Membrane Biology

Professor Tony Magee, Head of Group

Dr Martin Spitaler, FILM Facility Manager 

The Membrane Biology group is studying the functional role of lipid modifications of proteins, focussing on signal transduction pathways at the cell surface.  Our studies utilise a combination of biochemistry, immunology, molecular and cell biology, and state-of-the-art microscopy.

We have previously defined a role for the N-terminal dual acylation motif of Src family of protein tyrosine kinases (PTKs) in their targeting to glycolipid/cholesterol-enriched “lipid rafts” in the plasma membrane Figure 1. These are highly dynamic metastable lipid domains that concentrate lipid-modified proteins in both the outer leaflet (e.g. GPI-anchored proteins) and inner leaflet (e.g. Src PTKs, Ras) of the membrane.  We have shown that these structures are intimately involved in signal transduction from the T cell antigen receptor (TCR).  Confocal and multiphoton microscopes enable rafts to be imaged in live cells in real time during signalling events.  In collaboration with Profs. Paul French and Mark Neil, Dept of Physics and Prof. Dan Davis, Division of Cell & Molecular Biology, we have used fluorescence lifetime (FLIM) measurements to probe the local environment of lipid raft domains.  We are also studying the role of glycosphingolipids in T cells in TCR signalling (collaboration with Dr. Liz Jury, UCL, and Dr. Terry Butters, University of Oxford).  In collaboration with Prof. John Seddon and Dr. Rob Law, Dept. of Chemistry, we are studying fundamental properties of model lipid raft membranes.  Our in vitro studies have been extended to the in vivo situation by analysing the dynamics of rafts during the process of contact formation between T cells and their targets.

Two families of putative protein acyltransferases (PATs) have been identified in recent years, acting on intracellular proteins (DHHC-CRD family) and extracellular secreted signalling proteins of the Hedgehog and Wingless families (e.g. Hhat and Porc respectively, MBOAT family proteins) Figure 2.

We have determined the expression pattern of the DHHC-CRD family (23 members in man) in haematopoietic cell types and used RNAi to assess their role in the S-acylation of the key T cell tyrosine kinase Lck.  We have identified the DHHC2 protein as a PAT for Lck and raised antibodies to it (with Dan Davis).  The role of regulated S-acylation of proteins is also being addressed; we have raised specific antibodies to acylprotein thioesterases for use in studies of deacylation of S-acylated proteins.

 

We are also interested in acylation and cholesteroylation of secreted signalling proteins, e.g. human sonic hedgehog (Shh), and have raised antibodies to the multispanning membrane protein Hhat, the PAT for Shh.  We are analysing its function in Shh secretion and activity.  As Shh is involved in the growth of many tumours Hhat is a potential drug target for human cancer, including Pancreatic Ductal Adenocarcinoma (PDAC); some of our work on PDAC is funded by the Pancreatic Cancer Research Fund (PCRF http://www.pcrf.org.uk/). These studies have been greatly facilitated by collaboration with Dr. Ed Tate (Dept. of Chemistry) on the use of click chemistry with fatty acid and cholesterol analogues to allow rapid analysis of acylated and cholesteroylated proteins.

Membrane Biology group members:

Post-docs: Dr. Antonios Konitsiotis (PCRF-funded) and Dr. Martin Spitaler (FILM Facility Manager)

PhD students: Biljana Jovanovic, Mirella Koleva, Arwen Tyler

MRes/PhD students: Paulina Ciepla and Naoko Masumoto