Award details

Regulation of tight junction assembly and function by the heat shock protein Apg-2

Reference	BB/C514458/1
Principal Investigator / Supervisor	Professor Karl Matter
Co-Investigators / Co-Supervisors	Professor Maria Balda
Institution	University College London
Department	Institute of Ophthalmology
Funding type	Research
Value (£)	244,675
Status	Completed
Type	Research Grant
Start date	01/04/2005
End date	30/06/2008
Duration	39 months

Abstract

Epithelial cells interact with their neighbours and the extracellular matrix using molecular complexes composed of transmembrane proteins and distinct sets of cytoplasmic plaque proteins that serve as connections to the cytoskeleton. These proteins form a complex protein network composed of multimeric protein complexes. The proper establishment of these interactions is essential for the normal development and differentiation of polarised epithelia. Their dysfunction and destabilisation critically contributes to many diseases such as epithelial tumorigenesis and metastasis. The most apical structures of the epithelial junctional complex that joins individual cells in epithelial tissues are tight junctions. Tight junctions function as diffusion barriers that restrict transepithelial diffusion of solutes and participate in the regulation of the signal transduction pathways that control cell proliferation and differentiation. This proposal focuses on Apg-2, an Hsp-110 family member that we have identified since it binds to the SH3 domain of ZO-1, a tight junction-associated adaptor. ZO-1 also binds to the junction-associated transcription factor ZONAB, which binds to the SH3 domain as well. Our unpublished data suggest that Apg-2 negatively regulates the ZO-1/ZONAB interaction and thereby functions as a stimulator of ZONAB. Moreover, preliminary data based on cell lines that allow the regulated depletion of Apg-2 expression by RNA interference suggest that Apg-2 regulates junction assembly and becomes recruited to cell-cell junctions under conditions that destabilise tight junctions such as heat shock. We therefore postulate that Apg-2 functions as a factor that guides tight junction assembly (similar to classical molecular chaperones) and that this function becomes stimulated during conditions that cause junctional destabilisation. Our first two aims focus on the role of Apg-2 in junction assembly and the formation of different types of ZO-1-based multimeric protein complexes during normal junction assembly and during heat shock, a stress condition that destabilises the junctional complex. These experiments are largely based on biochemical assays to study protein complex formation, and immunofluorescence as well as function assays to monitor recruitment of proteins to the junctional complex and assembly of functionally active tight junctions. The third aim builds on other preliminary observations obtained with depleted cell lines, which suggest that Apg-2 and ZO-1 modulate the cellular stress response during heat shock and hyperosmotic stress. We propose a set of experiments based on techniques such as reporter gene assays, tests of MAP kinase activation and signalling, as well as cDNA arrays to determine how tight junctions regulate the cellular stress response and how Apg-2 activates ZO-1ZONAB signalling in response to heat shock and hyperosmotic shock. The expected results will help us to understand how the complex protein networks that form intercellular junctions assemble and how they signal to cells during cellular stress conditions to maintain epithelial tissue integrity.

Summary

unavailable

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