Development - EGFR signaling pathway

EGFR signaling pathway

Epidermal growth factor receptor (EGFR) belongs to the ERBB family of receptor tyrosine kinases that contains four closely related members EGFR and ERBB2-4. They couple the binding of the extracellular growth factor ligands to intracellular signaling pathways that regulate diverse biologic responses, including proliferation, differentiation, cell motility, and survival [1].

Six ligands of EGFR are known. These are Epidermal growth factor (EGF), Amphiregulin, Transforming growth factor alpha (TGF-alpha), Betacellulin, Heparin binding EGF-like growth factor (HB-EGF), and Epiregulin [2].

ErbB2 is a unique member of the ERBB family in that it does not bind any of the known ligands with high affinity. However, it is the preferred heterodimeric partner for other EGFRs [1].

The ligand-induced receptor dimerization and subsequent autophosphorylation of distinct tyrosine residues creates docking sites for various membrane-targeted proteins. The cytoplasmic mediators that bind to EGFR phosphotyrosine residues are either the adaptor proteins, such as SHC transforming protein 1 (Shc), Growth factor receptor-bound protein 2 (GRB2), Cas-Br-M ecotropic retroviral transforming sequence (c-Cbl), Docking protein 2 (DOK2) and NCK adaptor protein 1 (NCK1), or enzymes, such as Phospholipase C gamma 1 (PLC-gamma 1), v-Src sarcoma viral oncogene homolog (c-Src) and PTK2 protein tyrosine kinase 2 (FAK1).

The adaptors Shc and GRB2 recruit the exchange factor Son of sevenless homolog 1 (SOS) and form the complex consisting of Shc, GRB2 and SOS. Activated SOS activates small GTPase v-Ha-ras Harvey rat sarcoma viral oncogene homolog (H-Ras) by its conversion from the inactive GDP-bounding state to the active GTP-bounding state. The activated H-Ras stimulates v-Raf-1 murine leukemia viral oncogene homolog 1 (c-Raf-1) / Mitogen-activated protein kinase kinase 1 and 2 (MEK1 and MEK2)/ Mitogen-activated protein kinase 1 and 3 (ERK1/2) kinase cascade that leads to activation of the transcription factors ELK1 member of ETS oncogene family (Elk-1), v-Myc myelocytomatosis viral oncogene homolog (c-Myc), and v-Fos FBJ murine osteosarcoma viral oncogene homolog (c-Fos) [3].

The adaptor DOK2 associates with the GTPase-activating protein RAS p21 protein activator 1 (p120GAP) that reinforces intrinsic GTPase activity of H-Ras, thereby inactivating H-Ras. As a result, DOK2 can attenuate activation of the EGF-stimulated mitogen-activated protein kinase (MAPK) cascade [4].

The adaptor NCK1 couples EGFR stimulation to the activation of another MAPK-cascade, the JNK kinase cascade. NCK1 recruits p21-Activated kinase 1 (PAK1). NCK1/ PAK1 complex binds Mitogen-activated protein kinase kinase kinase 10 (MLK2) and activates the JNK cascade consisting of MLK2/ Mitogen-activated protein kinase kinase 4 and 7 (MEK4 and MKK7) / Mitogen-activated protein kinase 8 and 9 (JNK1 and JNK2). The recruitment of the cascade to the activated membrane receptor localizes MLK2 on the plasma membrane where it is activated by its known upstream effectors, such as Ras-related C3 botulinum toxin substrate 1 (Rac1). Stimulation of JNK cascade results in activation of the transcription factors Elk-1, Jun oncogene (c-Jun) and some others [5]. Dual specificity phosphatases 1 and 4 (MKP-1 and MKP-2) attenuate activation of the JNK cascade [6].

The adaptor GRB2 also binds via its SH3 domain with proline-rich regions of the c-Cbl protein. c-Cbl is tyrosine-phosphorylated by tyrosine kinase upon stimulation via the EGF receptor. EGF stimulation induces the association of c-Cbl with the regulatory p85 subunit of the Phosphatidylinositol 3-kinase (PI3K reg class IA (p85)) [7].

Activated PI3K cat class IA converts Phosphatidylinositol 4,5-biphosphate (PtdIns(4,5)P2) to Phosphatidylinositol 3,4,5-triphosphate (PtdIns(3,4,5)P3). The latter is a second messenger involved in regulation of various processes [8]. PtdIns(3,4,5)P3 associates with the inner surface of the plasma membrane and promotes the recruitment of proteins with pleckstrin homology (PH) domains. One of such proteins is serine/threonine kinase v-AKT murine thymoma viral oncogene homolog (AKT). It is the essential mediator of various cell processes, such as apoptosis, cell cycle, protein synthesis, regulation of metabolism [9].

Enzymes such as PLC-gamma 1 or the cytoplasmic tyrosine kinase c-Src tie EGFR activation to the generation of secondary messengers and calcium metabolism or to mitogenic signaling cascades, respectively. EGFR recruits and phosphorylates PLC-gamma 1 [10]. Phosphorylated PLC-gamma 1 generates Diacylglycerol (DAG) and Inositol-1,4,5-trisphosphate (IP3) from PtdIns(4,5)P2 [11]. DAG activates many isoforms of Protein kinase C (PKC), including conventional isoforms alpha, beta, and gamma (PKC-alpha, PKC-beta, and PKC-gamma), as well as PKC-epsilon and PKC-theta. PKC-alpha, PKC-beta, PKC-gamma, and PKC-epsilon phosphorylate and activate c-Raf-1, thereby amplifying H-Ras/ MEK1 and MEK2/ ERK1/2 kinase cascade [12], [13]. PKC-theta activates Nuclear factor NF-kappa-B inhibitor kinase beta (IKK-beta) resulting in activation of the Nuclear factor NF-kappa-B (NF-kB) [14].

The cytoplasmic tyrosine kinase c-Src is involved in important cellular processes such as mitogenic signaling or cytoskeletal organization. Substrates of the EGF-stimulated c-Src include the EGFR itself, transcription factors of the Signal transducer and activator of transcription family, such as Signal transducer and activator of transcription 3 (STAT3), Shc, cytoskeletal components and some other proteins [15].

EGFR via Janus kinase 1 and 2 (JAK1 and JAK2) complex with STAT1 and STAT3 induces cell migration [16]

EGF is one of modulators of epithelial-to-mesenchymal transition (EMT). Excessive or inappropriate EGF stimulation leads to EMT during tumor development [17], [18]. EGF leads to EMT during tissue development, for example epicardial tissue [19]. EGF in conjunction with hydrocortisone induces EMT during postovulatory functional changes of ovarian surface epithelium [20]. Normally, EGF enhances Transforming growth factor beta (TGF-beta) signaling and induces EMT only in conjunction with TGF-beta 1 [21], [22], [23]. The common pathway for EGF-dependent EMT are ERK1/2 [20], [21] and PI3K [22] activation. EGF via PI3K activates Integrin-linked kinase (ILK) signaling and Glycogen synthase kinase 3 beta (GSK3 beta) and AKT-dependent activation of Cyclin E and Cyclin-dependent kinase 2 (Cdk2) and thus promotes proliferation during EMT. Also, Via GSK3 beta-dependent manner, probably via c-Jun (a protein of AP-1 complex) [24] promotes Matrix metallopeptidases (MMP-2 and MMP-9) expression. MMP expression usually initiated during EMT process [20].

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