Actin assembly in filopodia and lamellipodia: Regulation of the Arp2/3 Complex by Scar and IRSp53

Lead Research Organisation: Beatson Institute for Cancer Research
Department Name: Research Group Y35 (Karen Blyth)

Abstract

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Technical Summary

We aim to determine how actin assembly is regulated in response to activation of various receptors, including during cell spreading. In particular, we want to know how the actin nucleating complex, Arp2/3, is controlled by Scar proteins and by IRSp53 to regulate the balance between filopodia and lamellipodia assembly in platelets and fibroblasts. We have four main aims. Firstly, we have recently discovered that Scar1 null mice have abnormal platelets that do not spread properly on extracellular matrix. Thus, Scar1 appears to be an essential controller of actin dynamics in response to integrin engagement in platelets. We aim to fully characterise the role of Scar1 in platelets. Secondly, we have also found abnormalities in Scar1 null fibroblasts that are distinct from those previously reported by another group. We will determine the role of Scar1 in fibroblast spreading and responses to growth factors using a combination of imaging and biochemical methods. Thirdly, we will also investigate connections between Scars and the exocytic machinery, specifically, the interaction between Scar2 and Sec3. Fourthly, we have solved the crystal structure of a key actin bundling protein and ligand of Scar1, IRSp53 IMD. We will determine how IRSp53 connects to Scar proteins as a mediator of filopod assembly using biophysical and cell biological methods. Our studies are relevant to haemostatic diseases, cancer, inflammation, neurological diseases and wounding. Arp2/3 complex subunits have been implicated in gastric cancers (Kaneda, 2002 #229); Scar proteins may be targets for congenital neurological diseases and cell migration/motility are essential to all of these processes (Kitamura 2003a, 2003b).

Publications

10 25 50
 
Description AICR Project Grant
Amount £100,000 (GBP)
Funding ID 110-119 
Organisation Association for International Cancer Research 
Sector Charity/Non Profit
Country United Kingdom
Start 02/2010 
End 02/2013
 
Description AICR project grant
Amount £170,000 (GBP)
Organisation Association for International Cancer Research 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2010 
End 03/2013
 
Description BHF Project Grant
Amount £100,000 (GBP)
Funding ID FS/09/034/27756 
Organisation British Heart Foundation (BHF) 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2009 
End 10/2012
 
Description CRUK Institute Programme Beatson Institute
Amount £900,000 (GBP)
Funding ID 15673 
Organisation Cancer Research UK 
Sector Charity/Non Profit
Country United Kingdom
Start 11/2012 
End 11/2017
 
Description CRUK Institute Programme Beatson Institute
Amount £900,000 (GBP)
Organisation Cancer Research UK 
Sector Charity/Non Profit
Country United Kingdom
Start 11/2007 
End 11/2012
 
Title Antibodies 
Description We have created several useful antibody reagents that are now commercially available- please see "collaborations section" 
Type Of Material Antibody 
Year Produced 2006 
Provided To Others? Yes  
Impact This material has helped many groups achieve primary research goals, such as publications and new knowledge about the actin cytoskeleton and cell motility. 
 
Title DNA expression plasmids 
Description We have developed hundreds of DNA expression plasmids for research use. We routinely make these available to other researchers upon request. We receive around 50 requests per year for our plasmids. 
Type Of Material Technology assay or reagent 
Year Produced 2006 
Provided To Others? Yes  
Impact Many scientific publications and discoveries have arisen from use of our DNA reagents. We have also developed numerous collaborations with other groups around the world. 
 
Description Collaboration with Ines Anton, CNB Madrid 
Organisation Spanish National Centre for Biotechnology
Country Spain 
Sector Public 
PI Contribution A student from Ines lab, Esther Garcia, came to our group for several months on an EMBO fellowshipvand a fellowship from the CNB in Spain to study the role of WIP in cell migration and invasion in breast cancer cells. This has led to several publications involving Ines Anton and Esther Garcia. Our group trained Esther in doing cell invasion assays and measuring migration quantitatively.
Collaborator Contribution Ines and Esther provided expertise to study WIP and in the study of breast cancer cells.
Impact We have had 3 publications so far and a 4th has just been accepted in the Journal of Cell Science
Start Year 2011
 
Description Collaboration with Ruoss and Bender 
Organisation University Hospital Zürich
Country Switzerland 
Sector Hospitals 
PI Contribution We will provide knockout mice for cytoskeletal genes that this group will study to determine whether the platelets are affected. My work with the BHF funded projects on platelets was hugely important to my being a part of this new collaboration with the hospital lab in Wuerzburg. I am the co-supervisor of the PhD student who is doing this project, so I will provide supervision and expertise throughout her PhD.
Collaborator Contribution Our partners will perform research to determine the effect of deletion of cytoskeletal genes (CYFIP1 and Strumpellin) on platelet and megakaryocyte development and function. They will perform most of the wet lab work and we will provide expertise and reagents as needed.
Impact No outputs yet.
Start Year 2015
 
Description Development of antibodies to Arp2 and Arp3 
Organisation Sigma-Aldrich
Country United States 
Sector Private 
PI Contribution We developed the antigen expression in E coli and we tested the antiserum for the company
Collaborator Contribution These antibodies are made available to our group free of charge
Impact We developed antibodies to Arp2 and Arp3 subunits of the Arp2/3 complex with Sigma Israel. This collaboration has led to two very useful reagents that are now available both to our group and to the wider research community.
 
Description Development of antibodies to p34-Arc 
Organisation Upstate Biotechnology, Inc.
Country United States 
Sector Private 
PI Contribution We developed an antibody against p34-Arc of the Arp2/3 complex and tested antiserum for the company.
Collaborator Contribution It has maintained a supply of excellent antibody that we can use free of charge for our work.
Impact We have developed a polyclonal antibody that is now sold by Upstate Biotech (Millipore). This is extremely useful to us, as it also ensures a long-term supply of a valuable reagent to our group.
Start Year 2006
 
Description Drug discovery: fascin and metastasis 
Organisation MRC-Technology
Country United Kingdom 
Sector Private 
PI Contribution WE have set up in vitro and in vivo models to study fascin function in invasion and migration as well as cancer metastasis. We have set up and performed assays on candidate compounds. We have provided expertise and DNA/protein reagents as well as cells to both Beatson Drug Discovery and MRC-T.
Collaborator Contribution We have set up a programme between Beatson Drug Discovery, my group and MRC-T to screen for new inhibitors of fascin-1, a protein implicated in migration and metastasis in epithelial cancers. Each group has contributed experimental work, reagents and intellectually to this collaboration.
Impact This collaboration has been very central to my research programme on invasion and metastasis. It is multidisciplinary and involves chemistry, biophysical studies (X-ray crystallography and NMR), cell biology and mouse genetic studies.
Start Year 2010
 
Description Use of patterned substrates for the study of cell polarity 
Organisation CYTOO
Country France 
Sector Private 
PI Contribution We will be given free samples of materials from CYTOO to study cell polarity on micro-patterned cell matrix coverslips. We will be a beta test site for these materials. This will allow us access to these state-of-the art materials which may be very valuable for our research.
Collaborator Contribution We will be given free samples of materials from CYTOO to study cell polarity on micro-patterned cell matrix coverslips. We will be a beta test site for these materials. This will allow us access to these state-of-the art materials which may be very valuable for our research.
Impact We were given free samples of materials from CYTOO to study cell polarity on micro-patterned cell matrix coverslips. We will be a beta test site for these materials. This will allow us access to these state-of-the art materials which may be very valuable for our research.
Start Year 2009
 
Description CRUK Relay for life 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact I gave a talk to cancer survivors and to all interested members of the public - e.g. families and supporters- about basic scientific research on cell migration and its importance for medical research and disease studies.

I have volunteered to do this activity every year. I don't think you can measure the impact, but I hope that it is obvious that communicating to the public about cell motility research and how it is important for medical issues such as cancer will have an impact on society.
Year(s) Of Engagement Activity 2008,2009
 
Description Meeting with CRUK charity workers 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact 10 volunteers attended a talk and tour of the institute and we had a coffee time for discussion and questions

We received positive feedback form the volunteers about the day
Year(s) Of Engagement Activity 2009,2010
 
Description Visit to Undergraduate College Alma, MI, USA 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Undergraduate students
Results and Impact 100 undergraduates and several faculty at this undergraduate college in Michigan attended my seminar about cancer metastasis and cell migration. We also held a careers workshop, which was attended by more than 100 first year undergraduates. My talks hopefully inspired these students to consider a career in scientific research.

We will set up an internship for an Alma College student to visit my laboratory here in Glasgow in the summer and receive training and experience in cell biology and cancer research.
Year(s) Of Engagement Activity 2012