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Divalent cations potentiate TRPV1 channel by lowering the heat activation threshold.

ABSTRACT: Transient receptor potential vanilloid type 1 (TRPV1) channel responds to a wide spectrum of physical and chemical stimuli. In doing so, it serves as a polymodal cellular sensor for temperature change and pain. Many chemicals are known to strongly potentiate TRPV1 activation, though how this is achieved remains unclear. In this study we investigated the molecular mechanism underlying the gating effects of divalent cations Mg(2+) and Ba(2+). Using a combination of fluorescence imaging and patch-clamp analysis, we found that these cations potentiate TRPV1 gating by most likely promoting the heat activation process. Mg(2+) substantially lowers the activation threshold temperature; as a result, a significant fraction of channels are heat-activated at room temperature. Although Mg(2+) also potentiates capsaicin- and voltage-dependent activation, these processes were found either to be not required (in the case of capsaicin) or insufficient (in the case of voltage) to mediate the activating effect. In support of a selective effect on heat activation, Mg(2+) and Ba(2+) cause a Ca(2+)-independent desensitization that specifically prevents heat-induced channel activation but does not prevent capsaicin-induced activation. These results can be satisfactorily explained within an allosteric gating framework in which divalent cations strongly promote the heat-dependent conformational change or its coupling to channel activation, which is further coupled to the voltage- and capsaicin-dependent processes.

SUBMITTER: Cao X

PROVIDER: S-EPMC3874569 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature

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Divalent cations potentiate TRPV1 channel by lowering the heat activation threshold.

Cao Xu X Ma Linlin L Yang Fan F Wang Kewei K Zheng Jie J

The Journal of general physiology 20131216 1

Transient receptor potential vanilloid type 1 (TRPV1) channel responds to a wide spectrum of physical and chemical stimuli. In doing so, it serves as a polymodal cellular sensor for temperature change and pain. Many chemicals are known to strongly potentiate TRPV1 activation, though how this is achieved remains unclear. In this study we investigated the molecular mechanism underlying the gating effects of divalent cations Mg(2+) and Ba(2+). Using a combination of fluorescence imaging and patch-c ...[more]

PMID: 24344247

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Project description:Among the best-studied interactions between soil phagocytic predators and a human-pathogenic fungus is that of Acanthamoeba castellanii and Cryptococcus neoformans The experimental conditions used in amoeba-fungus confrontation assays can have major effects on whether the fungus or the protozoan is ascendant in the interaction. In the presence of Mg2+ and Ca2+ in phosphate-buffered saline (PBS), C. neoformans was consistently killed when incubated with A. castellaniiA. castellanii survived better in the presence of Mg2+ and Ca2+, even when incubated with C. neoformans In the absence of Mg2+ and Ca2+, C. neoformans survived when incubated with A. castellanii, and the percentage of dead amoebae was higher than when incubated without yeast cells. These results show that the presence of Mg2+ and Ca2+ can make a decisive contribution toward tilting the outcome of the interaction in favor of the amoeba. Of the two metals, Mg2+ had a stronger effect than Ca2+ The cations enhanced A. castellanii activity against C. neoformans via enhanced phagocytosis, which is the major mechanism by which amoebae kill fungal cells. We found no evidence that amoebae use extracellular killing mechanisms in their interactions with C. neoformans In summary, the presence of Mg2+ and Ca2+ enhanced the cell adhesion on the surfaces and the motility of the amoeba, thus increasing the chance for contact with C. neoformans and the frequency of phagocytosis. Our findings imply that the divalent cation concentration in soils could be an important variable for whether amoebae can control C. neoformans in the environment.IMPORTANCE The grazing of soil organisms by phagocytic predators such as amoebae is thought to select for traits that enable some of them to acquire the capacity for virulence in animals. Consequently, knowledge about the interactions between amoebae and soil microbes, such as pathogenic fungi, is important for understanding how virulence can emerge. We show that the interaction between an amoeba and the pathogenic fungus C. neoformans is influenced by the presence in the assay of magnesium and calcium, which potentiate amoebae. The results may also have practical applications, since enriching soils with divalent cations may reduce C. neoformans numbers in contaminated soils.

Project description:As a key cellular sensor, the TRPV1 cation channel undergoes a gating transition from a closed state to an open state in response to various physical and chemical stimuli including noxious heat. Despite years of study, the heat activation mechanism of TRPV1 gating remains enigmatic at the molecular level. Toward elucidating the structural and energetic basis of TRPV1 gating, we have performed molecular dynamics (MD) simulations (with cumulative simulation time of 3 μs), starting from the high-resolution closed and open structures of TRPV1 solved by cryo-electron microscopy. In the closed-state simulations at 30°C, we observed a stably closed channel constricted at the lower gate (near residue I679), while the upper gate (near residues G643 and M644) is dynamic and undergoes flickery opening/closing. In the open-state simulations at 60°C, we found higher conformational variation consistent with a large entropy increase required for the heat activation, and both the lower and upper gates are dynamic with transient opening/closing. Through ensemble-based structural analyses of the closed state versus the open state, we revealed pronounced closed-to-open conformational changes involving the membrane proximal domain (MPD) linker, the outer pore, and the TRP helix, which are accompanied by breaking/forming of a network of closed/open-state specific hydrogen bonds. By comparing the closed-state simulations at 30°C and 60°C, we observed heat-activated conformational changes in the MPD linker, the outer pore, and the TRP helix that resemble the closed-to-open conformational changes, along with partial formation of the open-state specific hydrogen bonds. Some of the residues involved in the above key hydrogen bonds were validated by previous mutational studies. Taken together, our MD simulations have offered rich structural and dynamic details beyond the static structures of TRPV1, and promising targets for future mutagenesis and functional studies of the TRPV1 channel. Proteins 2016; 84:1938-1949. © 2016 Wiley Periodicals, Inc.

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Divalent cations potentiate TRPV1 channel by lowering the heat activation threshold.

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Divalent cations potentiate TRPV1 channel by lowering the heat activation threshold.

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