Kees Venema - Academia.edu (original) (raw)

Papers by Kees Venema

Research paper thumbnail of Estrés Abiótico y Homeostasis Iónica

Research paper thumbnail of Gibberellic Acid and Indole Acetic Acid Improves Salt Tolerance in Transgenic Tomato Plants Overexpressing LeNHX4 Antiporter

Research paper thumbnail of Transporters and Pumps in Plant Signaling

Signaling and Communication in Plants, 2011

The use of general descriptive names, registered names, trademarks, etc. in this publication does... more The use of general descriptive names, registered names, trademarks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use.

Research paper thumbnail of Overexpression of LeNHX2 and SlSOS2 increases salt tolerance and fruit production in double transgenic tomato plants

Plant Physiology and Biochemistry, 2019

Research paper thumbnail of Unravelling salt tolerance mechanisms in plants: From lab to field

Plant Physiology and Biochemistry, 2022

Research paper thumbnail of Na/K-Atpase and Related Transport Atpases

Research paper thumbnail of Loss of function of the chloroplast membrane K+/H+ antiporters AtKEA1 and AtKEA2 alters the ROS and NO metabolism but promotes drought stress resilience

Plant Physiology and Biochemistry, 2021

Research paper thumbnail of Overexpression of LeNHX4 improved yield, fruit quality and salt tolerance in tomato plants (Solanum lycopersicum L.)

Molecular Biology Reports, 2020

The function of the tomato K+, Na+/H+ antiporter LeNHX4 has been analyzed using 35S-driven gene c... more The function of the tomato K+, Na+/H+ antiporter LeNHX4 has been analyzed using 35S-driven gene construct for overexpressing a histagged LeNHX4 protein in Solanum lycopersicum L. Compared to wild-type plants, the expression of LeNHX4 was enhanced in most of plants transformed with a gene construct for LeNHX4 overexpression although some plants showed a decreased LeNHX4 expression. Overexpression of LeNHX4 was associated to an increased fruit size while silencing of this gene was related to a decreased fruit size. We have investigated the effect of LeNHX4 overexpression on fruit production and quality and we have also evaluated salt tolerance in two different overexpression lines by measuring proline, protein and glucose concentrations in tomato leaves grown either under control (0 mM NaCl) or saline (125 mM NaCl) conditions. Plants overexpressing LeNHX4 showed a higher amount of fruits than WT plants and accumulated higher contents of sugars and cations (Na+ and K+). The application of 125 mM NaCl, affected negatively fruit production and quality of WT plants. However the transgenic lines overexpressing LeNXH4 increased fruit quality and yield. In relation to salt tolerance, overexpression lines showed higher levels of leaf proline, glucose and proteins under NaCl treatment. The overexpression of LeNHX4 in tomato plants, improved salinity tolerance and increased fruit yield and quality under both normal and salinity stress conditions.

Research paper thumbnail of Envelope K+/H+ Antiporters AtKEA1 and AtKEA2 Function in Plastid Development

Plant physiology, Sep 1, 2016

It is well established that thylakoid membranes of chloroplasts convert light energy into chemica... more It is well established that thylakoid membranes of chloroplasts convert light energy into chemical energy, yet the development of chloroplast and thylakoid membranes is poorly understood. Loss of function of the two envelope K(+)/H(+) antiporters AtKEA1 and AtKEA2 was shown previously to have negative effects on the efficiency of photosynthesis and plant growth; however, the molecular basis remained unclear. Here, we tested whether the previously described phenotypes of double mutant kea1kea2 plants are due in part to defects during early chloroplast development in Arabidopsis (Arabidopsis thaliana). We show that impaired growth and pigmentation is particularly evident in young expanding leaves of kea1kea2 mutants. In proliferating leaf zones, chloroplasts contain much lower amounts of photosynthetic complexes and chlorophyll. Strikingly, AtKEA1 and AtKEA2 proteins accumulate to high amounts in small and dividing plastids, where they are specifically localized to the two caps of the...

Research paper thumbnail of P-Type H + - and Ca 2+ -ATPases in Plant Cells

Annals of the New York Academy of Sciences, 1997

Plasma membrane H+-ATPase and Na+/K+-ATPase belong to the same subfamily of ion pumps that has be... more Plasma membrane H+-ATPase and Na+/K+-ATPase belong to the same subfamily of ion pumps that has been termed Pz-ATPases. I A phylogenetic tree showing the evolutionary relation between members of the P-type ATPase superfamily is presented in FIGURE 1. Plasma membrane proton pumps appear to constitute a monophyletic group of P-type ATPases and are present in plants, fungi, algae, protozoa, and archaea. This wide distribution among species suggests that P-type H+-ATPases appeared very early in evolution. On the contrary, Na+/K+-ATPases probably appeared late in evolution because they are restricted to animals, who evolved in the sea and were dependent on systems to extrude sodium. However, in plants and animals respectively, H+-ATPase and Na+lK+-ATPase serve analogous functions. Thus, in plants (and

Research paper thumbnail of Changing the ion binding specificity of the plant plasma membrane proton ATPase

Research paper thumbnail of Improved yield, fruit quality, and salt resistance in tomato co-overexpressing LeNHX2 and SlSOS2 genes

Physiology and Molecular Biology of Plants

The K+, Na+/H+ antiporter LeNHX2 and the regulatory kinase SlSOS2 are important determinants of s... more The K+, Na+/H+ antiporter LeNHX2 and the regulatory kinase SlSOS2 are important determinants of salt tolerance in tomato plants and their fruit production ability. In this work, we have analyzed the effects of LeNHX2 and SlSOS2 co-overexpression on fruit production, quality in tomato plants (Solanum lycopersicum L. cv. MicroTom), and analyzed physiological parameters related to salt tolerance. Plants overexpressing LeNHX2, SlSOS2 or both were grown in greenhouse. They were treated with 125 mM NaCl or left untreated and their salt tolerance was analyzed in terms of plant biomass and fruit yield. Under NaCl cultivation conditions, transgenic tomato plants overexpressing either SlSOS2 or LeNHX2 or both grew better and showed a higher biomass compared to their wild-type plants. Proline, glucose and protein content in leaves as well as pH and total soluble solid (TSS) in fruits were analyzed. Our results indicate that salinity tolerance of transgenic lines is associated with an increased proline, glucose and protein content in leaves of plants grown either with or without NaCl. Salt treatment significantly reduced yield, pH and TSS in fruits of WT plants but increased yield, pH and TSS in fruits of transgenic plants, especially those overexpressing both LeNHX2 and SlSOS2. All these results indicate that the co-overexpression of LeNHX2 and SlSOS2 improve yield and fruit quality of tomato grown under saline conditions.

Research paper thumbnail of Deletion of the N‐terminal domain of the yeast vacuolar (Na + ,K + )/H + antiporter Vnx1p improves salt tolerance in yeast and transgenic Arabidopsis

Yeast

Cation/proton antiporters play a major role in the control of cytosolic ion concentrations in pro... more Cation/proton antiporters play a major role in the control of cytosolic ion concentrations in prokaryotes and eukaryotes organisms. In yeast, we previously demonstrated that Vnx1p is a vacuolar monovalent cation/H + exchanger showing Na + /H + and K + / H + antiporter activity. We have also shown that disruption of VNX1 results in an almost complete abolishment of vacuolar Na + /H + exchange, but yeast cells overexpressing the complete protein do not show improved salinity tolerance. In this study, we have identified an autoinhibitory N-terminal domain and have engineered a constitutively activated version of Vnx1p, by removing this domain. Contrary to the wild type protein, the activated protein has a pronounced effect on yeast salt tolerance and vacuolar pH. Expression of this truncated VNX1 gene also improves Arabidopsis salt tolerance and increases Na + and K + accumulation of salt grown plants thus suggesting a biotechnological potential of activated Vnx1p to improve salt tolerance of crop plants.

Research paper thumbnail of Plastidial transporters KEA1 and KEA2 at the inner envelope membrane adjust stromal pH in the dark

New Phytologist

Photosynthesis and carbon fixation depend critically on the regulation of pH in chloroplast compa... more Photosynthesis and carbon fixation depend critically on the regulation of pH in chloroplast compartments in day light and at night. While it is established that an alkaline stroma is required for carbon fixation, it is not known how alkaline stromal pH is formed, maintained or regulated. We tested whether two envelope transporters, AtKEA1 and AtKEA2, directly affected stromal pH in isolated Arabidopsis chloroplasts using the fluorescent probe BCECF. External K+ induced alkalinization of the stroma was observed in chloroplasts from WT plants but not from kea1kea2 mutants, suggesting KEA1 and KEA2 mediate K+ uptake/H+ loss to modulate stromal pH. While light stimulated alkalinization of the stroma was independent of KEA1 and KEA2, the rate of decay to neutral pH in the dark is delayed in kea1kea2 mutants. However, the dark-induced loss of a pH gradient across the thylakoid membrane was similar in WT and mutant chloroplasts. This indicates that proton influx from the cytosol mediated by envelope K+ /H+ antiporters contributes to adjustment of stromal pH upon light to dark transitions.

Research paper thumbnail of Etude quantitative du transport facilite de k#+ au niveau de la membrane plasmique vegetale a l'aide de vesicules isolees et de sondes fluorescentes

Research paper thumbnail of Proton Gradients and Proton-Dependent Transport Processes in the Chloroplast

Frontiers in plant science, 2016

Proton gradients are fundamental to chloroplast function. Across thylakoid membranes, the light i... more Proton gradients are fundamental to chloroplast function. Across thylakoid membranes, the light induced -proton gradient is essential for ATP synthesis. As a result of proton pumping into the thylakoid lumen, an alkaline stromal pH develops, which is required for full activation of pH-dependent Calvin Benson cycle enzymes. This implies that a pH gradient between the cytosol (pH 7) and the stroma (pH 8) is established upon illumination. To maintain this pH gradient chloroplasts actively extrude protons. More than 30 years ago it was already established that these proton fluxes are electrically counterbalanced by Mg(2+), K(+), or Cl(-) fluxes, but only recently the first transport systems that regulate the pH gradient were identified. Notably several (Na(+),K(+))/H(+) antiporter systems where identified, that play a role in pH gradient regulation, ion homeostasis, osmoregulation, or coupling of secondary active transport. The established pH gradients are important to drive uptake of e...

Research paper thumbnail of Protein architecture and core residues in unwound α-helices provide insights to the transport function of plant AtCHX17

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2016

Using Arabidopsis thaliana AtCHX17 as an example, we combine structural modeling and mutagenesis ... more Using Arabidopsis thaliana AtCHX17 as an example, we combine structural modeling and mutagenesis to provide insights on its protein architecture and transport function which is poorly characterized. This approach is based on the observation that protein structures are significantly more conserved in evolution than linear sequences, and mechanistic similarities among diverse transporters are emerging. Two homology models of AtCHX17 were obtained that show a protein fold similar to known structures of bacterial Na(+)/H(+) antiporters, EcNhaA and TtNapA. The distinct secondary and tertiary structure models highlighted residues at positions potentially important for CHX17 activity. Mutagenesis showed that asparagine-N200 and aspartate-D201 inside transmembrane5 (TM5), and lysine-K355 inside TM10 are critical for AtCHX17 activity. We reveal previously unrecognized threonine-T170 and lysine-K383 as key residues at unwound regions in the middle of TM4 and TM11 α-helices, respectively. Mutation of glutamate-E111 located near the membrane surface inhibited AtCHX17 activity, suggesting a role in pH sensing. The long carboxylic tail of unknown purpose has an alternating β-sheet and α-helix secondary structure that is conserved in prokaryote universal stress proteins. These results support the overall architecture of AtCHX17 and identify D201, N200 and novel residues T170 and K383 at the functional core which likely participates in ion recognition, coordination and/or translocation, similar to characterized cation/H(+) exchangers. The core of AtCHX17 models according to EcNhaA and TtNapA templates faces inward and outward, respectively, which may reflect two conformational states of the alternating access transport mode for proteins belonging to the plant CHX family.

Research paper thumbnail of Plant NHX cation/proton antiporters

Plant Signaling Behavior, 2009

Addendum to: This manuscript has been published online, prior to printing. Once the issue is comp... more Addendum to: This manuscript has been published online, prior to printing. Once the issue is complete and page numbers have been assigned, the citation will change accordingly.

Research paper thumbnail of Heterologously expressed protein phosphatase calcineurin downregulates plant plasma membrane H +-ATPase activity at the post-translational level

Febs Letters, Oct 8, 2004

To investigate the effects of calcineurin expression on cellular ion homeostasis in plants, we ha... more To investigate the effects of calcineurin expression on cellular ion homeostasis in plants, we have obtained a transgenic cell culture of tomato, expressing constitutively activated yeast calcineurin. Transgenic cells exhibited reduced growth rates and proton extrusion activity in vivo. We show that reduction of plasma membrane H þ -ATPase activity by expression of calcineurin is the basis for the observed phenotypes. Transgenic calli and cell suspensions displayed also increased salt tolerance and contained slightly higher Ca 2þ and K þ levels. This demonstrates that calcineurin can modulate ion homeostasis in plants as it does in yeast by affecting the activity of primary ion transporters.

Research paper thumbnail of Conserved and diversified gene families of monovalent cation/h(+) antiporters from algae to flowering plants

Frontiers in plant science, 2012

All organisms have evolved strategies to regulate ion and pH homeostasis in response to developme... more All organisms have evolved strategies to regulate ion and pH homeostasis in response to developmental and environmental cues. One strategy is mediated by monovalent cation-proton antiporters (CPA) that are classified in two superfamilies. Many CPA1 genes from bacteria, fungi, metazoa, and plants have been functionally characterized; though roles of plant CPA2 genes encoding K(+)-efflux antiporter (KEA) and cation/H(+) exchanger (CHX) families are largely unknown. Phylogenetic analysis showed that three clades of the CPA1 Na(+)-H(+) exchanger (NHX) family have been conserved from single-celled algae to Arabidopsis. These are (i) plasma membrane-bound SOS1/AtNHX7 that share ancestry with prokaryote NhaP, (ii) endosomal AtNHX5/6 that is part of the eukaryote Intracellular-NHE clade, and (iii) a vacuolar NHX clade (AtNHX1-4) specific to plants. Early diversification of KEA genes possibly from an ancestral cyanobacterium gene is suggested by three types seen in all plants. Intriguingly, ...

Research paper thumbnail of Estrés Abiótico y Homeostasis Iónica

Research paper thumbnail of Gibberellic Acid and Indole Acetic Acid Improves Salt Tolerance in Transgenic Tomato Plants Overexpressing LeNHX4 Antiporter

Research paper thumbnail of Transporters and Pumps in Plant Signaling

Signaling and Communication in Plants, 2011

The use of general descriptive names, registered names, trademarks, etc. in this publication does... more The use of general descriptive names, registered names, trademarks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use.

Research paper thumbnail of Overexpression of LeNHX2 and SlSOS2 increases salt tolerance and fruit production in double transgenic tomato plants

Plant Physiology and Biochemistry, 2019

Research paper thumbnail of Unravelling salt tolerance mechanisms in plants: From lab to field

Plant Physiology and Biochemistry, 2022

Research paper thumbnail of Na/K-Atpase and Related Transport Atpases

Research paper thumbnail of Loss of function of the chloroplast membrane K+/H+ antiporters AtKEA1 and AtKEA2 alters the ROS and NO metabolism but promotes drought stress resilience

Plant Physiology and Biochemistry, 2021

Research paper thumbnail of Overexpression of LeNHX4 improved yield, fruit quality and salt tolerance in tomato plants (Solanum lycopersicum L.)

Molecular Biology Reports, 2020

The function of the tomato K+, Na+/H+ antiporter LeNHX4 has been analyzed using 35S-driven gene c... more The function of the tomato K+, Na+/H+ antiporter LeNHX4 has been analyzed using 35S-driven gene construct for overexpressing a histagged LeNHX4 protein in Solanum lycopersicum L. Compared to wild-type plants, the expression of LeNHX4 was enhanced in most of plants transformed with a gene construct for LeNHX4 overexpression although some plants showed a decreased LeNHX4 expression. Overexpression of LeNHX4 was associated to an increased fruit size while silencing of this gene was related to a decreased fruit size. We have investigated the effect of LeNHX4 overexpression on fruit production and quality and we have also evaluated salt tolerance in two different overexpression lines by measuring proline, protein and glucose concentrations in tomato leaves grown either under control (0 mM NaCl) or saline (125 mM NaCl) conditions. Plants overexpressing LeNHX4 showed a higher amount of fruits than WT plants and accumulated higher contents of sugars and cations (Na+ and K+). The application of 125 mM NaCl, affected negatively fruit production and quality of WT plants. However the transgenic lines overexpressing LeNXH4 increased fruit quality and yield. In relation to salt tolerance, overexpression lines showed higher levels of leaf proline, glucose and proteins under NaCl treatment. The overexpression of LeNHX4 in tomato plants, improved salinity tolerance and increased fruit yield and quality under both normal and salinity stress conditions.

Research paper thumbnail of Envelope K+/H+ Antiporters AtKEA1 and AtKEA2 Function in Plastid Development

Plant physiology, Sep 1, 2016

It is well established that thylakoid membranes of chloroplasts convert light energy into chemica... more It is well established that thylakoid membranes of chloroplasts convert light energy into chemical energy, yet the development of chloroplast and thylakoid membranes is poorly understood. Loss of function of the two envelope K(+)/H(+) antiporters AtKEA1 and AtKEA2 was shown previously to have negative effects on the efficiency of photosynthesis and plant growth; however, the molecular basis remained unclear. Here, we tested whether the previously described phenotypes of double mutant kea1kea2 plants are due in part to defects during early chloroplast development in Arabidopsis (Arabidopsis thaliana). We show that impaired growth and pigmentation is particularly evident in young expanding leaves of kea1kea2 mutants. In proliferating leaf zones, chloroplasts contain much lower amounts of photosynthetic complexes and chlorophyll. Strikingly, AtKEA1 and AtKEA2 proteins accumulate to high amounts in small and dividing plastids, where they are specifically localized to the two caps of the...

Research paper thumbnail of P-Type H + - and Ca 2+ -ATPases in Plant Cells

Annals of the New York Academy of Sciences, 1997

Plasma membrane H+-ATPase and Na+/K+-ATPase belong to the same subfamily of ion pumps that has be... more Plasma membrane H+-ATPase and Na+/K+-ATPase belong to the same subfamily of ion pumps that has been termed Pz-ATPases. I A phylogenetic tree showing the evolutionary relation between members of the P-type ATPase superfamily is presented in FIGURE 1. Plasma membrane proton pumps appear to constitute a monophyletic group of P-type ATPases and are present in plants, fungi, algae, protozoa, and archaea. This wide distribution among species suggests that P-type H+-ATPases appeared very early in evolution. On the contrary, Na+/K+-ATPases probably appeared late in evolution because they are restricted to animals, who evolved in the sea and were dependent on systems to extrude sodium. However, in plants and animals respectively, H+-ATPase and Na+lK+-ATPase serve analogous functions. Thus, in plants (and

Research paper thumbnail of Changing the ion binding specificity of the plant plasma membrane proton ATPase

Research paper thumbnail of Improved yield, fruit quality, and salt resistance in tomato co-overexpressing LeNHX2 and SlSOS2 genes

Physiology and Molecular Biology of Plants

The K+, Na+/H+ antiporter LeNHX2 and the regulatory kinase SlSOS2 are important determinants of s... more The K+, Na+/H+ antiporter LeNHX2 and the regulatory kinase SlSOS2 are important determinants of salt tolerance in tomato plants and their fruit production ability. In this work, we have analyzed the effects of LeNHX2 and SlSOS2 co-overexpression on fruit production, quality in tomato plants (Solanum lycopersicum L. cv. MicroTom), and analyzed physiological parameters related to salt tolerance. Plants overexpressing LeNHX2, SlSOS2 or both were grown in greenhouse. They were treated with 125 mM NaCl or left untreated and their salt tolerance was analyzed in terms of plant biomass and fruit yield. Under NaCl cultivation conditions, transgenic tomato plants overexpressing either SlSOS2 or LeNHX2 or both grew better and showed a higher biomass compared to their wild-type plants. Proline, glucose and protein content in leaves as well as pH and total soluble solid (TSS) in fruits were analyzed. Our results indicate that salinity tolerance of transgenic lines is associated with an increased proline, glucose and protein content in leaves of plants grown either with or without NaCl. Salt treatment significantly reduced yield, pH and TSS in fruits of WT plants but increased yield, pH and TSS in fruits of transgenic plants, especially those overexpressing both LeNHX2 and SlSOS2. All these results indicate that the co-overexpression of LeNHX2 and SlSOS2 improve yield and fruit quality of tomato grown under saline conditions.

Research paper thumbnail of Deletion of the N‐terminal domain of the yeast vacuolar (Na + ,K + )/H + antiporter Vnx1p improves salt tolerance in yeast and transgenic Arabidopsis

Yeast

Cation/proton antiporters play a major role in the control of cytosolic ion concentrations in pro... more Cation/proton antiporters play a major role in the control of cytosolic ion concentrations in prokaryotes and eukaryotes organisms. In yeast, we previously demonstrated that Vnx1p is a vacuolar monovalent cation/H + exchanger showing Na + /H + and K + / H + antiporter activity. We have also shown that disruption of VNX1 results in an almost complete abolishment of vacuolar Na + /H + exchange, but yeast cells overexpressing the complete protein do not show improved salinity tolerance. In this study, we have identified an autoinhibitory N-terminal domain and have engineered a constitutively activated version of Vnx1p, by removing this domain. Contrary to the wild type protein, the activated protein has a pronounced effect on yeast salt tolerance and vacuolar pH. Expression of this truncated VNX1 gene also improves Arabidopsis salt tolerance and increases Na + and K + accumulation of salt grown plants thus suggesting a biotechnological potential of activated Vnx1p to improve salt tolerance of crop plants.

Research paper thumbnail of Plastidial transporters KEA1 and KEA2 at the inner envelope membrane adjust stromal pH in the dark

New Phytologist

Photosynthesis and carbon fixation depend critically on the regulation of pH in chloroplast compa... more Photosynthesis and carbon fixation depend critically on the regulation of pH in chloroplast compartments in day light and at night. While it is established that an alkaline stroma is required for carbon fixation, it is not known how alkaline stromal pH is formed, maintained or regulated. We tested whether two envelope transporters, AtKEA1 and AtKEA2, directly affected stromal pH in isolated Arabidopsis chloroplasts using the fluorescent probe BCECF. External K+ induced alkalinization of the stroma was observed in chloroplasts from WT plants but not from kea1kea2 mutants, suggesting KEA1 and KEA2 mediate K+ uptake/H+ loss to modulate stromal pH. While light stimulated alkalinization of the stroma was independent of KEA1 and KEA2, the rate of decay to neutral pH in the dark is delayed in kea1kea2 mutants. However, the dark-induced loss of a pH gradient across the thylakoid membrane was similar in WT and mutant chloroplasts. This indicates that proton influx from the cytosol mediated by envelope K+ /H+ antiporters contributes to adjustment of stromal pH upon light to dark transitions.

Research paper thumbnail of Etude quantitative du transport facilite de k#+ au niveau de la membrane plasmique vegetale a l'aide de vesicules isolees et de sondes fluorescentes

Research paper thumbnail of Proton Gradients and Proton-Dependent Transport Processes in the Chloroplast

Frontiers in plant science, 2016

Proton gradients are fundamental to chloroplast function. Across thylakoid membranes, the light i... more Proton gradients are fundamental to chloroplast function. Across thylakoid membranes, the light induced -proton gradient is essential for ATP synthesis. As a result of proton pumping into the thylakoid lumen, an alkaline stromal pH develops, which is required for full activation of pH-dependent Calvin Benson cycle enzymes. This implies that a pH gradient between the cytosol (pH 7) and the stroma (pH 8) is established upon illumination. To maintain this pH gradient chloroplasts actively extrude protons. More than 30 years ago it was already established that these proton fluxes are electrically counterbalanced by Mg(2+), K(+), or Cl(-) fluxes, but only recently the first transport systems that regulate the pH gradient were identified. Notably several (Na(+),K(+))/H(+) antiporter systems where identified, that play a role in pH gradient regulation, ion homeostasis, osmoregulation, or coupling of secondary active transport. The established pH gradients are important to drive uptake of e...

Research paper thumbnail of Protein architecture and core residues in unwound α-helices provide insights to the transport function of plant AtCHX17

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2016

Using Arabidopsis thaliana AtCHX17 as an example, we combine structural modeling and mutagenesis ... more Using Arabidopsis thaliana AtCHX17 as an example, we combine structural modeling and mutagenesis to provide insights on its protein architecture and transport function which is poorly characterized. This approach is based on the observation that protein structures are significantly more conserved in evolution than linear sequences, and mechanistic similarities among diverse transporters are emerging. Two homology models of AtCHX17 were obtained that show a protein fold similar to known structures of bacterial Na(+)/H(+) antiporters, EcNhaA and TtNapA. The distinct secondary and tertiary structure models highlighted residues at positions potentially important for CHX17 activity. Mutagenesis showed that asparagine-N200 and aspartate-D201 inside transmembrane5 (TM5), and lysine-K355 inside TM10 are critical for AtCHX17 activity. We reveal previously unrecognized threonine-T170 and lysine-K383 as key residues at unwound regions in the middle of TM4 and TM11 α-helices, respectively. Mutation of glutamate-E111 located near the membrane surface inhibited AtCHX17 activity, suggesting a role in pH sensing. The long carboxylic tail of unknown purpose has an alternating β-sheet and α-helix secondary structure that is conserved in prokaryote universal stress proteins. These results support the overall architecture of AtCHX17 and identify D201, N200 and novel residues T170 and K383 at the functional core which likely participates in ion recognition, coordination and/or translocation, similar to characterized cation/H(+) exchangers. The core of AtCHX17 models according to EcNhaA and TtNapA templates faces inward and outward, respectively, which may reflect two conformational states of the alternating access transport mode for proteins belonging to the plant CHX family.

Research paper thumbnail of Plant NHX cation/proton antiporters

Plant Signaling Behavior, 2009

Addendum to: This manuscript has been published online, prior to printing. Once the issue is comp... more Addendum to: This manuscript has been published online, prior to printing. Once the issue is complete and page numbers have been assigned, the citation will change accordingly.

Research paper thumbnail of Heterologously expressed protein phosphatase calcineurin downregulates plant plasma membrane H +-ATPase activity at the post-translational level

Febs Letters, Oct 8, 2004

To investigate the effects of calcineurin expression on cellular ion homeostasis in plants, we ha... more To investigate the effects of calcineurin expression on cellular ion homeostasis in plants, we have obtained a transgenic cell culture of tomato, expressing constitutively activated yeast calcineurin. Transgenic cells exhibited reduced growth rates and proton extrusion activity in vivo. We show that reduction of plasma membrane H þ -ATPase activity by expression of calcineurin is the basis for the observed phenotypes. Transgenic calli and cell suspensions displayed also increased salt tolerance and contained slightly higher Ca 2þ and K þ levels. This demonstrates that calcineurin can modulate ion homeostasis in plants as it does in yeast by affecting the activity of primary ion transporters.

Research paper thumbnail of Conserved and diversified gene families of monovalent cation/h(+) antiporters from algae to flowering plants

Frontiers in plant science, 2012

All organisms have evolved strategies to regulate ion and pH homeostasis in response to developme... more All organisms have evolved strategies to regulate ion and pH homeostasis in response to developmental and environmental cues. One strategy is mediated by monovalent cation-proton antiporters (CPA) that are classified in two superfamilies. Many CPA1 genes from bacteria, fungi, metazoa, and plants have been functionally characterized; though roles of plant CPA2 genes encoding K(+)-efflux antiporter (KEA) and cation/H(+) exchanger (CHX) families are largely unknown. Phylogenetic analysis showed that three clades of the CPA1 Na(+)-H(+) exchanger (NHX) family have been conserved from single-celled algae to Arabidopsis. These are (i) plasma membrane-bound SOS1/AtNHX7 that share ancestry with prokaryote NhaP, (ii) endosomal AtNHX5/6 that is part of the eukaryote Intracellular-NHE clade, and (iii) a vacuolar NHX clade (AtNHX1-4) specific to plants. Early diversification of KEA genes possibly from an ancestral cyanobacterium gene is suggested by three types seen in all plants. Intriguingly, ...