Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi


Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101504


Historical Background

The Shc family of proteins consists four members, ShcA, ShcB, ShcC, and ShcD, of which the best characterized to date is ShcA (Rozakis-Adcock et al. 1992). ShcA, or simply Shc, was identified in 1992 as an adaptor protein which linked the activated EGFR (epidermal growth factor receptor) to Ras and the MAP (mitogen-activated protein) kinase cascade (Ravichandran 2001). Shc is expressed as three isoforms which have the molecular weights of 66, 52, and 46 kDa, respectively. These isoforms are designated according to their molecular weight. p66Shc is the longest isoform of the three and also consists of the collagen homology domain (CH2), which is unique to it.

P66Shc Structure

All proteins of the Shc family share a unique and highly conserved domain organization, having an N-terminal PTB domain and a C-terminal SH2 domain. Collagen homology (CH1) is proline-rich central domain which in humans has three phosphorylatable tyrosine residues 239/240 and 317 (Fig. 1). When these residues are phosphorylated, they lead to recruitment of Grb2/Sos to the plasma membrane (Ravichandran 2001). These two Grb2 binding sites have been proposed to couple cell surface receptors either to fos-dependent cell proliferation (Y317) or to myc-dependent cell survival (YY239/240) in some mammalian cell types (Gotoh et al. 1996). Since the proportion of proline residues is high in the CH1 domain, they are prone to SH3 domain-dependent interactions, thus strengthening the concept and capacity of Shc proteins to function as molecular adaptors. p66Shc which is the longest isoform has an additional N-terminal CH domain, termed CH2, which can be phosphorylated at a serine residue at position 36 (S36) (Fig. 1). Mouse-based studies lacking p66Shc have revealed an unforeseen role of p66Shc in apoptotic responses to oxidative stress and in aging (Migliaccio et al. 1999).
p66Shc, Fig. 1

Schematic structure of p66Shc protein

P66Shc and Cell Signaling

p46Shc and p52Shc are ubiquitously expressed, while p66Shc is expressed at different level in various tissues. p66Shc, upon stimulation by growth factors, is tyrosine phosphorylated by receptor tyrosine kinases. p66Shc upon tyrosine phosphorylation binds Grb2 and is unable to activate the Ras-MAPK-Fos pathways (Migliaccio et al. 1997, 1999). Ras signaling pathway is also inhibited when p66Shc is overexpressed upon stimulation by growth factors or cytokines (Migliaccio et al. 1997; Okada et al. 1997). Stimulation of MEK (MAPK-ERK kinase)/ERK (extracellular signal-regulated kinase) pathway by insulin growth factor 1 (IGF-1) is also inhibited by p66Shc. This pathway is required by the actin cytoskeleton phosphorylation in skeleton muscle myoblasts (Nemoto et al. 2006). Another adaptor protein E3b1 together with Eps8 also plays a role in regulating the phosphorylation, reducing ubiquitylation, and increasing the stability of p66Shc protein through Rac1. Sos1 can exist in a complex with Eps8/E3b1, and the complex of Sos1/Eps8/E3b1 leads to the activation of Rac1 (Innocenti et al. 2002). p66Shc plays the role of a switch to dissociate Sos1 from the Grb2/Sos1 pool to Eps8/E3b1 pool. This in turn leads to Rac1 activation and as a result leads to an increase in the generation of oxidants (Migliaccio et al. 1997). Increased binding of p66Shc to activated EGFR and Grb2 occurs during severe oxidative stress. This binding leads to the dissociation of the Sos1 adaptor protein from the EGFR-recruited signaling complex. These events lead to the termination of the Ras/MEK/ERK activation, which further lead to varied downstream effects including cell proliferation, cell differentiation, apoptosis, carcinogenesis, etc. (Arany et al. 2008) (Fig. 2).
p66Shc, Fig. 2

Schematic representation of the role of p66Shc in cell signaling and downstream effects

Consequences of P66Shc Phosphorylations

Under oxidative stress the S36 residue of p66Shc is phosphorylated. Treatment with an iron-containing porphyrin, hemin, increased the phosphorylation of p66Shc at the S36 residue. In hemin-treated K562 erythroleukemic cells, p66Shc was transcriptionally activated through the ARE (antioxidant response element)-Nrf2 (NF-E2-related factor 2) pathway (Miyazawa and Tsuji 2014). In human colon carcinoma cell line RKO and in diploid human dermal fibroblasts, suppression in the production of ROS was seen in shRNA-mediated knockdown of p66Shc, and an increase in the production of ROS was observed upon overexpression of a recombinant p66Shc. These effects were not seen in the electron transport chain-deficient ρ0-RKO cells. These ρ0-RKO cells are mitochondrial DNA-depleted cells.

Till now most of the studies have demonstrated that p66Shc shows a proapoptotic and pro-oxidative role. Up to 30% extension in life span was seen in p66Shc knockdown mice. These knockdown mice also showed better handling of ROS and oxidative stress (Migliaccio et al. 1999). Mutagenesis of S36 to alanine results in a p66Shc variant that is unable to induce apoptosis. Therefore, S36 appears to be a critical regulatory site for the apoptotic activity and oxidative stress response of p66Shc (Galimov et al. 2014). This phosphorylation of the S36 residue of p66Shc in response to oxidative stress is done by PKCβ, which is activated by oxidative stress. Oxidative stress gives a stimulus to mouse thymocytes, peripheral blood lymphocytes, and splenic T-cells, and these cells then attain the ability to express p66Shc. Phosphorylation of p66Shc on the S36 residue has been shown to have a proapoptotic effect in different cell lines. The responses of S36 phosphorylation depends on the kinase(s) that mediate this process. The kinase(s) may differ (MAPK, stress-activated JNK, and P38) depending on the cell type or the type of inducement. When faced with oxidative stress conditions, S36 on p66Shc is phosphorylated which leads to the activation of p66Shc (Fig. 3). This activated p66Shc sensitizes to apoptotic stimuli after it is dissociated from an inhibitory complex. Apoptotic death induced by oxidative stress is dependent on p53, and knockout of either p53 or p66Shc leads to resistance. This suggests that p66Shc is downstream of p53 in the pathway leading from ROS to apoptosis. The S54 site in the CH2 domain of p66Shc is important for its stability. The proteolytic PEST signal sequence of p66Shc is disguised by the phosphorylation of S54 in the CH2 domain and the T386 in the CH1 domain of p66Shc. These phosphorylations are mediated by Rac1, and these Rac1-mediated phosphorylations lead to increase in the stability of p66Shc adaptor protein (Khanday et al. 2006). Conversely, it was also seen that p66Shc leads to the activation of Rac1 through the mediation of exchange factor. Recently, it was seen that in SH-SY5Y cells or in the mice cortex, exogenous H2S inhibited the mitochondrial ROS production and, as a result, phosphorylation of p66Shc and lead to sulfhydration of p66Shc on C59 residue. ROS are known to regulate canonical Wnt signaling. The canonical Wnt ligand, Wnt3a, also induces phosphorylation of p66Shc in endothelial cells. The knockdown of p66Shc inhibited both β-catenin dephosphorylation that is stimulated by Wnt3a and β-catenin-dependent transcription. This inhibition was seen to be reversed by the overexpression of p66Shc, independent of Wnt3a. β-catenin dephosphorylation brought about by exogenous H2O2 was also seen to be mediated by p66Shc (Vikram et al. 2014). Many stimuli that are engaged in tyrosine phosphorylation of p66Shc also bring about the phosphorylation of S138 in the PTB domain. The association of p66Shc adaptor protein in tyrosine phosphorylation signaling pathway is well known. Shc adaptor proteins, primarily p66Shc, transmit activated tyrosine phosphorylation signaling, which points to their possible role in growth regulation including carcinogenesis and metastasis (Fig. 3).
p66Shc, Fig. 3

Causes and effects of phosphorylation p66Shc residues


p66Shc, an adaptor protein, is the longest isoform of the ShcA family. p66Shc has an additional CH domain at the N-terminal, called the CH2 domain, which is not present in the other isoforms. This CH2 domain contains a very crucial S36 residue which is phosphorylated in response to oxidative stress and plays a role in apoptosis. This adaptor protein has been shown to be involved in mediating and executing the post effects of oxidative stress, and increasing body of evidence is pinpointing to its role in carcinogenesis as well. This multitasking protein is involved in regulating different networks of cell signaling. On one hand, it shows an increased expression profile in different cancers and has a positive role in cell proliferation and migration, whereas on the other hand, it promotes apoptosis under oxidative stress conditions by acting as a sensor of ROS. p66Shc by regulating intracellular ROS levels plays a crucial role in regulating longevity and cell senescence. These multifaceted properties of p66Shc make it a perfect candidate protein for further studies in various cancers and aging-related diseases. p66Shc can be targeted in terms of it being used as a possible therapeutic target in various diseases.


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© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Department of BiotechnologyUniversity of KashmirSrinagarIndia