Professor Elizabeth Smythe

LizProfessor of Molecular Cell Biology
Director, Centre for Membrane Interactions and Dynamics

Department of Biomedical Science
The University of Sheffield
Western Bank
Sheffield S10 2TN
United Kingdom

Room: C09 Florey building
Telephone: +44 (0) 114 222 4635
Email: e.smythe@sheffield.ac.uk

General

Career history

  • 2002-present: Professor of Molecular Cell Biology, University of Sheffield
  • 1992-2002: MRC Senior Fellow and Principal Investigator, Wellcome Trust BioCentre, University of Dundee
  • 1989-1992: NATO/SERC postdoctoral fellow, The Scripps Research Institute, California
  • 1986-1989: Postdoctoral research assistant, University of Dundee.
  • 1986: PhD, Trinity College, Dublin
  • 1982: B.A. (Mod), Trinity College, Dublin

Research interests

Our lab is interested in the molecular mechanisms of cargo sorting along the endocytic pathway with particular emphasis on the regulation of the clathrin coated vesicle cycle by rab5 and reversible phosphorylation. We are also interested in the interplay between endocytic trafficking and signaling pathways.

My research group is part of the Centre for Membrane Interactions and Dynamics (CMIAD).

CMIAD

Current activities

Member of:

  • the Faculty of 1000 (2001-)
  • the Editorial Board of the Biochemical Journal (2004-2007) Renewed (2007-2011)
  • the MRC College of Experts (2006-2010)
  • Executive Secretary of the British Society for Cell Biology (2006-2011)

Funding

  • MRC
  • BHF
  • EU

Recent publications

Full publications list

Research Overview

Functions of membrane microdomains on the endocytic pathway: integration of endocytosis and signalling

We are interested in the assembly and dynamics of membrane microdomains that are specialised for signalling on the endocytic pathway. Clathrin coated pits are microdomains responsible for the uptake of a wide variety of cargo. Using live-cell microscopy we are exploring how signalling cargo modulates clathrin coated pit dynamics via posttranslational modifications of the clathrin coat and how this impacts on downstream signalling.

The small GTPase, rab5, regulates many events on the early endocytic  pathway including cargo selection, vesicle uncoating, endosomal fusion, and signalling. Guanine nucleotide exchange factors (GEFs) convert rab5 to its active GTP-bound ‘on’ state and they are also proposed to drive the spatial and temporal assembly of rab5 functional domains. We are using a variety of biochemical and morphological approaches to explore the function of Rme-6, a GEF that acts early in the endocytic pathway to integrate trafficking and signalling.

During embryonic development a surprisingly small number of signals are used repeatedly to form a whole organism. These same signals are also used in homeostasis. This means that cells need to respond to signals in a context specific manner. We are exploring how endocytosis contributes to context specific signalling using JAK/STAT signalling in Drosophila cells as a paradigm.

In addition to the molecular cell biology approaches discussed above we wish to understand how biophysical properties of membranes, in particular membrane tension, contributes to the regulation of microdomain formation. In the longer term our aim is to translate our findings into 3-D systems and ultimately whole organisms to understand the relationship between endocytosis and signalling.

Figure 1

Positions Available

PhD Opportunities

1. Rme-6 as an integrator of endocytosis and signalling in cancer cells

The endocytic pathway regulates intracellular signalling in a variety of ways and when there are defects in this cross-talk, this can give rise to a variety of diseases including cancer. Rab5 is a major regulator of the early endocytic pathway, which cycles between an inactive, cytoplasmic GDP form and an active membrane-associated GTP form. Rab5 guanine nucleotide exchange factors (GEFs) catalyse the conversion of rab5 into its active form where it interacts with a variety of effectors which allow it to perform many different cellular functions, including signalling (Stenmark, 2009).

Our lab is particularly interested in Rme-6 which is a rab5 GEF that integrates signalling and trafficking of receptor tyrosine kinases such as EGFR and Tie2 by modulating flux through the endocytic pathway. Using SILAC mass spectrometry we have identified the interactome of Rme-6 in HeLa cells following EGF stimulation and have identified a number of novel binding partners.

The aim of this PhD project will  be to extend our current understanding of Rme-6 in the integration of endocytosis and signalling through examining the role of these novel binding partners of Rme-6 that interact in a cargo specific manner.

The project will utilise a variety of molecular cell biology approaches, including CRISPr technology to knockout genes of interest as well as knocking in tagged wild-type and mutant proteins, endocytic and signalling assays as well as high resolution light microscopy in fixed and living cells. There will be particular emphasis on the role of Rme-6 in cancer cells.

References

Stenmark, H.
Rab GTPases as coordinators of vesicle traffic.
Nat Rev Mol Cell Biol. 2009 10:513-25.

Semerdjieva S, Shortt B, Maxwell E, Singh S, Fonarev P, Hansen J, Schiavo G, Grant BD, Smythe E.
Coordinated regulation of AP2 uncoating from clathrin-coated vesicles by rab5 and hRME-6.
J Cell Biol. 2008 183:499-511.

Keywords: Cell Biology / Development

2. Structural analysis of Rme-6, a rab5GEF that integrates endocytosis and signalling

Co-Supervisor: Professor Per Bullough

The ras family of small molecular weight GTPases act as molecular switches. In their active GTP conformation, they interact with a variety of effectors to perform a range of physiological functions. Conversion to the GTP conformation is mediated by guanine nucleotide exchange factors (GEFs) while GTPases are inactivated by GTP hydrolysis, facilitated by GTPase activating proteins (GAPs). Ras is the founding member of this family and has many roles in intracellular signalling and mutations in ras can result in cancer.

The rab family of small GTPases regulates many aspects of membrane trafficking and rab5 is considered to be a master regulator of the early endocytic pathway (Stenmark, 2009). Rme-6 is a multidomain protein containing an N-terminal rasGAP domain and a C-terminal rab5 GEF domain connected by a flexible linker. We have evidence that Rme-6 modulates endocytic flux of EGFR resulting in modulation of its downstream signalling by acting as a signalling scaffold.

We have explored the structure of hRme-6 and our recent results have shown that hRme-6 exists as a higher order multimer in vivo and in vitro. Furthermore we have shown its GEF and GAP activities are autoinhibited and require specific spatial activation in cells (2). Recent studies have identified the hinge region as key to regulation of Rme-6 conformation and relief of autoinhibition. We can purify Rme-6 and negative staining and EM analysis have provided a preliminary structure.

The aim of this project will be to refine this structure using cryo-EM approaches. We are particularly existed to understand the role of the hinge region in effecting conformational changes in Rme-6.

References

Stenmark, H.
Rab GTPases as coordinators of vesicle traffic.
Nat Rev Mol Cell Biol. 2009 10:513-25.

Semerdjieva S, Shortt B, Maxwell E, Singh S, Fonarev P, Hansen J, Schiavo G, Grant BD, Smythe E.
Coordinated regulation of AP2 uncoating from clathrin-coated vesicles by rab5 and hRME-6.
J Cell Biol. 2008 183:499-511.

Keywords: Biochemistry, Cell Biology / Development, Structural Biology

For more information about these projects and how to apply, see our PhD Opportunities section.

PhD Opportunities