Interaction of platelet receptors with native collagen sequences and their downstream signalling effects

Lead Research Organisation: University of Cambridge
Department Name: Biochemistry

Abstract

Platelets are small cells found in the blood throughout the whole body. Normally, platelets do nothing and stay in a resting state for about ten days after which they die and are replaced. However, when a blood vessel is damaged, the platelets become active and respond immediately like an emergency service. They stick to the injured vessel and form a clot which prevents loss of blood and begins the process of healing. Unfortunately though, there are times when platelets block blood vessels in a manner which is harmful to the person. Over time, coronary arteries, which supply blood to the heart, can become obstructed with deposits of fat and cholesterol. These fatty deposits are more brittle than the elastic arteries and sometimes crack under stress. When this happens the platelets stick to the damaged area and can completely block the artery causing a heart attack. In both the damaged vessel and the cracked coronary artery, the platelets stick to a protein in the tissue called collagen. Collagen is very strong and acts like a soft tissue skeleton holding all the various body parts together. When platelets stick to collagen they become active and then stick to each other as well.

Our research looks at the way collagen sticks to and activates platelets. We already know that collagen sticks to particular proteins that are found on the surface of the platelet. Two of the most important of these proteins are called GPVI and ??2?O1. GPVI and ??2?O1 only stick to particular areas of the collagen. To find these areas we have made a series of 57 mini-collagens which resemble small segments of natural collagen found in our blood vessels. We now know that platelets only stick to a small number of these mini-collagens. This enables us to focus on the detailed properties of the collagen which makes it stick to the platelet.

Designing new treatments is a complex process and requires a detailed understanding of the processes which we want to influence. Our research attempts to provide that understanding which may assist in the design of new medicines to treat heart disease. Although the process of research is time-consuming, the advantage of looking in detail is that we often discover new things which we were not expecting to see, which, in the long run, gives us even more options for treating real people with heart disease, which is our ultimate goal.

Technical Summary

GPVI and alpha2beta1 are major collagen receptors on platelets which interact at different locations on the collagen molecule. GPVI binds to GPO motifs and alpha2beta1 to motifs containing GER. We have synthesized 57 peptides representing the full triple-helical portion of collagen III. Murine platelets bind efficiently to peptides III-04 and III-30 from this Toolkit. Binding to III-04 is dependent on GPVI and alpha2beta1, but binding to III-30 is only dependent on GPVI. We propose that the binding of platelets to III-04 involves the formation of a GPVI/alpha2beta1 receptor complex with previously unrecognised signalling consequences. We aim to confirm the formation of this complex, and characterise the signalling that results. Peptide III-30 represents a native GPVI binding sequence in collagen. We will identify which residues, in addition to the GPOs, contribute to the GPVI binding motif and how they interact with the binding site of GPVI.

Publications

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Farndale RW (2007) Platelet receptor recognition and cross-talk in collagen-induced activation of platelets. in Journal of thrombosis and haemostasis : JTH

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Guerrero JA (2008) In vivo relevance for platelet glycoprotein Ibalpha residue Tyr276 in thrombus formation. in Journal of thrombosis and haemostasis : JTH

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Jarvis GE (2013) A golden approach to ion channel inhibition. in Trends in pharmacological sciences

 
Title GpVI binding peptide 
Description Triple-helical GpVI-binding peptide derived from a native human collagen sequence 
Type Of Material Technology assay or reagent 
Year Produced 2007 
Provided To Others? Yes  
Impact Characterisation of the interaction between LAIR and collagen. Currently generating crystal structure for the peptide in collaboration. 
 
Description Babraham Institute, FcRgamma 
Organisation Babraham Institute
Country United Kingdom 
Sector Private 
PI Contribution Supply of genetically modified mice
Impact Publication: 18755982 Collaboration ongoing in other projects using our GM mice
Start Year 2007
 
Description Birmingham, UK 
Organisation University of Birmingham
Department College of Life and Environmental Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Signalling work carried out on ADAP project
Collaborator Contribution Collaborative work on a study investigating the role of ADAP in platelet function. Scientists in Birmingham have processed samples prepared in Cambridge.
Impact Collaborators in Birmingham have carried out intracellular signalling work to contribute to an ongoing study.
Start Year 2009
 
Description Little Rock, GpVI-/- 
Organisation University of Arkansas
Department University of Arkansas for Medical Sciences
Country United States 
Sector Academic/University 
PI Contribution Training in and conducting adhesion assays.
Collaborator Contribution Provision of genetically-modified mice
Impact Publication: 18339097 Data from GM mice in Little Rock has not yet been published.
Start Year 2007
 
Description Manchester, III-30 crystals 
Organisation University of Manchester
Department Faculty of Life Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Our laboratory supplies the peptides designed for crystallisation
Collaborator Contribution Collaborators carry out crystallisation and structural studies on triple-helical peptides
Impact To date, crystals have been obtained as well as preliminary diffraction data. The structural work is ongoing.
Start Year 2007
 
Description Pathology, Cambridge 
Organisation University of Cambridge
Department Department of Pathology
Country United Kingdom 
Sector Academic/University 
PI Contribution Supply of peptides and expertise in their use and experimental design.
Collaborator Contribution Supply of genetically-modified mice Use of flow cytometer
Impact Collaboration is ongoing. A paper was presented at the XXII ISTH in Boston in 2009.
Start Year 2007