Grb2 regulation of the actin-based cytoskeleton is required for ligand-independent EGF receptor-mediated oncogenesis

Abstract

Mutations within members of the EGF/ErbB receptor family frequently release the oncogenic potential of these receptors, resulting in the activation of downstream signaling events independent of ligand regulatory constraints. We previously have demonstrated that the signal transduction events originating from S3-v-ErbB, a ligandindependent, oncogenic EGF receptor mutant, are qualitatively distinct from the ligand-dependent mitogenic signaling pathways associated with the wild-type EGF receptor. Specifically, expression of S3-v-ErbB in primary fibroblasts results in anchorage-independent growth, increased invasive potential, and the formation of a transformation-specific phosphoprotein signaling complex, all in a Ras-independent manner. Here we demonstrate the transformation-specific interaction between two components of this complex: the adaptor protein Grb2 and the cytoskeletal regulatory protein caldesmon. This interaction is mediated via both the amino-terminal SH3 and central SH2 domains of Grb2, and the aminoterminal (myosin-binding) domain of caldesmon. Expression of a dominant-negative Grb2 deletion mutant, which lacks the carboxy-terminal SH3 domain, in fibroblasts expressing S3-v-ErbB results in a reduction in phosphoprotein complex formation, the loss of anchorageindependent growth, and a reduction in invasive potential.Together, these results demonstrate a Ras-independent role for Grb2 in modulating cytoskeletal function during ligand-independent EGF receptor-mediated transformation, and provide further support for the hypothesis that ligand-independent oncogenic signaling is qualitatively distinct from ligand-dependent mitogenic signaling by the EGF receptor.

Cyclin-Dependent Kinase 5 (CDK5) Controls Melanoma Cell Motility, Invasiveness, and Metastatic Spread—Identification of a Promising Novel therapeutic target

“…Tyrosine phosphorylation of caldesmon has been found to cause its dissociation from actin, thereby contributing to actin microfilament disassembly [54,55] . In a recent study by Boerner et al, only phosphotyrosine-27 of caldesmon was found to be required for binding to either the amino-terminal SH3 or central SH2 domains of Grb2 [56] . Our data are in line with a recent report by Quintavalle et al suggesting that CDK5 has the potential to directly phosphorylate caldesmon in src-3 T3 and SCC61 cells, thereby inhibiting cytoskeleton remodeling, formation of invadopodia, and invasive behavior [51] .…”

Phosphorylation of caldesmon during smooth muscle contraction and cell migration or proliferation

“…Although some of these issues have indeed been clarified for smooth muscle CaD, those concerning non-muscle CaD still remain unresolved after almost two decades. Significantly, the concept of thin filament regulation in non-muscle cells is being supported by quite a few recent reports [7,[9][10][11][12]. Thus it seems timely to revisit this problem now.…”

“…Whether, for example, phospho-CaD plays a role in recruiting essential partners to the focal adhesions remains to be seen. Furthermore, it was reported that l-CaD is associated with Grb2, Shc and Sos, and in this complex l-CaD was found to be tyrosine-phosphorylated [9,61]. The significance of this also awaits further investigation.…”

Caldesmon and the Regulation of Cytoskeletal Functions

“…Furthermore, CaD is associated with Grb2, Shc and Sos; in this complex CaD was found to be tyrosine-phosphorylated. 199,200 More recently, it has also been suggested that the interaction between phospho-CaD and Tm is further influenced by Hsp27. 201 The significance of all these findings awaits further investigation.…”