linked, and sporadic. An increasing body of evidence implicates miRNA activity in
Pathogenic mutations associated with SCN have been identified in mediating both normal and abnormal myelopoiesis (reviewed by
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a number of genes including HAX1, ELANE (neutrophil elastase or Pelosi et al ). MiRNAs have been shown to activate or be activated
NE, previously ELA2), GFI1, WAS, CSF3R, and G6PC3 (reviewed by by myeloid-specific transcription factors such as C/EBPα and
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Coffman ). Regardless of the mode of inheritance or gene mutation, Gfi-1. For example, mir-223 is thought to be a direct target of C/
newborns with SCN have severe neutropenia. In contrast, patients EBPα and its expression increases during granulopoiesis. Ablating
with cyclic neutropenia (CN) have regular and consistent oscillations mir-223 in mice results in the expansion of granulocyte precursor
of their circulating neutrophils, with a mean cycle of 21 days. Patients cells resulting from a cell autonomous increase in the number of