Regulation of the ligand-dependent activation of the epidermal growth factor receptor by calmodulin

Hongbing Li; Svetlana Panina; Amandeep Kaur; María J. Ruano; Pablo Sánchez-González; Peter Jonas Marstrand la Cour; Alexander Stephan; Uffe Høgh Olesen; Martin Werner Berchtold; Antonio Villalobo

doi:10.1074/jbc.M111.317529

Regulation of the ligand-dependent activation of the epidermal growth factor receptor by calmodulin

Hongbing Li, Svetlana Panina, Amandeep Kaur, María J. Ruano, Pablo Sánchez-González, Peter Jonas Marstrand la Cour, Alexander Stephan, Uffe Høgh Olesen, Martin Werner Berchtold, Antonio Villalobo

Cell Biology and Physiology

27 Citations (Scopus)

Abstract

Calmodulin (CaM) is the major component of calcium signaling pathways mediating the action of various effectors. Transient increases in the intracellular calcium level triggered by a variety of stimuli lead to the formation of Ca ²⁺/CaM complexes, which interact with and activate target proteins. In the present study the role of Ca ²⁺/CaM in the regulation of the ligand-dependent activation of the epidermal growth factor receptor (EGFR) has been examined in living cells. We show that addition of different cell permeable CaM antagonists to cultured cells or loading cells with a Ca ²⁺ chelator inhibited ligand-dependent EGFR auto(trans) phosphorylation. This occurred also in the presence of inhibitors of protein kinase C, CaM-dependent protein kinase II and calcineurin, which are known Ca ²⁺- and/or Ca ²⁺/CaM-dependent EGFR regulators, pointing to a direct effect of Ca ²⁺/CaM on the receptor. Furthermore, we demonstrate that down-regulation of CaM in conditional CaM knock out cells stably transfected with the human EGFR decreased its ligand-dependent phosphorylation. Substitution of six basic amino acid residues within the CaM-binding domain (CaM-BD) of the EGFR by alanine resulted in a decreased phosphorylation of the receptor and of its downstream substrate phospholipase Cγ1. These results support the hypothesis that Ca ²⁺/CaM regulates the EGFR activity by directly interacting with the CaM-BD of the receptor located at its cytosolic juxtamembrane region.

Original language	English
Journal	Journal of Biological Chemistry
Volume	287
Issue number	5
Pages (from-to)	3273-3281
Number of pages	9
ISSN	0021-9258
DOIs	https://doi.org/10.1074/jbc.M111.317529
Publication status	Published - 27 Jan 2012

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10.1074/jbc.M111.317529

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title = "Regulation of the ligand-dependent activation of the epidermal growth factor receptor by calmodulin",

abstract = "Calmodulin (CaM) is the major component of calcium signaling pathways mediating the action of various effectors. Transient increases in the intracellular calcium level triggered by a variety of stimuli lead to the formation of Ca 2+/CaM complexes, which interact with and activate target proteins. In the present study the role of Ca 2+/CaM in the regulation of the ligand-dependent activation of the epidermal growth factor receptor (EGFR) has been examined in living cells. We show that addition of different cell permeable CaM antagonists to cultured cells or loading cells with a Ca 2+ chelator inhibited ligand-dependent EGFR auto(trans) phosphorylation. This occurred also in the presence of inhibitors of protein kinase C, CaM-dependent protein kinase II and calcineurin, which are known Ca 2+- and/or Ca 2+/CaM-dependent EGFR regulators, pointing to a direct effect of Ca 2+/CaM on the receptor. Furthermore, we demonstrate that down-regulation of CaM in conditional CaM knock out cells stably transfected with the human EGFR decreased its ligand-dependent phosphorylation. Substitution of six basic amino acid residues within the CaM-binding domain (CaM-BD) of the EGFR by alanine resulted in a decreased phosphorylation of the receptor and of its downstream substrate phospholipase Cγ1. These results support the hypothesis that Ca 2+/CaM regulates the EGFR activity by directly interacting with the CaM-BD of the receptor located at its cytosolic juxtamembrane region.",

author = "Hongbing Li and Svetlana Panina and Amandeep Kaur and Ruano, {Mar{\'i}a J.} and Pablo S{\'a}nchez-Gonz{\'a}lez and {la Cour}, {Peter Jonas Marstrand} and Alexander Stephan and Olesen, {Uffe H{\o}gh} and Berchtold, {Martin Werner} and Antonio Villalobo",

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doi = "10.1074/jbc.M111.317529",

language = "English",

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T1 - Regulation of the ligand-dependent activation of the epidermal growth factor receptor by calmodulin

AU - Li, Hongbing

AU - Panina, Svetlana

AU - Kaur, Amandeep

AU - Ruano, María J.

AU - Sánchez-González, Pablo

AU - la Cour, Peter Jonas Marstrand

AU - Stephan, Alexander

AU - Olesen, Uffe Høgh

AU - Berchtold, Martin Werner

AU - Villalobo, Antonio

PY - 2012/1/27

Y1 - 2012/1/27

N2 - Calmodulin (CaM) is the major component of calcium signaling pathways mediating the action of various effectors. Transient increases in the intracellular calcium level triggered by a variety of stimuli lead to the formation of Ca 2+/CaM complexes, which interact with and activate target proteins. In the present study the role of Ca 2+/CaM in the regulation of the ligand-dependent activation of the epidermal growth factor receptor (EGFR) has been examined in living cells. We show that addition of different cell permeable CaM antagonists to cultured cells or loading cells with a Ca 2+ chelator inhibited ligand-dependent EGFR auto(trans) phosphorylation. This occurred also in the presence of inhibitors of protein kinase C, CaM-dependent protein kinase II and calcineurin, which are known Ca 2+- and/or Ca 2+/CaM-dependent EGFR regulators, pointing to a direct effect of Ca 2+/CaM on the receptor. Furthermore, we demonstrate that down-regulation of CaM in conditional CaM knock out cells stably transfected with the human EGFR decreased its ligand-dependent phosphorylation. Substitution of six basic amino acid residues within the CaM-binding domain (CaM-BD) of the EGFR by alanine resulted in a decreased phosphorylation of the receptor and of its downstream substrate phospholipase Cγ1. These results support the hypothesis that Ca 2+/CaM regulates the EGFR activity by directly interacting with the CaM-BD of the receptor located at its cytosolic juxtamembrane region.

AB - Calmodulin (CaM) is the major component of calcium signaling pathways mediating the action of various effectors. Transient increases in the intracellular calcium level triggered by a variety of stimuli lead to the formation of Ca 2+/CaM complexes, which interact with and activate target proteins. In the present study the role of Ca 2+/CaM in the regulation of the ligand-dependent activation of the epidermal growth factor receptor (EGFR) has been examined in living cells. We show that addition of different cell permeable CaM antagonists to cultured cells or loading cells with a Ca 2+ chelator inhibited ligand-dependent EGFR auto(trans) phosphorylation. This occurred also in the presence of inhibitors of protein kinase C, CaM-dependent protein kinase II and calcineurin, which are known Ca 2+- and/or Ca 2+/CaM-dependent EGFR regulators, pointing to a direct effect of Ca 2+/CaM on the receptor. Furthermore, we demonstrate that down-regulation of CaM in conditional CaM knock out cells stably transfected with the human EGFR decreased its ligand-dependent phosphorylation. Substitution of six basic amino acid residues within the CaM-binding domain (CaM-BD) of the EGFR by alanine resulted in a decreased phosphorylation of the receptor and of its downstream substrate phospholipase Cγ1. These results support the hypothesis that Ca 2+/CaM regulates the EGFR activity by directly interacting with the CaM-BD of the receptor located at its cytosolic juxtamembrane region.

U2 - 10.1074/jbc.M111.317529

DO - 10.1074/jbc.M111.317529

M3 - Journal article

C2 - 22157759

SN - 0021-9258

VL - 287

SP - 3273

EP - 3281

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 5

ER -

Regulation of the ligand-dependent activation of the epidermal growth factor receptor by calmodulin

Abstract

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