scholarly journals Stapled peptides as a new technology to investigate protein–protein interactions in human platelets

2018 ◽  
Vol 9 (20) ◽  
pp. 4638-4643 ◽  
Author(s):  
Jessica Iegre ◽  
Niaz S. Ahmed ◽  
Josephine S. Gaynord ◽  
Yuteng Wu ◽  
Kara M. Herlihy ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Protein Interactions ◽  
New Technology ◽  
Human Platelets ◽  
Protein Protein Interactions ◽  
Stapled Peptides ◽  
Traditional Methods

We describe the first application of stapled peptides in human platelets. Bim BH3 stapled peptides are used to overcome the limitations of traditional methods and uncover a new role for Bim in platelet activation.

CHAPTER 8. Double-click Stapled Peptides for Inhibiting Protein–Protein Interactions

2017 ◽  
pp. 164-187 ◽  
Author(s):  
K. Sharma ◽  
D. L. Kunciw ◽  
W. Xu ◽  
M. M. Wiedmann ◽  
Y. Wu ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Stapled Peptides ◽  
Inhibiting Protein

scholarly journals Titelbild: Light-Regulated Stapled Peptides to Inhibit Protein-Protein Interactions Involved in Clathrin-Mediated Endocytosis (Angew. Chem. 30/2013)

2013 ◽  
Vol 125 (30) ◽  
pp. 7759-7759
Author(s):  
Laura Nevola ◽  
Andrés Martín-Quirós ◽  
Kay Eckelt ◽  
Núria Camarero ◽  
Sébastien Tosi ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Stapled Peptides

C-Cbl-PI3K Interaction Is Important for Platelet Outside-In Signaling

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2014-2014
Author(s):  
Claudia Lorena Buitrago ◽  
Satya P. Kunapuli ◽  
Archana Sanjay
Keyword(s):  
Actin Cytoskeleton ◽  
Protein Interactions ◽  
Conflicts Of Interest ◽  
Physiological Role ◽  
Human Platelets ◽  
Scaffolding Protein ◽  
Protein Protein Interactions ◽  
Clot Retraction ◽  
Knock Out

Abstract Abstract 2014 Platelet activation by outside-in signaling is initiated by the binding of fibrinogen to alphaIIbbeta3, an integrin only expressed in platelets and megakaryocytes. Signals transduced by alphaIIbbeta3 regulate actin cytoskeleton resulting in filopodia and lamellipodia formation, cell spreading and retraction. c-Cbl protein is abundantly expressed in platelets and functions as E3 ubiquitin ligase and scaffolding protein to mediate protein-protein interactions. Importantly, c-Cbl tyrosine 731 has been shown to interact with p85 subunit of phosphotidylinositol 3-kinase (PI3K) modulating the actin cytoskeleton. Although previous reports showed c-Cbl activation downstream of alphaIIbbeta3, the mechanisms and implications of this activation or the downstream targets remain to be elucidated. We have studied the role of c-Cbl in platelet outside-in signaling: Using human platelets we have demonstrated that c-Cbl Y700, Y731 and Y774 residues undergoes tyrosine phosphorylation upon platelet adhesion to immobilized fibrinogen. These phosphorylation events are completely inhibited in the presence of the pan Src Family Kinases (SFKs) inhibitor (PP2) suggesting that c-Cbl is phosphorylated downstream of SFKs. Spleen tyrosine kinase (Syk) is also involved in this signaling pathway since its inhibition significantly reduce c-Cbl phosphorylation at residues Y774 and Y700; interestingly, tyrosine 731 phosphorylation, which allows the interaction with the p85-subunit of PI3K, is not affected by Syk inhibition. The physiological role of c-Cbl in platelet outside-in signaling was studied using c-Cbl knock-out mice. We found that in contrast to WT platelets, c-Cbl KO platelets had a significantly reduced spreading over a fibrinogen-coated surface. Furthermore, clot retraction analysis demonstrated that c-Cbl KO platelets retraction time was delayed when compared to WT platelets, suggesting a retraction defect. To further elucidate the physiological role of c-Cbl-PI3K interaction we used a knock-in mouse in which the c-Cbl residue Y 731 was substituted with phenylalanine (Y731F) thereby abolishing the PI3K binding site on c-Cbl. Importantly, platelets from Y731F mice showed spreading and clot retraction defect that were comparable with the c-Cbl KO. These result indicates that in large part, the role of c-Cbl in platelets outside-in signaling is determined by its interaction with PI3K. In conclusion, we have demonstrated that c-Cbl plays an important role in platelet outside-in signaling, and its interaction with PI3K through tyrosine 731 is of pivotal importance in platelet spreading and retraction. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Cover Picture: Light-Regulated Stapled Peptides to Inhibit Protein-Protein Interactions Involved in Clathrin-Mediated Endocytosis (Angew. Chem. Int. Ed. 30/2013)

2013 ◽  
Vol 52 (30) ◽  
pp. 7607-7607
Author(s):  
Laura Nevola ◽  
Andrés Martín-Quirós ◽  
Kay Eckelt ◽  
Núria Camarero ◽  
Sébastien Tosi ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Stapled Peptides

Light-Regulated Stapled Peptides to Inhibit Protein-Protein Interactions Involved in Clathrin-Mediated Endocytosis

2013 ◽  
Vol 125 (30) ◽  
pp. 7858-7862 ◽  
Author(s):  
Laura Nevola ◽  
Andrés Martín-Quirós ◽  
Kay Eckelt ◽  
Núria Camarero ◽  
Sébastien Tosi ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Stapled Peptides

scholarly journals Unprotected Peptide Macrocyclization and Stapling via A Fluorine-Thiol Displacement Reaction

2020 ◽  
Author(s):  
Md Shafiqul Islam ◽  
Samuel L. Junod ◽  
Si Zhang ◽  
Zakey Yusuf Buuh ◽  
Yifu Guan ◽  
...  
Keyword(s):  
Cellular Uptake ◽  
Protein Interactions ◽  
Cell Permeability ◽  
Displacement Reaction ◽  
Protein Protein Interactions ◽  
Cancer Cell Growth ◽  
Stapled Peptides ◽  
Peptide Substrates ◽  
New Approach ◽  
Dynamics Simulations

AbstractStapled peptides serve as a powerful tool for probing protein-protein interactions, but its application has been largely impeded by the limited cellular uptake. Here we report the discovery of a facile peptide macrocyclization and stapling strategy based on a fluorine thiol displacement reaction (FTDR), which renders a class of peptide analogues with enhanced stability, affinity, and cell permeability. This new approach enabled selective modification of the orthogonal fluoroacetamide side chains in unprotected peptides, with the identified 1,3-benzenedimethanethiol linker promoting alpha helicity of a variety of peptide substrates, as corroborated by molecular dynamics simulations. The cellular uptake of these stapled peptides was universally enhanced compared to the classic ring-closing metathesis (RCM) stapled peptides. Pilot mechanism studies suggested that the uptake of FTDR-stapled peptides may involve multiple endocytosis pathways. Consistent with the improved cell permeability, the FTDR-stapled lead Axin analogues demonstrated better inhibition of cancer cell growth than the RCM-stapled analogues.Graphical Abstract

scholarly journals Immune versus thrombotic stimulation of platelets differentially regulates signalling pathways, intracellular protein-protein interactions, and α-granule release

2009 ◽  
Vol 102 (07) ◽  
pp. 97-110 ◽  
Author(s):  
Sybille Rex ◽  
Lea M. Beaulieu ◽  
David H. Perlman ◽  
Olga Vitseva ◽  
Price S. Blair ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Protein Interactions ◽  
Fibrin Clot ◽  
Protein Release ◽  
Factor Xiiia ◽  
Signalling Pathways ◽  
Protein Protein Interactions ◽  
Platelet Surface ◽  
Granule Release ◽  
Stimulation Of

SummaryIn addition to haemostasis, platelets mediate inflammation and clearance of bacteria from the bloodstream. As with platelet-platelet interactions, platelet-bacteria interactions involve cytoskeletal rearrangements and release of granular content. Stimulation of the immune Toll-like receptor 2 (TLR2) on the platelet surface, activates phosphoinositide-3-kinase (PI3K) and causes platelet activation and plateletdependent thrombosis. It remains unknown if platelet activation by immune versus thrombotic pathways leads to the differential regulation of signal transduction, protein-protein interactions, and α-granule release, and the physiological relevance of these potential differences. We investigated these processes after immune versus thrombotic platelet stimulation. We examined selected signalling pathways and found that phosphorylation kinetics of Akt, ERK1/2 and p38 differed dramatically between agonists. Next, we investigated platelet protein-protein interactions by mass spectrometry (MS)-based proteomics specifically targeting cytosolic factor XIIIa (FXIIIa) because of its function as a cytoskeleton-crosslinking protein whose binding partners have limited characterisation. Four FXIIIa-binding proteins were identified, two of which are novel interactions: FXIIIa-focal adhesion kinase (FAK) and FXIIIa-gelsolin. The binding of FAK to FXIIIa was found to be altered differentially by immune versus thrombotic stimulation. Lastly, we studied the effect of thrombin versus Pam3CSK4 stimulation on α-granule release and observed differential release patterns for selected granule proteins and decreased fibrin clot formation compared with thrombin. The inhibition of PI3K caused a decrease in protein release after Pam3CSK4 -but not after thrombin-stimulation. In summary, stimulation of platelets by either thrombotic or immune receptors leads to markedly different signalling responses and granular protein release consistent with differential contribution to coagulation and thrombosis.

scholarly journals Designing stapled peptides to inhibit protein‐protein interactions: An analysis of successes in a rapidly changing field

2020 ◽  
Author(s):  
Marie T. J. Bluntzer ◽  
James O'Connell ◽  
Terry S. Baker ◽  
Julien Michel ◽  
Alison N. Hulme
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Stapled Peptides
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