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72 Part I Molecular and Cellular Basis of Hematology

receptors, called Frizzled (Fz), to initiate the canonical Wnt signaling grouped into four classes: (1) steroid receptors, which include recep-
transduction pathway. At the plasma membrane, binding of Wnt tors for glucocorticoids, mineralocorticoids, progesterone, androgen,
ligands to Fz receptors connect through direct binding to several and estrogen; (2) retinoid X receptor heterodimers, such as thyroid
intracellular proteins including Disheveled (Dsh), glycogen synthase receptor, retinoic acid receptor, vitamin D receptor, and peroxisome
kinase-3β (GSK3β), Axin, and adenomatous polyposis coli (APC), proliferator-activated receptors; (3) dimeric orphan receptors, such
inhibiting proteasome-mediated degradation of the transcriptional as COUPTF or HNF4; and (4) monomeric orphan receptors, such
protein β-catenin. This degradation is regulated through GSK3β- as NGFI. The cognate ligands for orphan receptors have yet to be
mediated phosphorylation of β-catenin. As a consequence, β-catenin identified.
accumulates in the cytoplasm and translocates to the nucleus, where
it interacts with transcription factors such as lymphoid enhancer-
binding factor 1 (LEF)/TCF to modulate gene expression. G Protein–Coupled Receptor and Chemokine Signaling
GPCR Signaling
Notch Signaling
The GPCR superfamily comprises a large collection of proteins,
Notch ligands are plasma single-pass transmembrane proteins named with approximately 2000 annotated genes in the human genome
Delta-like and Jagged. Thus, cells expressing the ligands are adjacent (~10% of the entire genome). GPCRs are involved in a large array
to cells expressing the Notch receptors, which are also transmembrane of physiologic functions, including platelet aggregation and leukocyte
proteins. The Notch receptor interacts with a Notch ligand on a chemotaxis. GPCRs are single polypeptides with seven-pass trans-
contacting cell; this interaction produces Notch receptor cleavage, membrane domains containing both cytoplasmic and extracellular
which releases the Notch intracellular domain (NICD). The NICD regions. Ligands for GPCRs are very diverse and include proteins
translocates to the nucleus where it binds to several DNA-binding or peptides, amino acids, lipids, and nucleotides that bind at the
proteins including CBF1/Suppressor of Hairless/LAG-1 (CSL). As cell surface where GPCRs are localized. In spite of its vast size and
a result of this interaction between NICD and CSL, changes in variety of activational ligands, the GPCR superfamily relies upon
Notch target genes occur. In contrast to the other signaling pathways three main intracellular signaling cascades for communicating recep-
discussed in this chapter that mainly function through phosphoryla- tor activation: the cyclic adenosine monophosphate (cAMP)/protein
tion, there is no amplification from the initial Notch ligand binding kinase A (PKA), the phosphatidylinositol/phospholipase C, and the
to the receptor. Moreover, this core pathway is modulated through Rho GTPase-based cascades.
auxiliary proteins that influence the response to the Notch ligand. GPCRs are coupled to a heterotrimeric G protein formed from
Among these proteins are acute myeloid leukemia 1 (AML1), dis- three unique subunits (α, β, and γ) that are membrane bound. The
coidin domain receptor family (DDR1), NECD, Notch extracellular G-α subunit contains a GTPase domain, which is capable of hydrolyz-
domain, and CBF1-interacting protein. ing GTP to GDP. When bound to GDP, the complex is functionally
inactive, with the G-α subunit remaining tightly associated with the
other subunits of the GPCR complex. Upon ligand binding to the
Hedgehog Signaling GPCR, structural conformational changes produce the release of
GDP from the heterotrimeric complex, allowing GTP to bind to the
Hedgehog (Hh) signaling is a ligand-dependent signaling pathway. G-α subunit. In this GTP-bound form, the G-α subunit dissociates
There are three different protein ligands—Sonic, Desert, and from the G-β and G-γ subunits with which it interacts. The G-α
Indian—that are secreted and produce an N-terminal active fragment. subunit then proceeds to interact with its downstream cognate targets
Indian appears to be highly expressed in hematopoietic tissue. These to affect a particular signal response, depending upon the GPCR and
ligands bind to Patch transmembrane receptors and are internalized, the specific G-α subunit isoform. Among these second-messenger
and Smoothened (a GPCR member) translocates to the plasma effectors are the cAMP/PKA pathway, ion channels, Rho GTPase,
membrane of the primary cilium and promotes activation of the Gli MAPK, PI3K, and inositol-3-phosphate/diacylglycerol (InsP3/DAG)
family of zinc finger transcription factors. Hg targets include genes pathways. In the case of the cAMP pathway, adenylate cyclase is
involved in differentiation, apoptosis, and the cell cycle. Abnormal downstream of different GPCRs (e.g., adrenergic receptors) and is
activation of Hh signaling occurs in hematologic malignancies and activated by GTP-bound G-α. Adenylate cyclase converts ATP to
maintains stem cell expansion. Because these cells are resistant to cAMP, a freely diffusible second-messenger molecule. A key effector
conventional chemotherapy, Hh antagonism is considered a plausible of intracellular cAMP is PKA, an inactive tetrameric protein complex
target in these malignancies. consisting of two regulatory and two catalytic subunits. Binding of
cAMP to the regulatory subunits causes release and activation of
the catalytic subunits, which phosphorylate different cellular targets.
Nuclear Hormone Receptor Superfamily Among them are the transcription factor cAMP-responsive element
(CREB) and several ion channels. In addition to adenylate cyclase,
Nuclear hormones include steroid hormones (sex hormones, gluco- there are other common effectors downstream of GPCRs, such as
corticoids, and mineralocorticoids), sterol hormones (vitamin D and phospholipase C, a plasma membrane–bound enzyme that cleaves
its derivatives), thyroid hormones, and retinoids. These hormones phosphatidyl inositol (PIP2) into two products and messengers:
are lipophilic and need carrier proteins to be transported in the inositol triphosphate (IP3) and diacyl glycerol (DAG). IP3 can
blood. Due to this hydrophobicity, they can diffuse across the plasma diffuse through the cytoplasm and bind receptors in the endoplasmic
membrane to reach the receptor proteins inside the cells, either in the reticulum, resulting in calcium release to the cytoplasm. Importantly,
cytoplasm or in the nucleus. These receptors are called the nuclear calcium propagates the signaling cascade through different proteins
hormone receptor (NHR) superfamily. What distinguishes this recep- such as calcineurin and nuclear factor of activated T cell (NFAT)
tor family from those discussed previously is their ability to directly transcription factors (see Fig. 7.2), which are involved in, for example,
bind to DNA and coordinate gene expression, which effectively IL-2 gene expression. DAG at the plasma membrane binds and
makes them a form of transcription factor. NHRs contain a central activates, in conjunction with calcium, protein kinase C (PKC),
DNA-binding domain, which targets the receptor to DNA sequences which will phosphorylate other downstream targets. Rho guanine
known as hormone response elements. In addition, the C-terminal nucleotide exchange factor (RhoGEF) is also a target for some G-α
part of the receptor contains a ligand-binding domain where the subunits. Binding of the G-α subunit to Rho allosterically activates
ligand or hormone binds. Upon ligand binding, NHRs control the it, causing GTP to be preferentially bound. This, in turn, allows
expression of diverse sets of genes related to the hormonal response. RhoGEFs to activate Rho kinase, which is involved in the cytoskeletal
Based on the types of ligands that they can bind, NHRs can be reorganization necessary for changes in cell shape and motility.