p66^Shc Couples Mechanical Signals to RhoA through Focal Adhesion Kinase-Dependent Recruitment of p115-RhoGEF and GEF-H1

Abstract

Tissue cells respond to changes in tensional forces with proliferation or death through the control of RhoA. However, the response coupling mechanisms that link force with RhoA activation are poorly understood. We found that tension applied to fibronectin-coated microbeads caused recruitment of all three isoforms of the Shc adapter (p66 Shc , p52 Shc , and p46 Shc ) to adhesion complexes. The Shc PTB domain was necessary and sufficient for this recruitment, and screening studies revealed the direct interactions with the FERM domain of focal adhesion kinase (FAK) that were required for Shc translocation to adhesion complexes. The FAK/p66Shc complex specifically bound and activated the Rho guanyl exchange factors (GEFs) p115-RhoGEF and GEF-H1, leading to tension-induced RhoA activation. In contrast, the FAK/p52Shc complex bound SOS1 but not the Rho GEFs to mediate tension-induced Ras activation. Nuclear translocation and activation of the YAP/TAZ transcription factors on firm substrates required the FAK/p66Shc /Rho GEF complex, and both proliferation on firm substrates and anoikis in suspension required signaling through p66Shc and its associated Rho GEFs. These studies reveal the binary and exclusive assignment of p66 Shc and p52Shc to tension-induced Rho or Ras signals, respectively, and suggest an integrated role for the two Shc isoforms in coordinating the cellular response to mechanical stimuli.

p66ShcA functions as a contextual promoter of breast cancer metastasis

“…To define the mechanisms by which p66ShcA enhances metastasis, we therefore decided to examine the role of p66ShcA in breast cancer cell motility, partly guided by previous research showing that it can localize to focal adhesions and mediate signaling [14,16,17]. Our results demonstrate that p66ShcA loss severely reduces breast cancer motility in vitro due to the formation of larger and more stable cellular adhesions.…”

“…Ruling out an EMT as the primary mechanism by which p66ShcA increased breast cancer lung metastasis, we next examined its impact on the migratory properties of breast cancer cells. Previous research implicated p66ShcA as a mediator of focal adhesion signaling, an essential part of cellular motility [17,30]. We first assessed whether p66ShcA could promote cell migration by performing live-cell tracking experiments.…”

“…For example, p66ShcA restrains Ras signaling in lung cancer cells by reducing activation of Grb2/SOS signaling complexes [6,14]. In addition, p66ShcA suppresses an epithelial-to-mesenchymal transition (EMT) in lung cancer cells [15] and increases anoikis [16,17].…”

AMPA receptors modulate the reorganization of F‐actin in Bergmann glia cells through the activation of RhoA

“…which are well known for their ability to activate GEFs that mediate the activation of RhoA (Tanabe et al 2006;Iwanicki et al 2008;Kuroiwa et al 2011;Chandra et al 2012;Scott et al 2016;Wu et al 2016). To test the participation of these kinases in the modulation of RhoA, and in consequence, in the reorganization of the actin cytoskeleton of the Bergmann fibers, we first measured the activation of RhoA by immunoprecipitation using protein extracts obtained from adult cerebelar slices pre-treated with the PI3-K inhibitor wortmannin, the FAK inhibitor 14 (FI14) or the PKC inhibitor C1.…”

The mechanical microenvironment regulates ovarian cancer cell morphology, migration, and spheroid disaggregation

“…A central mediator of adhesion-dependent signaling, the role of FAK in mechanotransduction is regarded as being less in the generation of cellular tension and more in the transduction of signals downstream of mechanical cell-matrix interactions [65][66][67] . Nonetheless, its demonstrated importance in both functional mechanical signaling [68][69][70][71] and in the biology of EOC [72][73][74][75][76][77] prompted us to assess the effect of FAK inhibition on the EOC mechano-response. Unlike blebbistatin and fasudil, treatment of SKOV3 cells with PF-271 (also known as PF-562271), a potent and highly selective inhibitor of FAK, had no significant effect on cellular traction force ( Figure 8C and D) or stress fiber and phospho-MLC architecture ( Figure 8G and I), but significantly reduced phospho-FAK intensity while preserving or slightly enhancing focal adhesion size and aspect ratio ( Figure 8E and F), consistent with FAK's role in focal adhesion turnover 78 .…”