Proteomic analysis of resting and thrombin-stimulated platelets reveals the translocation and functional relevance of HIP-55 in platelets.

Abstract

The platelet surface is a dynamic interface that changes rapidly in response to stimuli to co-ordinate the formation of thrombi at sites of vascular injury. Tight control is essential as loss of organisation may result in the inappropriate formation of thrombi (thrombosis) or excessive bleeding. In this paper we describe the comparative analysis of resting and thrombin-stimulated platelet membrane proteomes and associated proteins to identify proteins important to platelet function. Surface proteins were labelled using a biotin tag and isolated by NeurtrAvidin affinity chromatography. Liquid phase IEF and SDS-PAGE were used to separate proteins, and bands of increased intensity in the stimulated platelet fractions were digested and identified by FT-ICR mass spectrometry. Novel proteins were identified along with proteins known to be translocated to the platelet surface. Furthermore, many platelet proteins revealed changes in location associated with function, including G6B and Hip-55. HIP-55 is an SH3-binding protein important in T-cell receptor signalling. Further analysis of HIP-55 revealed that this adaptor protein becomes increasingly associated with both Syk and integrin beta3 upon platelet activation. Analysis of HIP-55 deficient platelets revealed reduced fibrinogen binding upon thrombin stimulation, suggesting HIP-55 to be an important regulator of platelet function.

• Authors: Katherine L Tucker, William J Kaiser, Angela L Bergeron, Hongbo Hu, Jing-fei Dong, Tse-Hua Tan, Jonathan M Gibbins
• Journal: Proteomics (Proteomics) Vol. 9 Issue 18 Pg. 4340-54 (Sep 2009) ISSN: 1615-9861 [Electronic] Germany
• PMID: 19725075 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DBNL protein, human
• Integrin beta3
• Intracellular Signaling Peptides and Proteins
• Microfilament Proteins
• Proteins
• Proteome
• Biotin
• Protein-Tyrosine Kinases
• SYK protein, human
• Syk Kinase
• Thrombin

Topics

• Biotin (metabolism)
• Biotinylation
• Blood Platelets (drug effects, metabolism)
• Cell Communication (drug effects, physiology)
• Cell Membrane (metabolism)
• Chromatography, Affinity
• Electrophoresis, Polyacrylamide Gel
• Humans
• Integrin beta3 (metabolism)
• Intracellular Signaling Peptides and Proteins (metabolism)
• Isoelectric Focusing
• Microfilament Proteins (blood)
• Protein Transport
• Protein-Tyrosine Kinases (metabolism)
• Proteins (metabolism)
• Proteome (drug effects)
• Proteomics (methods)
• Reproducibility of Results
• Syk Kinase
• Tandem Mass Spectrometry
• Thrombin (pharmacology)
• src Homology Domains