Sweet taste receptor interacting protein CIB1 is an inhibitor of InsP3-mediated Ca2+-release

Life Sciences 2007 (2007) Proc Life Sciences, PC515

Poster Communications: Sweet taste receptor interacting protein CIB1 is an inhibitor of InsP3-mediated Ca2+-release

J. K. Hennigs1, N. Burhenne2, H. Schmale1

1. Institute for Molecular Cell Biology, Hamburg, Germany. 2. Institute for Cell Biochemistry and Clinical Neurobiology, Hamburg, Germany.

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In previous works of our group the calcium and integrin binding protein CIB1 has been identified and characterised as a specific interaction partner of the sweet taste receptor subunit TAS1R2 carboxyterminal domain. Sweet and umami taste is mediated by three G-protein-coupled subfamily C receptors of the TAS1R-family, namely TAS1R1, TAS1R2, and TAS1R3. These receptors feature the typical large N-terminal “venus-fly-trap module“ as well as a short intracellular C-terminus and function only as (hetero-)dimers recognising different classes of taste stimuli. While in humans the TAS1R1/TAS1R3 complex triggers mainly the perception of umami taste (taste of sodium glutamate), the TAS1R2/TAS1R3 exhibits the actual sweet receptor recognising natural ligands like sugars, D-amino acids, sweet tasting proteins and artificial sweeteners. The intracellular signalling cascade seems to involve a yet unidentified heterotrimeric G protein, phospholipase C-β2 generating InsP3, InsP3-receptor subtype 3 and the cation-conducting TRPM5 ion channel [1]. In the heterologous HEK293 sweet taste expression system the G protein chimera Gα16gust44 links the sweet taste receptor dimer to an InsP3-dependent Ca2+-release pathway. To further characterise the in vivo role of CIB1, which is endogenously expressed in HEK293 cells, we generated cell lines stably and transiently expressing shRNA against CIB1. Investigating the general aspects of CIB1 influence on intracellular calcium signalling, various imaging systems were used, .e.g. FLIPR and FURA-2 cuvette assays. Applications of ATP, UTP and carbachol – ligands which increase intracellular calcium in an InsP3– dependent manner via endogenous receptors, revealed a more pronounced Ca2+-elevation in HEK293 cells lacking CIB1 than in control cells. Overexpression of CIB1 lowered the Ca2+ response to UTP in comparison to mock transfected cells. Preincubating these cells with the SERCA inhibitor thapsigargin showed that the assayed changes of cytosolic calcium were due to its release from the endoplasmic reticulum. In addition to that, HEK293 cells transiently overexpressing CIB1 also showed an increased release of calcium from intracellular stores after application of thapsigargin. Taken together, this most likely corresponds with a general inhibitory function of CIB1 on InsP3 receptors in vivo.



Where applicable, experiments conform with Society ethical requirements.

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