Rac and PAK: a partnership at the interface between junction disassembly and increased cell motility

Lead Research Organisation: Imperial College London
Department Name: Life Sciences

Abstract

Tight adhesion between neighbouring cells plays an important role in the maintenance of tissue cohesion and architecture. During tumorigenesis, disruption of cell-cell adhesion is a key event for cell detachment from solid tumours and metastasis. We found that increased activity of Rac, a signalling protein, leads to a rapid destabilization of cell-cell contacts and increased migration and invasion of cells. Similar Rac hyper-activity is found in many different types of tumours and participate in malignant transformation. Here we propose to identify regulatory molecules that are recruited by Rac to help perturb cell-cell contact stability and investigate their potential as therapeutic targets during tumorigenesis.

Technical Summary

A major focus of research in cancer cell biology is to identify the regulatory mechanisms of the conversion of adherent cells in solid tumours into de-differentiated, highly invasive tumour cells. In many different tumour types there is a strong correlation between loss cadherin-dependent cell-cell contacts of cell-cell and increased metastatic potential.

We found that inappropriate Rac activation can disrupt cell-cell adhesion by inducing internalization and degradation of cadherins. Inappropriate activation of the small GTPase Rac is found in a broad range of tumours and result from different mechanisms commonly found in cancers. The mechanism via which Rac can perturb cell-cell contacts is not known. Interestingly, our recent results show that disruption of junctions by Rac can be reversed by inhibition of its target PAK and Rab11, a small GTPase that regulates early endocytic trafficking. This data is exciting in the context of tumour metastasis as Rac, PAK endocytic trafficking are also key players in cell motility. Consistent with our data, Rac and PAK have been implicated in increased motility and invasiveness of tumour cells. Thus the pathway identified in our study may participate in the coordination of junction destabilization and increased motility observed during metastasis.

The interplay between Rac, PAK and Rab11 is novel and significant to tumour progression. Here we will investigate the mechanisms required for junction disassembly and identify the:
1. Molecular mechanisms of destabilization of cadherin complexes by Rac and PAK;
2. Cellular processes by which Rac, PAK and Rab11 interfere with cell-cell contact stability;
3. Molecular mechanisms downstream of Rac and PAK that modulate Rab11 function.

This proposal will provide considerable insights into the intracellular mechanisms that lead to cadherin destabilization from junctions following Rac activation. Using biochemical and cell biology approaches, we expect to unravel a potential strategy to minimize cell detachment and de-differentiation during tumorigenesis.

Publications

10 25 50
 
Description Commentary on funding basic sciences
Geographic Reach National 
Policy Influence Type Citation in other policy documents
 
Description BHF Centre for Research Excellence
Amount £40,000 (GBP)
Organisation British Heart Foundation (BHF) 
Sector Charity/Non Profit
Country United Kingdom
Start 11/2009 
End 10/2010
 
Description Brunei PhD Studentship
Amount £151,000 (GBP)
Organisation Government of Brunei 
Sector Public
Country Brunei Darussalam
Start 02/2011 
End 01/2015
 
Description FP7 collaborative grant
Amount £275,743 (GBP)
Organisation European Commission 
Department Seventh Framework Programme (FP7)
Sector Public
Country European Union (EU)
Start 11/2008 
End 04/2012
 
Description Wellcome Trust Project grant
Amount £263,000 (GBP)
Funding ID WT094371MA 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 07/2011 
End 12/2014
 
Title Development of Machine Learning methods to quantify junction phenotypes driven by different stimuli (2017) 
Description A major road block in the analysis of junction phenotypes is the ability to recognize cell borders in a monolayer of epithelial cells, which is painstakinly done manually and with enorumous time invested in each image. Such detailed analysis is not feasible for high-throughput analysis in screens. Training datasets teach computer how to identify cell-cell contacts, which can then tbe deployed in large datasets with minimal input from user. Development of the program was done using a large dataset of images obtained under different conditions obtained via the distinct grants funding my research. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? No  
Impact Methodology is currenly being validated. It will be made available after thorough optimization, validation and adding a user-friendly interface. 
 
Title Quantitative fluorescence analysis 
Description New techniques to quantify fluorescence intensity 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact Increased the efficiency and accuracy of analysis 
 
Title Rab7 activation assay 
Description Methodology to detect levels of active Rab7 in cells following different treatment (growth factor stimulation or expression of different constructs) 
Type Of Material Technology assay or reagent 
Year Produced 2011 
Provided To Others? Yes  
Impact This methodology will allow dissecting - signalling processes that regulate Rab7 - mechanisms of lysosomal biogenesis and function - degradation 
 
Description Ahmadian - Armus 
Organisation Max Planck Society
Country Germany 
Sector Charity/Non Profit 
PI Contribution Technical and conceptual input - RabGAP assays
Collaborator Contribution novel techniques, visits from people in my lab to Germany
Impact paper Armus Current Biology
Start Year 2008
 
Description Bailly - PAK phosphorylation substrates 
Organisation University College London
Department Institute of Ophthalmology UCL
Country United Kingdom 
Sector Academic/University 
PI Contribution We identified a PAK1 substrate as Ajuba, a protein that localizes at junctions and regulates junction stability. BAilly's lab performed studies to address which cytoskeletal functions Ajuba has to help stabilization of cell-cell contacts.
Collaborator Contribution experiments performed in their lab; intellectual output
Impact Paper in press in JCB - Ajuba cross-talk Rac1 and PAK1
Start Year 2008
 
Description Longmore - Ajuba project 
Organisation Washington University in St Louis
Country United States 
Sector Academic/University 
PI Contribution We have provided the model system (epithelial cell-cell adhesion) and Dr. Longmore provided the reagents to investigate Ajuba function
Collaborator Contribution Two joint publications (2003, 2005) exchange of reagents, etc.
Impact Publications: 15728191 12417594
 
Description Ridley - RhoE 
Organisation King's College London
Department School of Biomedical Sciences KCL
Country United Kingdom 
Sector Academic/University 
PI Contribution We provided the model system (keratinocyte differentiation) and Prof Ridley provided reagents and expertise in RhoE
Collaborator Contribution Reagents, publications, dissemination of work
Impact Publication: 18923151
Start Year 2006
 
Description Sean Sun - Modelling of biophysical parameters that generate a mature cell-cell contact 
Organisation Johns Hopkins University
Country United States 
Sector Academic/University 
PI Contribution We provided the biological question and image datasets.
Collaborator Contribution Sean Sun - Hopkins Universitymodeller
Impact Manuscript submitted: Ranjan, K.; Alonso-Mardones, JS; Swiatlowska, P; Liu, S.;Rothery, S.; M. Stevens; Sun,S.; Gorelik, J & Braga, VMM. Cell confinement impacts on cortical elasticity, configuration and dynamics of junctions
Start Year 2015
 
Description Invited lecture "Pioneers in Cell Dynamics and Imaging", Cells in Motion Clusters of Excellence, University of Muenster, Germany 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Seminar presented at University of Munster
Year(s) Of Engagement Activity 2019
 
Description Key note speaker 3rd World Congress on Cancer Biology and Immunology. Milan Italy 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact presentation data & results
Year(s) Of Engagement Activity 2019
 
Description Lecture "Pioneers in Cell Dynamics and Imaging", Cells in Motion Clusters of Excellence, University of Muenster, Germany. 2018 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact networking
Year(s) Of Engagement Activity 2018
 
Description STEM World Summer School; Outreach at Imperial 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact About 15 international students (15-18 years old). We organised a half-day session to highlight the changes in cell behaviour that accompany tumour cell transformation, including microscope demonstration of cell preparation.

raise interest in academic research in medicine and biology
Year(s) Of Engagement Activity 2011
 
Description Talk Oncobiology Symposium, Brazil 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact networking, share ideas
Year(s) Of Engagement Activity 2017
 
Description Talk at Brazilian Embassy Networking event 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact Workshop "Innovation in Healthcare" - discussion on engagement and collaborations - how is it best fomented?
Year(s) Of Engagement Activity 2017
 
Description Talk conference UK membrane trafficking 2018 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Postgraduate students
Results and Impact Networking, discussion concepts and collaborations
Year(s) Of Engagement Activity 2014,2018
 
Description Workshop Women in Science 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Women Scientists discussing their career progression in Academia, what worked well, what could have been different
Year(s) Of Engagement Activity 2009
 
Description invited speaker Liverpool 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact invited to give a seminar at the Dept Cellular and Molecular Physiology, Liverpool, UK

networking with colleagues at the Dept.
Year(s) Of Engagement Activity 2012
 
Description invited speaker Martinsried, Germany 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact invited to give a seminar at the Dept Biochemistry, Max Planck Institute, Martinsried, Germany

finalise discussions for collaborative manuscript publication
Year(s) Of Engagement Activity 2012
 
Description speaker at Autophagy interest research meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Autophagy in London Special Interest Group, LIR Cancer Research Institute, London, UK

netwroking with autophagy academics - discussion of results
Year(s) Of Engagement Activity 2010,2012
 
Description talk Regulation and Function of Small GTPases, FASEB Conference, Miami - USA. 2017 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact networking
Year(s) Of Engagement Activity 2017