Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Epidermal Growth Factor (EGF)

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101919

Synonyms

Historical Background

Epidermal growth factor (EGF) is a growth factor best known for its ability to stimulate cell growth and proliferation. EGF was discovered as a result of the initial observations of Stanley Cohen and coworkers in the 1950s (Cohen 1986) in which the injection of extracts from mouse salivary glands into newborn mice resulted in precocious eyelid opening and tooth eruption. Subsequent studies revealed more generalized biological effects of salivary gland extracts, including enhancement of epidermal growth and keratinization. The factor mediating these effects was identified as a 6-kDa peptide of 53 amino acids and was thus named the epidermal growth factor. In later years and independent of these studies, urogastrone, long known as inhibitor of gastric acid secretion, was isolated from urine and found to be the human form of mouse EGF. EGF has now been detected in various additional body fluids and tissues including milk, urine, plasma, intestinal fluid, amniotic fluid, kidney, brain, prostate fluids, duodenum, and placenta and has diverse cellular effects ranging from mitogenesis to the promotion of hormone secretion to glucose metabolism and cell differentiation (Carpenter and Cohen 1979). For the ground-breaking work on EGF, Stanley Cohen was awarded the 1986 Nobel Prize in Physiology and Medicine together with another pioneer in growth factor biology, Rita Levi-Montalcini, who discovered the functionally distinct nerve growth factor (NGF).

EGF Family of Proteins

The EGF family of growth factors consists of at least 12 different EGF-like polypeptides that are encoded by distinct genes. Currently, the members include EGF, transforming growth factor-alpha (TGF-α), amphiregulin (AREG), epiregulin (EREG), betacellulin (BTC), heparin-binding epidermal growth factor-like growth factor (HB-EGF), epigen (EPGN), neuregulin 1–4 (NRG1–4), and teratocarcinoma-derived growth factor (Cripto-1) (Singh et al. 2016). Despite their diverse functions, common to all these growth factors is at least one repeat of a structural motif of 35–40 amino acids that forms the EGF domain. Characteristic for this domain are six conserved cysteines that are required for biological activity. The cysteines are arranged as disulfide bridges between Cys6 (C1) and Cys20 (C3), Cys14 (C2) and Cys31 (C4), and Cys33 (C5) and Cys42 (C6) with a hinge residue between C4 and C5, thereby forming a two-stranded beta-sheet (A-loop and B-loop) followed by a loop to a C-terminal short two-stranded sheet (Fig. 1). The subdomains between the cysteines found in EGF (XnCX7CX5CX10CXCX5GXRCXn; C, cysteine; G, glycine; R, arginine; X, any amino acid) can vary in amino acid sequence and length between different peptides and define receptor selectivity and affinity. The EGF domains are not limited to growth factors of the EGF family but can be found in a more or less conserved form in a large number of structurally and functionally unrelated proteins where they can mediate protein-protein interactions. The proteins with growth factor activity, however, can be distinguished by the presence of two additional conserved residues, one glycine and one arginine, within the EGF domain sequence.
Epidermal Growth Factor (EGF), Fig. 1

Epidermal growth factor (EGF). Schematic scheme of EGF mRNA (a) and pro-EGF (b). (c) Secondary structure of human EGF showing the six conserved cysteines, the hinge residue, and the conserved glycine and arginine found only in peptides with growth factor activity

Like most of its family members, EGF is synthesized as a precursor or pro-form which is critical for the control of receptor activation and allows cells to regulate the equilibrium between the cell-associated and diffusible form of the growth factor by ectodomain shedding (Fig. 2) (Sanderson et al. 2006). The gene encoding the 1207 amino acid human prepro-EGF is located on chromosome 4q25–q27 (Zeng and Harris 2014), and prepro-EGF is further processed to pro-EGF which is a glycosylated type I transmembrane protein. In addition to the EGF sequence that resides towards the carboxy terminus of the protein, pro-EGF has an additional eight EGF-like units which vary in homology to the EGF peptide and a 400 amino acid residue mid-portion that shares a 33% homology with the LDL (low-density lipoprotein) receptor. Pro-EGF undergoes proteolytic cleavage to release the growth factor in its soluble form in response to physiological stimuli. Activation of proteolytic enzymes such as ADAMs (“a disintegrin and metalloproteinase”) leads to the cleavage of pro-EGF at the juxtamembrane region, release of the ectodomain, and further maturation to the 6-kDa form of EGF which can activate its receptor in an autocrine, paracrine, or endocrine fashion. However, in contrast to HB-EGF and other members of the EGF-family, the molecular players in ectodomain shedding and possible functions specific to the cytoplasmic domain of pro-EGF or its large extracellular domain are less well characterized (Higashiyama et al. 2008). The impaired basolateral sorting of pro-EGF, which is caused by mutations in the EGF gene, causes autosomal recessive renal hypomagnesemia.
Epidermal Growth Factor (EGF), Fig. 2

Ectodomain shedding of epidermal growth factor (EGF). Extracellular stimuli lead to the activation of proteolytic enzymes that cleave pro-EGF to release the soluble ligand that can then bind to its receptor

EGF Receptors (EGFR)

The family of receptors for the EGF-related growth factors is comprised of four members, EGFR (ErbB1/HER1), ErbB2 (neu/HER2), ErbB3 (HER3), and ErbB4 (HER4). EGFR/ErbB receptors are glycosylated transmembrane proteins with an analogous structure of an extracellular ligand-binding domain, a single hydrophobic transmembrane domain, and a cytoplasmic tyrosine kinase-containing domain. While the intracellular tyrosine kinase domain is highly conserved among family members, the kinase domain of ErbB3/HER3 has several critical amino acid substitutions, and the receptor lacks kinase activity. The extracellular domains are less well conserved allowing for specific binding of the various EGF family members to their respective receptors. ErbB2/HER2 so far has no known direct ligand and undergoes heterodimerization with other family members. Thus, both ErbB2 and ErbB3 cannot signal by themselves but are able to dimerize with EGFR/ErbB1 and ErbB4 or signal as a highly mitogenic heterodimer of ErbB2 and ErbB3 (Fig. 3). EGFR/ErbBs can also be activated by ligand-independent mechanisms including G-protein-coupled receptors (GPCR), a process termed transactivation (Forrester et al. 2016), and by viruses and viral proteins (Zheng et al. 2014).
Epidermal Growth Factor (EGF), Fig. 3

Epidermal growth factor receptor family. Growth factors are grouped into families based on their ability to interact with specific EGFR/ErbB family members. X indicates the lack of a known ligand for ErbB2 or the lack of kinase activity in ErbB3

EGFR/HER1 (Dreux et al. 2006; Lemmon et al. 2014) is the primary receptor for EGF and is encoded by a gene located on the short arm of chromosome 7 (p11.2). The mature 170 kDa type 1 transmembrane protein consists of 1186 amino acid residues with a 621 amino acid highly N-glycosylated extracellular domain, a 23 amino acid transmembrane region, and a 542 amino acid cytoplasmic domain. Within this domain, the juxtamembrane region that is conserved between EGFR family members is involved in receptor downregulation, ligand-dependent internalization, and basolateral sorting. The EGFR ectodomain has four subdomains (I-IV) composed of two sets of tandem repeats with domains I and III providing ligand specificity and domains II and IV being highly enriched in cysteine regions that form several disulfide bonds. Together they create the ligand-binding pocket which in the absence of ligand is locked into an autoinhibitory confirmation thereby preventing the exposure of the dimerization motifs. Binding of EGF to EGFR or that of other members of the EGF growth factor family to their respective receptors increases the affinity of the receptor to the growth factor and results in a conformational shift leading to homodimeric or heterodimeric interactions between the different receptors and activation of the intracellular tyrosine kinase domain (Lemmon et al. 2014). Dimerization is accompanied by autophosphorylation of tyrosine residues in the cytoplasmic tails of the receptors which serve as docking stations for adaptor and signaling molecules leading to activation of the EGFR/ErbB signaling cascades. Within the cytoplasmic domain, the phosphorylation of specific tyrosine residues is linked to the binding of discrete signaling molecules thereby leading to the activation of defined intracellular signaling pathways and for termination of receptor signaling through endocytosis and targeting to lysosomes for degradation.

EGF Signaling

Cellular responses to EGF are varied and can, depending on cellular background and tissue environment, range from mitogenesis to cell motility to apoptosis or differentiation. Activation of the receptor by ligand binding leads to autophosphorylation of the receptor, the recruitment of signaling proteins, the assembly of signaling complexes, and the activation of signaling cascades (Lindsey and Langhans 2014). Most of the proteins recruited by the activated EGFR contain Src homology 2 (SH2) and phosphotyrosine-binding (PTB) domains. Many have the ability to recruit additional proteins or phospholipids to form signaling scaffolds and/or have intrinsic enzyme activities to activate downstream signaling pathways. The best known signaling cascades stimulated by EGF are the MAPK (mitogen-activated protein kinase) and the PI3K (phosphatidylinositol-3-kinase) pathways through the activation of the Src family of tyrosine kinases and Ras/Raf signaling (Fig. 4). Other well-characterized mediators of EGF signaling include PLCγ (phospholipase C-γ), PLD (phospholipase D), and STAT (signal transducer and activator of transcription) proteins as well as ROS (reactive oxygen species). Many of these signaling proteins are shared with other signaling pathways leading to an elaborate signaling network to integrate EGF signaling with other cellular stimulants. EGF signaling itself is tightly controlled and can be modulated by sequestering EGFR into subcellular compartments like caveolae or by feedback mechanisms such as PKC (protein kinase C) or PTPs (protein tyrosine phosphatases). The most prominent mechanism regulating the duration of EGFR signaling after ligand binding is receptor internalization via clathrin-mediated endocytosis (Tomas et al. 2014). After Cbl-mediated ubiquitination and Eps15-mediated binding to the AP-2 clathrin-adaptor protein complex, the activated receptor-ligand complex is shuttled along the endocytic pathway. The receptor-ligand complexes dissociate in the acidic endocytic vesicles and the ligand is degraded in the lysosomes. The receptor itself can be targeted for degradation in lysosomes or for receptor recycling to the plasma membrane. The rate of EGFR degradation can be modulated by its heterodimerization with ErbB2, leading to a more potent and prolonged signal compared to EGFR homodimers. Along the endocytic pathway, the EGF/EGFR complex still can fulfill distinct signaling functions depending on compartment-specific distribution of the receptor dimers.
Epidermal Growth Factor (EGF), Fig. 4

Epidermal growth factor signaling and crosstalk with other signaling pathways. The major signaling pathways activated by EGF are indicated. Boxes in blue indicate some of the signaling molecules that have been shown to intersect with EGF signaling

EGF in Development, Tissue Regeneration, and Tumor Formation

Much of our understanding of the developmental, physiological, and pathological roles of EGF has come from transgenic mouse models (Zeng and Harris 2014). Mice with widespread expression of human EGF have low birth weight and stunted growth and abnormalities in chondrocyte and bone development with hyperproliferation of osteoblasts. Adult male mice are sterile, but surprisingly do not develop tumors. Mice with overexpression of mouse EGF have abnormalities in fur development and behavioral changes in addition to the phenotypic changes observed with human EGF. Studies in pre-implantation embryos suggest that EGF is an important mitogenic factor during early development acting synergistically with other growth factors. However, EGF knockout mice have no overt phenotype suggesting that EGF family members are functionally redundant and can compensate for loss of EGF function. This is in stark contrast to the effect of EGFR knockout in mice that, depending on the genetic background, varies from embryonic lethality to death at birth or postnatal death with defects in bone, brain, heart, skin, and lungs and various other epithelia.

In addition to its function in embryonic stem cells during development, EGF also plays important roles in stem or progenitor cells during wound healing and tissue renewal. EGF has been shown to be important for stem cell function in a variety of tissues, including brain, heart, bone marrow, intestine, and skin and can function synergistically with other growth factors to expand stem cell populations (Zeng and Harris 2014). In combination with LIF (leukemia inhibitory factor) or CNTF (ciliary neurotrophic factor), EGF can reprogram acinar cells into insulin-producing beta-like cells in diabetic mice. In wound healing, the response to EGF is diverse and may differ between acute and chronic wounds. EGF is upregulated after acute wounding to promote epithelialization and downregulated in chronic wounds.

Most of our knowledge on EGF signaling and cancer has come from studies on the family of EGFR/ErbB receptors. Abnormal receptor expression, oncogenic EGFR mutations, decreased internalization, and other trafficking defects that lead to altered or prolonged signaling all have been shown to play a role in tumor development and progression. The complexity of the interactions between ligands of the EGF family of growth factors with the EGFR/ErbB family of receptors that can form diverse combinations of homo- and heterodimers and the observation that EGF overexpressing mice do not develop tumors point to a multifaceted relationship between EGF signaling, EGFR, and cancer. Studies on a direct role of EGF in tumor development are more limited. However, increased expression of EGF has been correlated with aggressive tumor growth and metastasis. EGF can increase tumor cell proliferation and migration through MAPK and PI3K signaling, increased expression of MMPs (matrix metalloproteases), and by promoting EGFR localization to the nucleus to activate gene transcription (Edwin et al. 2006; Tebbutt et al. 2013) which is consistent with EGF promoting oncogenesis via the activation of EGFR.

Summary

Since its isolation by Cohen and coworkers, EGF has likely become the best known growth factor in biology. Studies on EGF function, the identification of additional members of the EGF growth factor family, and the characterization of their receptors and associated signaling cascades has aided tremendously in our understanding of fundamental physiological processes such as development and tissue regeneration. Along this path, it has become clear that dysregulation of EGF function is associated with the pathogenesis of various diseases ranging from renal hypomagnesemia to cancer development. Still, the biology and regulation of EGF is not as well understood as that of some of the other members of the EGF family of growth factors.There is also a wealth of knowledge of the biological and pathological functions of EGFR, but studies show that EGF and EGFR can have biologically distinct functions. These gaps in knowledge will still have to be closed in order to develop novel therapeutic strategies that directly target EGF with high specificity and efficacy.

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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Nemours Center for Childhood Cancer ResearchNemours/Alfred I duPont Hospital for ChildrenWilmingtonUSA