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Counting functional IP3 receptors into the plasma membrane

Reference	BB/E004660/1
Principal Investigator / Supervisor	Professor Colin Taylor
Co-Investigators / Co-Supervisors
Institution	University of Cambridge
Department	Pharmacology
Funding type	Research
Value (£)	320,781
Status	Completed
Type	Research Grant
Start date	11/09/2006
End date	10/03/2010
Duration	42 months

Abstract

IP3 receptors (IP3R) are ubiquitously expressed and their role in mediating Ca release from intracellular stores is firmly established. Their roles in mediating Ca entry across the plasma membrane (PM) are more contentious. Using single-channel recording, we recently established that in B cells just two IP3R are reliably targeted to the PM where they provide the route for ~50% of the Ca entry evoked by activation of the B cell receptor. Irrespective of their expression level within the ER (400-13,000 IP3R), we reliably detect just ~2 IP3R within the PM. Using a combination of single-channel recording and expression of recombinant/mutant IP3R tagged to allow their detection with alpha-bungarotoxin, we will examine the features of the IP3R required for this precise 'counting' and the role of TrpC7 proteins. Our results will both establish how IP3R, perhaps in concert with TrpC7, are reliably counted into the PM in order to allow them safely to contribute to Ca entry, and more generally they will contribute to understanding how cells can reliably count such tiny numbers of proteins into a specific biological compartment.

Summary

The protective membrane that surrounds every cell provides the barrier needed to allow each cell to maintain a composition different from that of its environment, but it also isolates the cell interior from important extracellular signals. The latter problem is resolved by inserting proteins into the membrane that specifically recognise extracellular signals and then transduce them into intracellular signals capable of controlling cellular activity. Ca is the most important of these intracellular signals, not least because it is present at much lower concentration within the cell than on the opposite side of the membranes that either surround the cell or the organelles within it. The opening of Ca channels within these membranes is the most common means whereby extracellular stimuli evoke an increase in intracellular Ca concentration. For many years we have known that IP3 receptors are the most important and widely expressed Ca channels within the membranes of intracellular stores, but our recent work suggests that they are also functionally expressed in the membrane (PM) that surrounds the cell, where they provide the route for much of the Ca that flows into the cell from outside. To our great surprise, each cell absolutely reliably counts 1-3 IP3R into the PM from the many 1000s expressed within the cell. Such counting is important because otherwise IP3R in the PM might cause the cell to be overwhelmed with Ca and that would be toxic. But how can a cell so reliably count so few proteins into the membrane surrounding each cell? This project aims to answer that question.

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

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