Time resolved quantitative phosphoproteomics reveals distinct patterns of SHP2 dependence in EGFR signaling

Vemulapalli,; Chylek, Lily A.; Erickson, Alison R.; LaRochelle, Jonathan R.; Subramanian, Kartik; Mohseni, Morvarid; LaMarche, Matthew J.; Mg, Acker; Pk, Sorger; Gygi, SP; Sc, Blacklow

doi:10.1101/598664

Cited by 3 publications

(1 citation statement)

References 39 publications

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“…Several proteins interacting with SHP2 through its SH2 domains have been identified. Lists of more than 50 known or putative interacting proteins have been compiled in the past, − and several additional partners have been reported since then. − A database of the known interactions is available at . However, in many of these cases, the sites of interaction, the pY residues that bind specifically to the SHP2 N-SH2 domain, and the binding affinities have not been determined.…”

Section: Introductionmentioning

confidence: 99%

Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

Anselmi

Calligari

Hub

et al. 2020

J. Chem. Inf. Model.

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SH2 domain-containing tyrosine phosphatase 2 (SHP2), encoded by PTPN11 , plays a fundamental role in the modulation of several signaling pathways. Germline and somatic mutations in PTPN11 are associated with different rare diseases and hematologic malignancies, and recent studies have individuated SHP2 as a central node in oncogenesis and cancer drug resistance. The SHP2 structure includes two Src homology 2 domains (N-SH2 and C-SH2) followed by a catalytic protein tyrosine phosphatase (PTP) domain. Under basal conditions, the N-SH2 domain blocks the active site, inhibiting phosphatase activity. Association of the N-SH2 domain with binding partners containing short amino acid motifs comprising a phosphotyrosine residue (pY) leads to N-SH2/PTP dissociation and SHP2 activation. Considering the relevance of SHP2 in signaling and disease and the central role of the N-SH2 domain in its allosteric regulation mechanism, we performed microsecond-long molecular dynamics (MD) simulations of the N-SH2 domain complexed to 12 different peptides to define the structural and dynamical features determining the binding affinity and specificity of the domain. Phosphopeptide residues at position −2 to +5, with respect to pY, have significant interactions with the SH2 domain. In addition to the strong interaction of the pY residue with its conserved binding pocket, the complex is stabilized hydrophobically by insertion of residues +1, +3, and +5 in an apolar groove of the domain and interaction of residue −2 with both the pY and a protein surface residue. Additional interactions are provided by hydrogen bonds formed by the backbone of residues −1, +1, +2, and +4. Finally, negatively charged residues at positions +2 and +4 are involved in electrostatic interactions with two lysines (Lys89 and Lys91) specific for the SHP2 N-SH2 domain. Interestingly, the MD simulations illustrated a previously undescribed conformational flexibility of the domain, involving the core β sheet and the loop that closes the pY binding pocket.

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Section: Introductionmentioning

confidence: 99%

Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

Anselmi

Calligari

Hub

et al. 2020

J. Chem. Inf. Model.

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Dissecting protein tyrosine phosphatase signaling by engineered chemogenetic control of its activity

Fauser

Huyot

Matsche

et al. 2022

Journal of Cell Biology

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Protein tyrosine phosphatases (PTPases) are critical mediators of dynamic cell signaling. A tool capable of identifying transient signaling events downstream of PTPases is essential to understand phosphatase function on a physiological time scale. We report a broadly applicable protein engineering method for allosteric regulation of PTPases. This method enables dissection of transient events and reconstruction of individual signaling pathways. Implementation of this approach for Shp2 phosphatase revealed parallel MAPK and ROCK II dependent pathways downstream of Shp2, mediating transient cell spreading and migration. Furthermore, we show that the N-SH2 domain of Shp2 regulates MAPK-independent, ROCK II-dependent cell migration. Engineered targeting of Shp2 activity to different protein complexes revealed that Shp2-FAK signaling induces cell spreading whereas Shp2-Gab1 or Shp2-Gab2 mediates cell migration. We identified specific transient morphodynamic processes induced by Shp2 and determined the role of individual signaling pathways downstream of Shp2 in regulating these events. Broad application of this approach is demonstrated by regulating PTP1B and PTP-PEST phosphatases.

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Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

Anselmi

Calligari

Hub

et al. 2020

Preprint

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ABSTRACTSH2 domain-containing tyrosine phosphatase 2 (SHP2), encoded by PTPN11, plays a fundamental role in the modulation of several signaling pathways. Germline and somatic mutations in PTPN11 are associated with different rare diseases and hematologic malignancies, and recent studies have individuated SHP2 as a central node in oncogenesis and cancer drug resistance. SHP2 structure includes two Src homology 2 domains (N-SH2 and C-SH2) followed by a catalytic protein tyrosine phosphatase (PTP) domain. Under basal conditions, the N-SH2 domain blocks the active site, inhibiting phosphatase activity. Association of the N-SH2 domain with binding partners containing short amino acid motifs comprising a phosphotyrosine residue (pY) leads to N-SH2/PTP dissociation and SHP2 activation. Considering the relevance of SHP2 in signaling and disease and the central role of the N-SH2 domain in its allosteric regulation mechanism, we performed microsecond-long molecular dynamics simulations of the N-SH2 domain complexed to 12 different peptides, to define the structural and dynamical features determining the binding affinity and specificity of the domain. Phosphopeptide residues at position −2 to +5, with respect to pY, have significant interactions with the SH2 domain. In addition to the strong interaction of the pY residue with its conserved binding pocket, the complex is stabilized hydrophobically by insertion of residues +1, +3 and +5 in an apolar groove of the domain, and interaction of residue −2 with both the pY and a protein surface residue. Additional interactions are provided by hydrogen bonds formed by the backbone of residues −1, +1, +2 and +4. Finally, negatively charged residues at position +2 and +4 are involved in electrostatic interactions with two lysines (Lys89 and Lys91) specific of the SHP2 N-SH2 domain. Interestingly, the MD simulations illustrated a previously undescribed conformational flexibility of the domain, involving the core β-sheet and the loop that closes the pY binding pocket.

show abstract

Time resolved quantitative phosphoproteomics reveals distinct patterns of SHP2 dependence in EGFR signaling

Cited by 3 publications

References 39 publications

Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

Dissecting protein tyrosine phosphatase signaling by engineered chemogenetic control of its activity

Structural Determinants of Phosphopeptide Binding to the N-Terminal Src Homology 2 Domain of the SHP2 Phosphatase

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