Protein degradation (or stabilization) #251 | | |
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Activation of feedback inhibitory (or stimulatory) loops Adaptor proteins play a key role in organizing intracel- #253 | | |
lular signaling pathways. These proteins function as molecular connectors that physically link different enzymes and promote the assembly of complexes; adaptors can be integral membrane proteins or cytosolic proteins. A typical adaptor may contain a few specific domains (e.g., SH2 or SH3) that mediate protein–protein interactions. By influ- encing the proteins that are recruited to signaling com- plexes, adaptors can determine downstream signaling events. #254 | | |
By analogy with computer networks, the protein– protein complexes can be considered nodes and the bio- chemical events feeding into or emanating from these nodes can be thought of as hubs. Signal transduction can therefore be visualized as a kind of networking phenom- enon; understanding this higher-order complexity is the province of systems biology, involving a “marriage” of biology and computation. #255 | | |
Transcription Factors #256 | | |
Most signal transduction pathways ultimately influence cellular function by modulating gene transcription through the activation and nuclear localization of tran- scription factors. Conformational changes of transcription factors (e.g., following phosphorylation) can allow their translocation into the nucleus or can expose specific DNA or protein-binding motifs. Transcription factors may drive the expression of a relatively limited set of genes or may have much more widespread effects on gene expression. Among the transcription factors that regulate the expres- sion of genes that are needed for growth are MYC and JUN, whereas a transcription factor that triggers the expression of genes that lead to growth arrest is p53. Transcription factors have a modular design, often containing domains that bind DNA and others that interact with other proteins, such as components of the RNA polymerase complex required for transcription. #257 | | |
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DNA-binding domains permit specific binding to short DNA sequences. Whereas some transcription factor binding sites are found in promoters, close to the site where transcription starts, it is now appreciated that most transcription factors bind widely throughout genomes, including to long-range regulatory elements such as enhancers. Enhancers function by looping back to gene promoters, and therefore are spatially located close to the genes that they regulate, even though it terms of genomic sequence they may appear to be far away. These insights highlight the importance of chro- #259 | | |
matin organization in regulating gene expression, both normal and pathologic. #260 | | |
For a transcription factor to induce transcription, it must also possess protein:protein interaction domains that directly or indirectly recruit histone-modifying enzymes, chromatin-remodeling complexes, and (most importantly) RNA polymerase—the large multipro- tein enzymatic complex that is responsible for RNA synthesis. #261 | | |
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A major role of growth factors is to stimulate the activity of proteins that are required for cell survival, growth and division. Growth factor activity is mediated through binding to specific receptors, ultimately influencing the expression of genes that can: #265 | | |
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Promote entry of cells into the cell cycle #267 | | |
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Relieve blocks on cell-cycle progression (thus promot- ing replication) #269 | | |
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Enhance biosynthesis of cellular components (nucleic acids, proteins, lipids, carbohydrates) required for a mother cell to give rise to two daughter cells #273 | | |
Although some growth factors are proteins that “just” stimulate cell proliferation and/or survival, it is important to remember that they also can drive a host of other activi- ties, including migration, differentiation, and synthetic capacity. Some of the important growth factors relevant to tissue regeneration and repair are listed in Table 1.1 and described further in Chapter 3. #274 | | |
Growth factors can be involved in the proliferation of cells at steady state as well as after injury, when irrevers- ibly damaged cells must be replaced. Uncontrolled proliferation can result when the growth factor activity is #275 | | |
dysregulated, or when growth factor signaling pathways are altered to become constitutively active. Thus, many growth factor pathway genes are proto-oncogenes, and gain- of-function mutations in these genes can convert them into oncogenes capable of driving unfettered cell proliferation and tumor formation. The following discussion summa- rizes selected growth factors that are involved in the important proliferative processes of tissue repair and regeneration; by virtue of their proliferative effects they can also drive tumorigenesis. Although the growth factors described here all involve receptors with intrinsic kinase activity, other growth factors may signal through each of the various pathways shown in Fig. 1.12. #276 | | |
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Epidermal growth factor and transforming growth factor-α. Both of these factors belong to the EGF family and bind to the same receptors, explaining their shared biologic activities. EGF and TGF-α are produced by macrophages and a variety of epithelial cells, and are mitogenic for hepatocytes, fibroblasts, and a host of epithelial cells. The “EGF receptor family” includes four membrane receptors with intrinsic tyrosine kinase activity; the best- characterized is EGFR1, also known as ERB-B1, or simply EGFR. EGFR1 mutations and/or amplification fre- quently occur in a number of cancers including those of the lung, head and neck, breast, and brain. The ERBB2 receptor (also known as HER2) is overexpressed in a subset of breast cancers. To treat malignancies, many of these receptors have been successfully targeted by anti- bodies and small molecule antagonists. #278 | | |
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Hepatocyte growth factor. Hepatocyte growth factor (HGF; also known as scatter factor) has mitogenic effects on hepatocytes and most epithelial cells. HGF acts as a morphogen during embryonic development (i.e., it influences the pattern of tissue differentiation), pro- motes cell migration (hence its designation as scatter factor), and enhances hepatocyte survival. HGF is pro- duced by fibroblasts and most mesenchymal cells, as well as endothelium and nonhepatocyte liver cells. It is synthesized as an inactive precursor (pro-HGF) that is proteolytically activated by serine proteases released at sites of injury. The receptor for HGF is MET, which has intrinsic tyrosine kinase activity. It is frequently overex- pressed or mutated in tumors, particularly renal and thyroid papillary carcinomas. Consequently, MET inhibitors are being evaluated as cancer therapies. #280 | | |
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Platelet-derived growth factor. PDGF is a family of several closely related proteins, each consisting of two chains (designated by pairs of letters). Three isoforms of PDGF (AA, AB, and BB) are constitutively active, while PDGF-CC and PDGF-DD must be activated by proteo- lytic cleavage. PDGF is stored in platelet granules and is released on platelet activation. Although originally isolated from platelets (hence the name), it also is pro- duced by many other cells, including activated macro- phages, endothelium, smooth muscle cells, and a variety of tumors. All PDGF isoforms exert their effects by binding to two cell surface receptors (PDGFR α and β), both having intrinsic tyrosine kinase activity. PDGF induces fibroblast, endothelial, and smooth muscle cell proliferation and matrix synthesis, and is chemotactic for these cells (and inflammatory cells), thus promoting
recruitment of the cells into areas of inflammation and tissue injury. #282 | | |
Vascular endothelial growth factor. Vascular endothelial growth factors (VEGFs)—VEGF-A, -B, -C, and -D, and PIGF (placental growth factor)—are a family of homodi- meric proteins. VEGF-A is generally referred to simply as VEGF; it is the major factor responsible for angiogen- esis, inducing blood vessel development, after injury and in tumors. In comparison, VEGF-B and PIGF are involved in embryonic vessel development, and VEGF-C and -D stimulate both angiogenesis and lymphatic development (lymphangiogenesis). VEGFs also are involved in the maintenance of endothelial cells lining mature vessels. Its expression is highest in epithelial cells adjacent to fenestrated endothelium (e.g., podo- cytes in the kidney, pigment epithelium in the retina, and choroid plexus in the brain). VEGF induces angio- genesis by promoting endothelial cell migration and proliferation (capillary sprouting), and the formation of the vascular lumina. VEGFs also induce vascular dila- tion and increase vascular permeability. As might be anticipated, hypoxia is the most important inducer of VEGF production through pathways that involve acti- vation of the transcription factor hypoxia-inducible factor (HIF-1). Other VEGF inducers—produced at sites of inflammation or wound healing—include PDGF and TGF-α. #283 | | |
VEGFs bind to a family of receptor tyrosine kinases (VEGFR-1, -2, and -3). VEGFR-2 is highly expressed in endothelium and is the most important for angiogene- sis. Antibodies against VEGF are approved for the treat- ment of several tumors such as renal and colon cancers because cancers require angiogenesis for their spread and growth. Anti-VEGF antibodies also are used in the treatment of a number of ophthalmic diseases, includ- ing: “wet” age-related macular degeneration (AMD a disorder of inappropriate angiogenesis and vascular permeability that causes adult-onset blindness); the reti- nopathy of prematurity; and the leaky vessels that lead to diabetic macular edema. Finally, increased levels of soluble versions of VEGFR-1 (s-FLT-1) in pregnant women may contribute to preeclampsia (hypertension and proteinuria) by “sopping up” the free VEGF required for maintaining normal endothelium. #284 | | |
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Fibroblast growth factor. Fibroblast growth factor (FGF) is a family of growth factors with more than 20 members. Acidic FGF (aFGF, or FGF-1) and basic FGF (bFGF, or FGF-2) are the best characterized; FGF-7 is also referred to as keratinocyte growth factor (KGF). Released FGFs associate with heparan sulfate in the ECM, which serves as a reservoir for inactive factors that can be subsequently released by proteolysis (e.g., at sites of wound healing). FGFs transduce signals through four tyrosine kinase receptors (FGFR 1–4). FGFs contribute to wound healing responses, hematopoiesis, and development; bFGF has all the activities necessary for angiogenesis as well. #287 | | |
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Transforming growth factor-β. TGF-β has three isoforms (TGF-β1, TGF-β2, and TGF-β3) that belong to a family with about 30 members, including bone morphogenetic proteins (BMPs), activins, inhibins, and Müllerian inhib- iting substance. TGF-β1 has the most widespread distri- bution, and it is more commonly referred to simply as TGF-β. It is a homodimeric protein produced by mul- tiple cell types, including platelets, endothelium, and mononuclear inflammatory cells. TGF-β is secreted as a precursor that requires proteolysis to yield the biologi- cally active protein. There are two TGF-β receptors, both with serine/threonine kinase activity that induces the phosphorylation of several downstream cytoplasmic transcription factors called Smads. Phosphorylated Smads form heterodimers with Smad4, allowing nuclear translocation and association with other DNA-binding proteins to activate or inhibit gene transcription. TGF-β produces multiple and often opposing effects depend- ing on the tissue type and concurrent signals. Agents with such multiplicity of effects are called pleiotropic, and TGF-β is “pleiotropic with a vengeance.” Primarily, TGF-β drives scar formation by stimulating matrix syn- thesis through decreased matrix metalloproteinase (MMP) activity and increased activity of tissue inhibi- tors of proteinases (TIMPs). TGF-β also applies brakes to the inflammation that accompanies wound healing
by inhibiting lymphocyte proliferation and the activity of other leukocytes. #289 | | |
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The ECM is a network of interstitial proteins that consti- tutes a significant proportion of any tissue. Cell interac- tions with ECM are critical for development and healing, as well as for maintaining normal tissue architecture (Fig. 1.13). Much more than a simple “space filler” around cells, ECM serves several key functions: #291 | | |
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Mechanical support for cell anchorage and cell migration, and maintenance of cell polarity. #293 | | |
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Control of cell proliferation, by binding and displaying growth factors and by signaling through cellular recep- tors of the integrin family. The ECM provides a depot for a variety of latent growth factors that can be acti- #295 | | |
vated within a focus of injury or inflammation. #296 | | |
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Scaffolding for tissue renewal. Because maintenance of normal tissue structure requires a basement membrane or stromal scaffold, the integrity of the basement mem- brane or the stroma of parenchymal cells is critical for the organized regeneration of tissues. Thus, ECM disruption results in defective tissue regeneration and repair, for example, cirrhosis of the liver resulting from the collapse of the hepatic stroma in various forms of hepatitis. #298 | | |
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Establishment of tissue microenvironments. The basement membrane acts as a boundary between the epithelium and underlying connective tissue; it does not just provide support to the epithelium but is also functional, for example, in the kidney, forming part of the filtration apparatus. #300 | | |
