Analysis of the roles of the TRPV2 calcium channel.. (TRPV2PI3KMacrophage)
Analysis of the roles of the TRPV2 calcium channel in inflammation
Start date: 02 Mar 2009,
End date: 01 Sep 2011
Macrophages are key players in the front line of our immune system, responsible primarily for the recognition and destruction of bacterial and fungal pathogens. However, they are also involved in various inflammatory pathologies. Membrane receptors of chemokines control many aspects of macrophages functions by stimulating various 2nd messenger systems, important amongst these are PI3Ks and Ca2+. However the identity of the Ca2+ channels involved in Ca2+ influx and the cellular responses regulation by that Ca2+ are still poorly understood. The TRPV2 calcium channel is expressed in immune cells, up-regulated during their activation, interacts with some proteins involved in the regulation of the actin cytoskeleton and plays a role in chemotaxis. Moreover, PI3K can regulate TRPV2 activity. The complementary objectives of the project will be to i) define the PI3K-dependant signalling pathways leading to the translocation of TRPV2 to the plasma membrane and its activation, ii) determine the physiological roles of the channel in a native environment. Mouse primary macrophages expressing either i) a TRPV2-GFP construct containing an extracellular flag tag or ii) specific shRNA against TRPV2 will be obtained using the innovative technique of adoptive transfer of haematopoietic stem cells transduced with appropriate retrovirus. Thus we can analyse the subcellular localisation of the channel and its translocation under various conditions. The physiological functions of the channel in macrophages will be precisely analysed in TRPV2-suppressed macrophages using techniques routinely used in the host laboratory. This work will be the first analysis of the endogenous TRPV2 in primary immune cells and will determine if TRPV2 is an important component of the pro-inflammatory signalling networks of macrophages and hence may be a potential target for novel anti-inflammatories.
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