TRPM8 channels in supramolecular complexes with Inorganic Polyphosphate and Polyhydroxybutyrate The main objective of this proposal is to understand the nature, function and mechanism of TRP channels in supramolecular complexes. TRP channels are calcium-permeable cation channels that play significant roles in a great variety of physiological and pathological processes. In this project we will focus on studying one of the members of melastatin subfamily TRP channels, TRPM8. The cold and menthol receptor, TRPM8, expressed in sensory neurons and is involved in the pathway of thermosensation. The activity of TRPM8 channels is regulated by a variety of factors, including cool temperatures, ligand binding, voltage, pH, etc. reflecting diverse stimuli and mechanisms of these regulators. We propose that such a diversity of stimuli and mechanisms of these channels can be explained by the complexity of the TRPM8 protein itself. In our preliminary results we reveal that TRPM8 is associated with inorganic polyphosphate (polyP) and polyhydroxybutyrate (PHB), and furthermore this complex modifies functional properties of the channel. PolyP and PHB are ubiquitous homopolymers that possess important molecular properties in the formation of the simplest model of ion channel. The polyP/PHB complexes have been found in association with a number of membrane proteins, including ion channels and porins. Moreover, these complexes were shown to be important for normal functioning of their hosts. Here we postulate that TRPM8 exists in a supramolecular complex with polyP and PHB and that such an ensemble might be a common feature of other TRP channels. To test this hypothesis we will apply biochemical and biophysical analysis, electrophysiology, enzymology, molecular genetics, molecular spectroscopy and fluorescence spectroscopy methods.
The specific aims of this proposal are: (I) assess the association of polyhydroxybutyrate with TRPM8 and role of the polyester in temperature sensation of TRPM8;(II) examine the physiological role of inorganic polyphosphate in TRPM8 channel function. The proposed study will open new directions and insights on TRP channels as supramolecular complexes and it will be of an immense importance for understanding of the molecular mechanisms for such interactions and their regulation in the function of these channels during physiological and pathological processes.
Transient Receptor Potential (TRP) channels play significant roles in a great variety of physiological and pathological processes. TRP channels are involved in development of various diseases due to the multifunctional cellular responses carried by these proteins. The overall goal of this project is to unravel the molecular complexity of these channels, which will open new insights into the regulatory mechanisms of the channel activity of TRPs and will be helpful for drug design.
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|Uchida, Kunitoshi; Demirkhanyan, Lusine; Asuthkar, Swapna et al. (2016) Stimulation-dependent gating of TRPM3 channel in planar lipid bilayers. FASEB J 30:1306-16|
|Asuthkar, Swapna; Demirkhanyan, Lusine; Sun, Xiaohui et al. (2015) The TRPM8 protein is a testosterone receptor: II. Functional evidence for an ionotropic effect of testosterone on TRPM8. J Biol Chem 290:2670-88|
|BÃ¡rcena-Uribarri, IvÃ¡n; Benz, Roland; Winterhalter, Mathias et al. (2015) Pore forming activity of the potent RTX-toxin produced by pediatric pathogen Kingella kingae: Characterization and comparison to other RTX-family members. Biochim Biophys Acta 1848:1536-44|
|Asuthkar, Swapna; Elustondo, Pia A; Demirkhanyan, Lusine et al. (2015) The TRPM8 protein is a testosterone receptor: I. Biochemical evidence for direct TRPM8-testosterone interactions. J Biol Chem 290:2659-69|
|Asuthkar, Swapna; Velpula, Kiran Kumar; Elustondo, Pia A et al. (2015) TRPM8 channel as a novel molecular target in androgen-regulated prostate cancer cells. Oncotarget 6:17221-36|
|Sun, Xiaohui; Zakharian, Eleonora (2015) Regulation of the temperature-dependent activation of transient receptor potential vanilloid 1 (TRPV1) by phospholipids in planar lipid bilayers. J Biol Chem 290:4741-7|
|Chang, Dennis W; Nudell, Yoav A; Lau, Jenny et al. (2014) RTX toxin plays a key role in Kingella kingae virulence in an infant rat model. Infect Immun 82:2318-28|
|Zakharian, Eleonora (2013) Recording of ion channel activity in planar lipid bilayer experiments. Methods Mol Biol 998:109-18|
|Smithen, Matthew; Elustondo, Pia A; Winkfein, Robert et al. (2013) Role of polyhydroxybutyrate in mitochondrial calcium uptake. Cell Calcium 54:86-94|
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