β-Adducin siRNA disruption of the spectrin-based cytoskeleton in differentiating keratinocytes prevented by calcium acting through calmodulin/epidermal growth factor receptor/cadherin pathway

Wu, Jianghong; Masci, Paul P.; Chen, Chenfeng; Chen, Jiezhong; Lavin, Martin F.; Zhao, Kong‐Nan

doi:10.1016/j.cellsig.2014.10.001

Cited by 15 publications

(12 citation statements)

References 44 publications

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“…Apparently, non-phosphorylated CaM only interacts with the EGFR when is not phosphorylated at Tyr 1173 , while failing to interact with the receptor phosphorylated at this residue. In contrast, P-(Tyr138)-CaM interacts with the EGFR in both nonphosphorylated and phosphorylated states [55]. We have also demonstrated that both non-phosphorylated CaM and P-(Tyr)-CaM bind to the same cytosolic juxtamembrane region of the EGFR (aa 645-660).…”

Section: Discussionmentioning

confidence: 72%

The activating role of phospho-(Tyr)-calmodulin on the epidermal growth factor receptor

Stateva

Salas

Benguría

et al. 2015

Biochemical Journal

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The activity of calmodulin (CaM) is modulated not only by oscillations in the cytosolic concentration of free Ca 2 + , but also by its phosphorylation status. In the present study, the role of tyrosine-phosphorylated CaM [P-(Tyr)-CaM] on the regulation of the epidermal growth factor receptor (EGFR) has been examined using in vitro assay systems. We show that phosphorylation of CaM by rat liver solubilized EGFR leads to a dramatic increase in the subsequent phosphorylation of poly-L-(Glu:Tyr) (PGT) by the receptor in the presence of ligand, both in the absence and in the presence of Ca 2 + . This occurred in contrast with assays where P-(Tyr)-CaM accumulation was prevented by the presence of Ca 2 + , absence of a basic cofactor required for CaM phosphorylation and/or absence of CaM itself. Moreover, an antibody against CaM, which inhibits its phosphorylation, prevented the extra ligand-dependent EGFR activation. Addition of purified P-(Tyr)-CaM, phosphorylated by recombinant c-Src (cellular sarcoma kinase) and free of non-phosphorylated CaM, obtained by affinity-chromatography using an immobilized antiphospho-(Tyr)-antibody, also increased the ligand-dependent tyrosine kinase activity of the isolated EGFR toward PGT. Also a CaM(Y99D/Y138D) mutant mimicked the effect of P-(Tyr)-CaM on ligand-dependent EGFR activation. Finally, we demonstrate that P-(Tyr)-CaM binds to the same site (as non-phosphorylated CaM, located at the cytosolic juxtamembrane region of the EGFR. These results show that P-(Tyr)-CaM is an activator of the EGFR and suggest that it could contribute to the CaM-mediated ligand-dependent activation of the receptor that we previously reported in living cells.

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Section: Discussionmentioning

confidence: 72%

The activating role of phospho-(Tyr)-calmodulin on the epidermal growth factor receptor

Stateva

Salas

Benguría

et al. 2015

Biochemical Journal

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“…Src-family kinases including c-Src are known to phosphorylate CaM (reviewed in [ 20 ]) as also demonstrated by us [ 27 , 28 ]. It has been shown in keratinocytes that phospho-(Y138)-CaM does not co-immunoprecipitate with Src, while non-phosphorylated CaM does [ 25 ]. Our results using CaM(Y99D/Y138D) and CaM(Y99E/Y138E) suggest that diphospho-(Y99/Y138)-CaM may be able to interact with Src, in contrast to what it was reported with monophospho-(Y138)-CaM [ 25 ].…”

Section: Discussionmentioning

confidence: 99%

“…The direct interaction of CaM with a recombinant glutathione S-transferase (GST)-Src fusion protein was demonstrated to occur via a dual Ca 2+ -dependent and Ca 2+ -independent mechanism, although mutation of the proposed CaM-binding site located at the Src homology domain 2 (SH2) of c-Src only partially prevented CaM binding [ 24 ]. Interestingly, Src was shown to co-immunoprecipitate with CaM but not with tyrosine-phosphorylated CaM in keratinocytes [ 25 ]. Also, the unique and SH3 domains of c-Src has been shown to bind acidic lipids, usually present in the inner leaflet of the plasma membrane, and binding of Ca 2+ /CaM to the unique domain has been proposed to regulate this process [ 26 ].…”

Section: Introductionmentioning

confidence: 99%

Ca2+/Calmodulin and Apo-Calmodulin Both Bind to and Enhance the Tyrosine Kinase Activity of c-Src

Stateva

Salas²,

Anguita

et al. 2015

PLoS ONE

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Src family non-receptor tyrosine kinases play a prominent role in multiple cellular processes, including: cell proliferation, differentiation, cell survival, stress response, and cell adhesion and migration, among others. And when deregulated by mutations, overexpression, and/or the arrival of faulty incoming signals, its hyperactivity contributes to the development of hematological and solid tumors. c-Src is a prototypical member of this family of kinases, which is highly regulated by a set of phosphorylation events. Other factor contributing to the regulation of Src activity appears to be mediated by the Ca2+ signal generated in cells by different effectors, where the Ca2+-receptor protein calmodulin (CaM) plays a key role. In this report we demonstrate that CaM directly interacts with Src in both Ca2+-dependent and Ca2+-independent manners in vitro and in living cells, and that the CaM antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) inhibits the activation of this kinase induced by the upstream activation of the epidermal growth factor receptor (EGFR), in human carcinoma epidermoide A431 cells, and by hydrogen peroxide-induced oxidative stress, in both A431 cells and human breast adenocarcinoma SK-BR-3 cells. Furthermore, we show that the Ca2+/CaM complex strongly activates the auto-phosphorylation and tyrosine kinase activity of c-Src toward exogenous substrates, but most relevantly and for the first time, we demonstrate that Ca2+-free CaM (apo-CaM) exerts a far higher activatory action on Src auto-phosphorylation and kinase activity toward exogenous substrates than the one exerted by the Ca2+/CaM complex. This suggests that a transient increase in the cytosolic concentration of free Ca2+ is not an absolute requirement for CaM-mediated activation of Src in living cells, and that a direct regulation of Src by apo-CaM could be inferred.

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“…Experiments using CaM-(Y99D/Y138D) and CaM-(Y99E/Y138E), where the negative charges of aspartic and glutamic acids mimic the phosphates [27], suggest that diphospho-CaM with pTyr99/pTyr138 can interact with Src, as can non-phosphorylated CaM, but not monophospho-CaM with pTyr138. CaM with pTyr138 also does not co-immunoprecipitate with Src [55], confirming pTyr99-CaM as a key player. In line with these observations, Chaudhuri et al .…”

Section: Cam Phosphorylationmentioning

confidence: 99%

Calmodulin and PI3K Signaling in KRAS Cancers

et al. 2017

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Calmodulin (CaM) uniquely promotes signaling of oncogenic K-Ras; but not N-Ras or H-Ras. How CaM interacts with K-Ras and how this stimulates cell proliferation are among the most challenging questions in KRAS-driven cancers. Earlier data pointed to formation of a ternary complex consisting of K-Ras, PI3Kα and CaM. Recent data point to phosphorylated CaM binding to the SH2 domains of the p85 subunit of PI3Kα and activating it. Modeling suggests that the high affinity interaction between the phosphorylated CaM tyrosine motif and PI3Kα, can promote full PI3Kα activation by oncogenic K-Ras. Our up-to-date review discusses CaM’s role in PI3K signaling at the membrane in KRAS-driven cancers. This is significant since it may help development of K-Ras-specific pharmacology.

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β-Adducin siRNA disruption of the spectrin-based cytoskeleton in differentiating keratinocytes prevented by calcium acting through calmodulin/epidermal growth factor receptor/cadherin pathway

Cited by 15 publications

References 44 publications

The activating role of phospho-(Tyr)-calmodulin on the epidermal growth factor receptor

The activating role of phospho-(Tyr)-calmodulin on the epidermal growth factor receptor

Ca2+/Calmodulin and Apo-Calmodulin Both Bind to and Enhance the Tyrosine Kinase Activity of c-Src

Calmodulin and PI3K Signaling in KRAS Cancers

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