An aberrant K+ channel behavior in stomatal guard cells

Sensitivity to abscisic acid of guard-cell K+ channels is suppressed by abi1-1, a mutant Arabidopsis gene encoding a putative protein phosphatase

Armstrong F., Leung J., Grabov A., Brearley J., Giraudat J., Blatt M. R., (1995)

===

Proc Natl Acad Sci U S A. 92(21): 9520-9524 – doi: 10.1073/pnas.92.21.9520 – PMID: 7568166 – PMCID: PMC40833 –

https://pubmed.ncbi.nlm.nih.gov/7568166/

Abstract

Abscisic acid (ABA) modulates the activities of three major classes of ion channels–inward- and outward-rectifying K+ channels (IK,in and IK,out, respectively) and anion channels–at the guard-cell plasma membrane to achieve a net efflux of osmotica and stomatal closure. Disruption of ABA sensitivity in wilty abi1-1 mutants of Arabidopsis and evidence that this gene encodes a protein phosphatase suggest that protein (de)-phosphorylation contributes to guard-cell transport control by ABA. To pinpoint the role of ABI1, the abi1-1 dominant mutant allele was stably transformed into Nicotiana benthamiana and its influence on IK,in, IK,out, and the anion channels was monitored in guard cells under voltage clamp. Compared with guard cells from wild-type and vector-transformed control plants, expression of the abi1-1 gene was associated with 2- to 6-fold reductions in IK,out and an insensitivity of both IK,in and IK,out to 20 microM ABA. In contrast, no differences between control and abi1-1 transgenic plants were observed in the anion current or its response to ABA. Parallel measurements of intracellular pH (pHi) using the fluorescent dye 2′,7′-bis(2-carboxyethyl)-5-(and -6)-carboxyfluorescein (BCECF) in every case showed a 0.15- to 0.2-pH-unit alkalinization in ABA, demonstrating that the transgene was without effect on the pHi signal that mediates in ABA-evoked K+ channel control. In guard cells from the abi1-1 transformants, normal sensitivity of both K+ channels to and stomatal closure in ABA was recovered in the presence of 100 microM H7 and 0.5 microM staurosporine, both broad-range protein kinase antagonists. These results demonstrate an aberrant K+ channel behavior–including channel insensitivity to ABA-dependent alkalinization of pHi–as a major consequence of abi1-1 action and implicate AB11 as part of a phosphatase/kinase pathway that modulates the sensitivity of guard-cell K+ channels to ABA-evoked signal cascades.

An aberrant K+ channel behavior in stomata

 

 

Sensitivity to abscisic acid of guard cell K⁺ channels is suppressed by abi1-1, a mutant Arabidopsis gene encoding a putative protein phosphatase.

by Armstrong F.,Leung J., Grabov A., Brearley J., Giraudat J., Blatt M.R.(1995)

Blatt M.R.

in Proc. Natl Acad. Sci. USA, 92, 9520–9524. – DOI: 10.1073/pnas.92.21.9520

CrossRef | PubMed | CAS | ADS

http://www.pnas.org/content/92/21/9520

http://www.ncbi.nlm.nih.gov/pubmed/7568166

http://chemport.cas.org/cgi-bin/sdcgi?APP=ftslink&action=reflink&origin=wiley&version=1%2E0&coi=1%3aCAS%3a528%3aDyaK2MXoslCnurk%253D&md5=5ba6995c093f8524104e33099f953f76

http://adsabs.harvard.edu/abs/1995PNAS…92.9520A

Abstract

Abscisic acid (ABA) modulates the activities of three major classes of ion channels–inward- and outward-rectifying K+ channels (IK,in and IK,out, respectively) and anion channels–at the guard-cell plasma membrane to achieve a net efflux of osmotica and stomatal closure.

Disruption of ABA sensitivity in wilty abi1-1 mutants of Arabidopsis and evidence that this gene encodes a protein phosphatase suggest that protein (de)-phosphorylation contributes to guard-cell transport control by ABA.

To pinpoint the role of ABI1, the abi1-1 dominant mutant allele was stably transformed into Nicotiana benthamiana and its influence on IK,in, IK,out, and the anion channels was monitored in guard cells under voltage clamp. Compared with guard cells from wild-type and vector-transformed control plants, expression of the abi1-1 gene was associated with 2- to 6-fold reductions in IK,out and an insensitivity of both IK,in and IK,out to 20 microM ABA.

In contrast, no differences between control and abi1-1 transgenic plants were observed in the anion current or its response to ABA. Parallel measurements of intracellular pH (pHi) using the fluorescent dye 2′,7′-bis(2-carboxyethyl)-5-(and -6)-carboxyfluorescein (BCECF) in every case showed a 0.15- to 0.2-pH-unit alkalinization in ABA, demonstrating that the transgene was without effect on the pHi signal that mediates in ABA-evoked K+ channel control.

In guard cells from the abi1-1 transformants, normal sensitivity of both K+ channels to and stomatal closure in ABA was recovered in the presence of 100 microM H7 and 0.5 microM staurosporine, both broad-range protein kinase antagonists.

These results demonstrate an aberrant K+ channel behavior–including channel insensitivity to ABA-dependent alkalinization of pHi–as a major consequence of abi1-1 action and implicate AB11 as part of a phosphatase/kinase pathway that modulates the sensitivity of guard-cell K+ channels to ABA-evoked signal cascades.

CO2, K+ and anion channels and stomata

 

 

The effect of elevated CO₂ concentrations on K⁺ and anion channels of Vicia faba L. guard cells.

by Brearley J., Venis M. A., Blatt M. R . (1997)

• 10.1007/s004250050176

in Planta 203:145–154

CrossRef

http://link.springer.com/article/10.1007%2Fs004250050176

Abstract.

The effects of elevated CO₂ concentrations on stomatal movement, anion- and K⁺-channel activities were examined in guard cells from epidermal strips of Vicia faba. Membrane voltage was measured using intracellular, double-barrelled microelectrodes and ion-channel currents were recorded under voltage clamp during exposure to media equilibrated with ambient (350 μl · l⁻¹), 1000 μl · l⁻¹ and 10 000 μl · l⁻¹ CO₂ in 20% O₂ and 80% N₂.

The addition of 1000 μl · l⁻¹ CO₂ to the bathing solution caused stomata to close with a halftime of approx. 40 min, and with 10 000 μl · l⁻¹ CO₂closure occurred with a similar time course. Under voltage clamp, exposure to 1000 μl · l⁻¹ and 10 000 μl · l⁻¹ CO₂ resulted in a rapid increase (mean, 1.5 ± 0.2-fold, n = 8; range 1.3- to 2.5-fold) in the magnitude of current carried by outward-rectifying K⁺ channels (I_K,out).

The effect of CO₂ on I_K,outwas essentially complete within 30 s and was independent of clamp voltage, but was associated with 25–40% (mean, 30 ± 4%) decrease in the halftime for current activation. Exposure to CO₂ also resulted in a four-fold increase in background current near the free-running membrane voltage, recorded as the instantaneous current at the start of depolarising and hyperpolarising voltage steps, and a decrease in the magnitude of current carried by inward-rectifying K⁺ channels (I_K,in).

The effect of CO₂ on I_K,in was generally slower than on I_K,out; it was allied with a transient acceleration of its activation kinetics during the first 60–120 s of treatment; and it was associated with a negative shift in the voltage-sensitivity of gating over a period of 3–5 min.

Measurements carried out to isolate the background currents attributable to anion channels (I_Cl), using tetraethylammonium chloride and CsCl, showed that CO₂ also stimulated I_Cl and dramatically altered its relaxation kinetics.

Within the timeframe of CO₂ action at the membrane, no significant effect was observed on cytosolic pH, measured using the fluorescent dye 2′,7′-bis-(2-carboxyethyl)-5,6-carboxyflourescein (BCECF) and ratio fluorescence microphotometry.

These results are broadly consistent with the pattern of guard-cell response to abscisic acid, and indicate that guard cells control both anion and K⁺ channels to achieve net solute loss in CO₂.

By contrast with the effects of abscisic acid, however, the data indicate that CO₂ action is not mediated through changes in cytosolic pH and thereby implicate new and, as yet, unidentified pathway(s) for channel regulation in the guard cells.

The role of abi1-1, a mutant Arabidopsis gene, in stomata

 

Sensitivity to abscisic acid of guard-cell K+ channels is suppressed by abi1-1, a mutant Arabidopsis gene encoding a putative protein phosphatase

by Armstrong F., Leung J., Grabov A., Brearley J., Giraudat J., Blatt M.R.  (1995)

in Proceedings of the National Academy of Sciences, USA 1995;92:9520-9524. –

Abstract/FREE Full Text

Abstract

Abscisic acid (ABA) modulates the activities of three major classes of ion channels–inward- and outward-rectifying K+ channels (IK,in and IK,out, respectively) and anion channels–at the guard-cell plasma membrane to achieve a net efflux of osmotica and stomatal closure. Disruption of ABA sensitivity in wilty abi1-1 mutants of Arabidopsis and evidence that this gene encodes a protein phosphatase suggest that protein (de)-phosphorylation contributes to guard-cell transport control by ABA.

To pinpoint the role of ABI1, the abi1-1 dominant mutant allele was stably transformed into Nicotiana benthamiana and its influence on IK,in, IK,out, and the anion channels was monitored in guard cells under voltage clamp. Compared with guard cells from wild-type and vector-transformed control plants, expression of the abi1-1 gene was associated with 2- to 6-fold reductions in IK,out and an insensitivity of both IK,in and IK,out to 20 microM ABA.

In contrast, no differences between control and abi1-1 transgenic plants were observed in the anion current or its response to ABA. Parallel measurements of intracellular pH (pHi) using the fluorescent dye 2′,7′-bis(2-carboxyethyl)-5-(and -6)-carboxyfluorescein (BCECF) in every case showed a 0.15- to 0.2-pH-unit alkalinization in ABA, demonstrating that the transgene was without effect on the pHi signal that mediates in ABA-evoked K+ channel control. In guard cells from the abi1-1 transformants, normal sensitivity of both K+ channels to and stomatal closure in ABA was recovered in the presence of 100 microM H7 and 0.5 microM staurosporine, both broad-range protein kinase antagonists.

These results demonstrate an aberrant K+ channel behavior–including channel insensitivity to ABA-dependent alkalinization of pHi–as a major consequence of abi1-1 action and implicate AB11 as part of a phosphatase/kinase pathway that modulates the sensitivity of guard-cell K+ channels to ABA-evoked signal cascades.

See the text: PNAS

Read the full article: PNAS