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Ionic selectivity and thermal adaptations within the voltage-gated sodium channel family of alkaliphilic Bacillus.

DeCaen PG, Takahashi Y, Krulwich TA, Ito M, Clapham DE - Elife (2014)

Bottom Line: Increasing pH and temperature shifts their activation threshold towards their native resting membrane potential.Furthermore, we find drugs that target Bacillus Nav channels also block the growth of the bacteria.This work identifies some of the adaptations to achieve ion discrimination and gating in Bacillus Nav channels.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Howard Hughes Medical Institute, Boston Children's Hospital, Boston, United States.

ABSTRACT
Entry and extrusion of cations are essential processes in living cells. In alkaliphilic prokaryotes, high external pH activates voltage-gated sodium channels (Nav), which allows Na(+) to enter and be used as substrate for cation/proton antiporters responsible for cytoplasmic pH homeostasis. Here, we describe a new member of the prokaryotic voltage-gated Na(+) channel family (NsvBa; Non-selective voltage-gated, Bacillus alcalophilus) that is nonselective among Na(+), Ca(2+) and K(+) ions. Mutations in NsvBa can convert the nonselective filter into one that discriminates for Na(+) or divalent cations. Gain-of-function experiments demonstrate the portability of ion selectivity with filter mutations to other Bacillus Nav channels. Increasing pH and temperature shifts their activation threshold towards their native resting membrane potential. Furthermore, we find drugs that target Bacillus Nav channels also block the growth of the bacteria. This work identifies some of the adaptations to achieve ion discrimination and gating in Bacillus Nav channels.

No MeSH data available.


The effect of Nav antagonist on INa, bacteria motility and growth by drugs: lidocaine, tamoxifen and nifedipine.DOI:http://dx.doi.org/10.7554/eLife.04387.017
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fig7s1: The effect of Nav antagonist on INa, bacteria motility and growth by drugs: lidocaine, tamoxifen and nifedipine.DOI:http://dx.doi.org/10.7554/eLife.04387.017

Mentions: The stimuli that depolarize these bacteria from −180 mV to the more depolarized range where voltage-gated channels activate (−40 to −100 mV, depending on pH and temperature), are not known. We speculate that Ψrest declines as bacterial pumps are starved for internal H+ or Na+ to supply the hyperpolarizing extrusion pumps. Internal [Na+] would be rapidly recharged by activation of the voltage-gated monovalent cation channels. Since cation entry into alkaliphilic bacteria is at least partially dependent on Nav channels, we hypothesized that Nav channel antagonists would attenuate bacterial growth. Under voltage clamp, we observed that sodium current from Bacillus channels NavBp, NaChBac and NsvBa are blocked by known Nav channel antagonists, the local anesthetic lidocaine, the anti-hypertensive nifedipine, and the anti-estrogen tamoxifen, with similar potencies (Figure 7A,B and Figure 7—figure supplement 1). When these drugs were introduced into the culture media, growth of Bacillus species alcalophilus and pseudofirmus were severely impaired as measured by spectroscopic absorbance (Figure 7C,D). The measured half-inhibitory concentrations (IC50) of sodium current and bacterial growth were within a half-log unit (Figure 7—figure supplement 1), suggesting that growth inhibition was not an off-target effect. We also examined the effect of these drugs on two matrices of Bacillus motility: tumbling frequency and swim speed. tamoxifen, nifedipine and lidocaine increased tumbling frequency and decreased swim speed of B. pseudofirmus (Figure 7E, Figure 7—figure supplement 1). However, B. alcalophilus swim speed was not delayed by the three drugs and only tamoxifen and nifedipine increased tumbling frequency (EC50 = 70 μM and 375 μM, respectively).10.7554/eLife.04387.016Figure 7.Antagonism of Nav channels blocks the growth and motility of alkaliphilic Bacillus.


Ionic selectivity and thermal adaptations within the voltage-gated sodium channel family of alkaliphilic Bacillus.

DeCaen PG, Takahashi Y, Krulwich TA, Ito M, Clapham DE - Elife (2014)

The effect of Nav antagonist on INa, bacteria motility and growth by drugs: lidocaine, tamoxifen and nifedipine.DOI:http://dx.doi.org/10.7554/eLife.04387.017
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225499&req=5

fig7s1: The effect of Nav antagonist on INa, bacteria motility and growth by drugs: lidocaine, tamoxifen and nifedipine.DOI:http://dx.doi.org/10.7554/eLife.04387.017
Mentions: The stimuli that depolarize these bacteria from −180 mV to the more depolarized range where voltage-gated channels activate (−40 to −100 mV, depending on pH and temperature), are not known. We speculate that Ψrest declines as bacterial pumps are starved for internal H+ or Na+ to supply the hyperpolarizing extrusion pumps. Internal [Na+] would be rapidly recharged by activation of the voltage-gated monovalent cation channels. Since cation entry into alkaliphilic bacteria is at least partially dependent on Nav channels, we hypothesized that Nav channel antagonists would attenuate bacterial growth. Under voltage clamp, we observed that sodium current from Bacillus channels NavBp, NaChBac and NsvBa are blocked by known Nav channel antagonists, the local anesthetic lidocaine, the anti-hypertensive nifedipine, and the anti-estrogen tamoxifen, with similar potencies (Figure 7A,B and Figure 7—figure supplement 1). When these drugs were introduced into the culture media, growth of Bacillus species alcalophilus and pseudofirmus were severely impaired as measured by spectroscopic absorbance (Figure 7C,D). The measured half-inhibitory concentrations (IC50) of sodium current and bacterial growth were within a half-log unit (Figure 7—figure supplement 1), suggesting that growth inhibition was not an off-target effect. We also examined the effect of these drugs on two matrices of Bacillus motility: tumbling frequency and swim speed. tamoxifen, nifedipine and lidocaine increased tumbling frequency and decreased swim speed of B. pseudofirmus (Figure 7E, Figure 7—figure supplement 1). However, B. alcalophilus swim speed was not delayed by the three drugs and only tamoxifen and nifedipine increased tumbling frequency (EC50 = 70 μM and 375 μM, respectively).10.7554/eLife.04387.016Figure 7.Antagonism of Nav channels blocks the growth and motility of alkaliphilic Bacillus.

Bottom Line: Increasing pH and temperature shifts their activation threshold towards their native resting membrane potential.Furthermore, we find drugs that target Bacillus Nav channels also block the growth of the bacteria.This work identifies some of the adaptations to achieve ion discrimination and gating in Bacillus Nav channels.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Howard Hughes Medical Institute, Boston Children's Hospital, Boston, United States.

ABSTRACT
Entry and extrusion of cations are essential processes in living cells. In alkaliphilic prokaryotes, high external pH activates voltage-gated sodium channels (Nav), which allows Na(+) to enter and be used as substrate for cation/proton antiporters responsible for cytoplasmic pH homeostasis. Here, we describe a new member of the prokaryotic voltage-gated Na(+) channel family (NsvBa; Non-selective voltage-gated, Bacillus alcalophilus) that is nonselective among Na(+), Ca(2+) and K(+) ions. Mutations in NsvBa can convert the nonselective filter into one that discriminates for Na(+) or divalent cations. Gain-of-function experiments demonstrate the portability of ion selectivity with filter mutations to other Bacillus Nav channels. Increasing pH and temperature shifts their activation threshold towards their native resting membrane potential. Furthermore, we find drugs that target Bacillus Nav channels also block the growth of the bacteria. This work identifies some of the adaptations to achieve ion discrimination and gating in Bacillus Nav channels.

No MeSH data available.