Moonlighting functions of endocytic proteins during the cell cycle

Lead Research Organisation: University of Warwick
Department Name: Warwick Medical School

Abstract

Cells can be thought of as islands - they are closed to the outside world because they have a plasma membrane that doesn't let anything through. However, cells have evolved tiny portals for things to enter in a tightly regulated way. These portals are working all the time, but they are shut down when the cell needs to divide. It is important that no mistakes are made at this time. Otherwise the cell could die or it could start to grow in an unusual way leading to disease e.g. cancer. Nobody knows how this shutdown happens. We want to study the portals using a machine that can tell us all the pieces that make up a portal. If we compare portals from normal cells with those that are dividing this will tell us which pieces are missing in dividing cells. We can then find out how the cell takes these pieces away at the start of cell division. We think that this work is important to do. These portals are hijacked by viruses: they use portals as a way to infect cells. If we knew how the cell shuts down the portals, we could think of ways to switch them off using drugs. These drugs could be used as anti-viral treatments.

Technical Summary

Clathrin-coated vesicles (CCVs) are formed continuously in non-dividing cells. When the cell enters mitosis, membrane trafficking is shut down. This phenomenon has been acknowledged for almost fifty years, but the mechanism behind the shutdown has been enigmatic. It is thought that certain components of the clathrin coat are phosphorylated during mitosis and this prevents their engagement with the network of proteins responsible for driving CCV formation. In addition, there is an emerging concept in cell biology of proteins with alternative functions during mitosis. In this project, we will investigate the link between those proteins that are missing from coats in mitotic cells, those that are phosphorylated and those that carry out alternative functions. We propose a systematic and quantitative proteomic study of clathrin-coated membranes from cells in interphase and in mitosis. This will allow us to identify the proteins that are less abundant in clathrin coats from mitotic cells. Using live-cell imaging, we will determine at what stage in the cell cycle these proteins cease to be recruited to the coat and also investigate where in the cell the protein is localised during this time. Finally, we will discover whether these proteins are phosphorylated and whether or not this modification is responsible for the mitotic shutdown of CCV formation.

Planned Impact

The major stakeholders impacted by this research are: 1. International scientific community: cell biologists and biochemists 2. Local scientific community: systems biology research in Liverpool 3. Commercial sector Our strategy for engaging with these stakeholders is outlined in the Impact Plan.

Publications

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Fielding AB (2013) Mitotic inhibition of clathrin-mediated endocytosis. in Cellular and molecular life sciences : CMLS

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López-Murcia FJ (2014) Presynaptic clathrin levels are a limiting factor for synaptic transmission. in The Journal of neuroscience : the official journal of the Society for Neuroscience

Related Projects

Project Reference Relationship Related To Start End Award Value
BB/H015582/1 01/12/2010 31/01/2013 £187,132
BB/H015582/2 Transfer BB/H015582/1 22/07/2013 31/08/2014 £74,156
 
Description All the time, cells take up chunks of plasma membrane and recycle them. They use this recycling system to control many aspects of cell function: signalling, degradation, nutrient uptake etc. When cells begin to divide they switch off the uptake process. This had been known for half a century, but nobody knew how this happened. We have discovered the mechanism that cells use to inhibit this activity during cell division.
It turns out that when cells divide they swell so that their plasma membrane is under greater tension. Higher tension is a problem for taking up chunks of membrane, but the cell has a way around this which is to use its cytoskeleton to help uptake. However, in mitotic cells this is not available and so we see a decrease in uptake.
Exploitation Route Cells that are rapidly dividing, e.g. in cancer are less likely to take up drugs that enter via this pathway. The mechanism we have discovered is an opportunity to restart the uptake pathway to help drug delivery to rapidly dividing cells.
Sectors Pharmaceuticals and Medical Biotechnology

 
Description Our work has been well received in the academic sector. Papers arising from this award have been well-cited.
First Year Of Impact 2014
 
Description BBSRC Alert14 (50% University of Warwick)
Amount £1,490,400 (GBP)
Funding ID BB/M01228X/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2015 
End 01/2016
 
Description Open Access from a PI's perspective, Crick Institute Retreat 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact EuropePMC and I presented on how to make your research outputs freely accessible and how to disseminate outcomes widely

Lots of discussion followed the event at the reception and via email and twitter afterwards
Year(s) Of Engagement Activity 2014
 
Description Open Access from a Researcher's perspective, EuropePMC funder's Group, British Library 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Around 30 representatives from groups that fund EuropePMC assembled for the day to listen to developments at the repository. I talked about Open Access from a researcher's perspective.

Lively discussion with participants. Involvement in future events and studies on research outcome sharing.
Year(s) Of Engagement Activity 2012