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252 Part IV: Molecular and Cellular Hematology Chapter 17: Signal Transduction Pathways 253

Another example of the capacity of adaptor proteins to translate by IL-12 and STAT6 by IL-4 and IL-13. This same STAT-mediated “lin-
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extracellular signals into intracellular physiologic change is found in eage choice” is seen in pathologic hematopoiesis. Individuals more likely
the response to TNF ligands. The capacity of receptors that bear DDs to express high levels of STAT1 who acquire the pathologic Jak2 V617F
to induce apoptosis is dependent on the binding of the adaptor pro- mutation are more likely to develop essential thrombocythemia than
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tein FADD to the cytoplasmic domain of TNFR, which then recruits polycythemia vera. Consequently, because our understanding of the
and activates the initiating caspases 8 and 10, leading to activation of entirety of downstream signals is far from complete, the cytoplasmic
the executioner caspases 3, 6, and 7 (Chap. 15). 28,29,75 This extracellu- domains of cytokine receptors bear almost no homology other than that
lar signal-mediated apoptotic pathway stands in contrast to a second, required to engage JAKs, and there already exists a modest degree of
cell-intrinsic apoptosis pathway, in which DNA damage, cell-cycle signaling specificity, it is likely that although several cytokines engage
checkpoint defects, or loss of survival factors leads to enhanced expres- overlapping sets of signaling intermediaries, each will result in a unique
sion of proapoptotic bcl family members (bax, bad, bclXs, bid). Once set of signaling events. It is almost certain that the use of unbiased
proapoptotic proteins overcome the level of antiapoptotic family mem- screens of the entirety of signaling molecules will be required to deci-
bers (bc12, BclXL), mitochondrial TM potential declines, leading to pher all the interactions induced by ligand engagement of the multi-
leakage of cytochrome c and SMAC, the former engaging the apoptotic ple receptor families described in this chapter. Such efforts have been
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protease-activating factor (APAF) adaptor, thereby activating caspase 9 described for the epidermal growth factor receptor family, and should
and, subsequently, the executioner family of caspases, the latter inhibit- be highly informative in studies of hematopoietic signaling.
ing members of the IAP family that otherwise attenuate caspase action.
It should also be noted that although these two apoptosis pathways can
be discussed as distinct entities, merging at the level of caspase 3, they SIGNALING INSULATION
interact. For example, activation of caspase 8 by TNF family members Many of the kinases and other intermediaries that play important roles in
can also cleave bid to cause mitochondrial leakage of cytochrome c, signal transduction are not absolutely substrate specific; nevertheless, they
thereby engaging the cell intrinsic pathway, serving to amplify the extra- do participate in specific pathways free from interference from other path-
cellular signal pathway to programmed cell death. ways. Perhaps the best example of this is found in the mitogen-activated
Binding of TNF family members to their receptors does not always protein kinase (MAPK) pathway. At least three major MAPK pathways
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result in apoptosis. Although there are likely many mechanisms for operate in most cells, the p42/p44 ERK (extracellular response kinase),
this finding, one is mediated by the binding of adaptors. Different TNF p38, and JNK, each of which is triggered by distinct stimuli (mitogens
family receptors employ one of six TRAFs to engage and activate IKK, such as cytokines for ERK, inflammatory mediators and hypoxia for
which leads to the release of NF-κB, a transcription factor that induces p38, and stress and noxious stimuli for JNK), but all of which eventuate
expression of several prosurvival and proliferation-associated genes. 68 in the activation of a cascade of kinases, a MAPK kinase kinase (also
termed MEKK), which phosphorylates and activates a MAPK kinase
(also termed a MEK), and finally the MAPK. The MAPKKK (MAP
SIGNALING SPECIFICITY WITHIN EACH kinase kinase kinase) for ERK1/2 is Raf-1 and the MAPKK for ERK1/2
RECEPTOR FAMILY is MEK1; the MAPKKK for p38 is MEKK1 and the MAPKK is MKK3;
and for JNK they are MEKK1 and MKK4 or MKK7, respectively.
Once a large number of receptor/cytokine systems were identified and Because each of these kinases display only limited substrate specificity
tools to study some of their downstream signaling events developed, in vitro, it would be difficult to explain how MEKK1 activation does not
it became clear that most cytokine receptors stimulate a very similar lead to ERK activation without some mechanism to insulate the signals.
cadre of signaling events as other members of the same family. For Several scaffolding proteins have now been identified that assemble spe-
example, EPO, TPO, GH, GM-CSF, IL-6, and leptin all stimulate the cific MAPKKK, MAPKK, and MAPKs. By forming complexes of the
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phosphorylation of JAK2, yet lead to quite different cellular effects. cascade on pathway-specific scaffolding molecules, signaling integrity
One theory of hematopoiesis posits that growth factors merely serve is preserved. Moreover, once the MAPK is activated, additional scaf-
to prevent apoptosis; the stochastic induction of one or another set folding molecules can link the specific MAPK to its target transcription
of transcription factors is responsible for the distinct lineage differen- factors. Additional examples of “insulating” signaling scaffolds include
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tiation events of hematopoiesis. If this is true, then overlapping sig- those for NF-κB and the TNF receptor, the B-cell antigen receptor
naling events supported by a diverse range of cytokines might not be (termed BLNK), and protein kinase C (PKC) and integrins (termed
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surprising as they would subserve the same end point, inhibition of RACKs). 91
programmed cell death. However, it is also clear that some cytokines
and extracellular stimuli induce changes in critical transcription fac- EXTINGUISHING SIGNALS
tors, and that the fate of multipotent progenitor cells can be influenced
by the cytokines to which they are exposed; if so, each cytokine would In addition to initiating signaling by extracellular ligands, the cell must
need to induce distinct signals. Careful studies of signaling events have also be able to extinguish the stimulus to prepare for additional events
supported this hypothesis. For example, JAK3 is engaged only by cytok- and to guard against continuous cell growth. Several mechanisms have
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ine receptors that use γ , and although EPO activates the same JAK been identified that extinguish the signals initiated by extracellular
C
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as TPO (JAK2), the former leads to activation of STAT5, whereas the stimuli.
latter leads to STAT1, STAT3, and STAT5 activation, which targets a
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different set of genes. Moreover, engagement of integrin α β stimulates
5 1
EPO-induced erythroid development, whereas stimulation of integrin RECEPTOR DOWN-MODULATION
α β mediates signals that inhibit erythropoiesis and enhances TPO-in- Shortly after binding to ligand, HCRs and RTKs are rapidly internal-
4 1
duced megakaryocyte growth. 80,81 Additional examples of relative sig- ized, serving to down-modulate further signaling. Receptor inter-
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naling specificity that separates sets of cytokines are the predominance nalization is dependent on membrane clathrin, which represents a
of STAT5 activation by IL-2, compared with STAT1 and STAT3 by the major mechanism of endocytosis of cell-surface proteins, and on at least
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closely related IL-21, and the almost exclusive engagement of STAT4 one element of ligand-induced signaling. The sites on hematopoietic
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