Award details

Molecular mechanisms regulating intracellular targeting and trafficking of angiotensin-converting enzyme (ACE) and its novel homologue ACE2

Reference	BB/D001781/1
Principal Investigator / Supervisor	Professor Anthony Turner
Co-Investigators / Co-Supervisors	Professor Nigel Hooper, Professor Daniel Lambert
Institution	University of Leeds
Department	Inst of Molecular & Cellular Biology
Funding type	Research
Value (£)	199,229
Status	Completed
Type	Research Grant
Start date	01/09/2005
End date	31/08/2008
Duration	36 months

Abstract

A wide variety of proteolytic enzymes are expressed on the cell surface where they are poised to exert their physiological effects by modulating the activity of circulating peptides. Two such enzymes are the closely related metallopeptidases angiotensin-converting enzyme (ACE) and its close homologue, ACE2. Both enzymes are expressed by the epithelial cells of many tissues. Both proteins play important roles in the regulation of the cardiovascular system by modifying angiotensin peptides. Whilst much effort has gone into elucidating the biochemical properties and physiological roles of ACE and ACE2, surprisingly little is known of the mechanisms regulating their expression on the cell surface, particularly in the case of ACE2. Furthermore, no information is available on the intracellular targeting of either protein in polarised epithelial cells. Hence, we propose in this study to investigate the mechanisms regulating the cell surface expression and intracellular trafficking of ACE and ACE2 in polarised epithelial cells. We have preliminary data indicating that ACE and ACE2, when expressed heterologously in polarised MDCKII cells, have different membrane localisation. We intend to develop this study to another polarised epithelial cell line, Caco-2, and identify motifs within the proteins which determine their subcellular sorting. In addition we shall examine the mechanisms by which the levels of both proteins are regulated within the plasma membrane. To this end we already have evidence that ACE and ACE2 are proteolytically shed from the plasma membrane by distinct sheddases. We plan to investigate the molecular basis of this difference using the chimaeric proteins and deletion mutants outlined above. It is already known that ACE ectodomain shedding is influenced by phosphorylation of its cytoplasmic tail and ectodomain, and we will therefore determine, by metabolic labelling and immunoprecipitation, whether ACE2 is similarly regulated. In addition we shall seek toidentify any proteins which interact with ACE2 and modulate its subcellular distribution and/or ectodomain shedding, and whether the level of N-glycosylation is similarly important, as has been shown for ACE. The data obtained from these studies will provide long-awaited information regarding the cellular regulation of these two critical metallopeptidases.

Summary

The cells of the human body have to perform an array of different functions, often in very varied environments. In order to cope with this, cells in different environments have developed complex adaptations in their structure. One type of cell, termed an epithelial cell, covers the external surfaces of the body (including the surfaces of the lungs, intestines and other organs exposed to the outside environment). The surface of these cells which faces the outside environment has evolved to be quite different to the surface in contact with the cells surrounding it. These surfaces, termed the apical and basolateral membranes, respectively, have to perform very different functions. One of the ways in which they do this is to have different proteins present on the two membranes which carry out different functions. The surfaces of most cells contain a variety of proteins which are involved in sending messages to other cells, relaying messages to the nucleus of their own cell, or altering the function of other proteins (for example hormones) circulating in the blood. Two important proteins which perform the latter function in epithelial cells are the closely related angiotensin-converting enzyme (ACE) and angiotensin-converting enzyme-2 (ACE2). These proteins are present on the surface of epithelial cells and maintain blood pressure and heart function by modifying circulating peptides. Whilst a lot of effort has gone into investigating what ACE and ACE2 do, little is known about the processes which control where in an epithelial cell they are located, how they get there, and how, once there, they are regulated. We have evidence that despite their structural similarity, the membrane location of ACE and ACE2 in epithelial cells is quite different. In this study we aim to further investigate the processes controlling the cell surface expression of these two important proteins.

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search