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Nigel M Crawford - One of the best experts on this subject based on the ideXlab platform.
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the arabidopsis chl1 protein plays a major role in high affinity Nitrate Uptake
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Rongchen Wang, Dong Liu, Nigel M CrawfordAbstract:The CHL1 (NRT1) gene of Arabidopsis encodes a Nitrate-inducible Nitrate transporter that is thought to be a component of the low-affinity (mechanism II) Nitrate-Uptake system in plants. A search was performed to find high-affinity (mechanism I) Uptake mutants by using chlorate selections on plants containing Tag1 transposable elements. Chlorate-resistant mutants defective in high-affinity Nitrate Uptake were identified, and one had a Tag1 insertion in chl1, which was responsible for the phenotype. Further analysis showed that chl1 mutants have reduced high-affinity Uptake in induced plants and are missing a saturable component of the constitutive, high-affinity Uptake system in addition to reduced low-affinity Uptake. The contribution of CHL1 to constitutive high-affinity Uptake is higher when plants are grown at more acidic pH, conditions that increase the level of CHL1 mRNA. chl1 mutants show reduced membrane depolarization in root epidermal cells in response to low (250 μM) and high (10 mM) concentrations of Nitrate. Low levels of Nitrate (100 μM) induce a rapid increase in CHL1 mRNA. These results show that CHL1 is an important component of both the high-affinity and the low-affinity Nitrate-Uptake systems and indicate that CHL1 may be a dual-affinity Nitrate transporter.
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chl1 encodes a component of the low affinity Nitrate Uptake system in arabidopsis and shows cell type specific expression in roots
The Plant Cell, 1996Co-Authors: Nienchen Huang, Nigel M Crawford, Chiensung Chiang, Yifang TsayAbstract:The Arabidopsis CHL1 (AtNRT1) gene confers sensitivity to the herbicide chlorate and encodes a Nitrate-regulated Nitrate transporter. However, how CHL1 participates in Nitrate Uptake in plants is not yet clear. In this study, we examined the in vivo function of CHL1 with in vivo Uptake measurements and in situ hybridization experiments. Under most conditions tested, the amount of Nitrate Uptake by a chl1 deletion mutant was found to be significantly less than that of the wild type. This Uptake deficiency was reversed when a CHL1 cDNA clone driven by the cauliflower mosaic virus 35S promoter was expressed in transgenic chl1 plants. Furthermore, tissue-specific expression patterns showed that near the root tip, CHL1 mRNA is found primarily in the epidermis, but further from the root tip, the mRNA is found in the cortex or endodermis. These results are consistent with the involvement of CHL1 in Nitrate Uptake at different stages of root cell development. A functional analysis in Xenopus oocytes indicated that CHL1 is a low-affinity Nitrate transporter with a K(m) value of approximately 8.5 mM for Nitrate. This finding is consistent with the chlorate resistance phenotype of chl1 mutants. However, these results do not fit the current model of a single, constitutive component for the low-affinity Uptake system. To reconcile this discrepancy and the complex Uptake behavior observed, we propose a "two-gene" model for the low-affinity Nitrate Uptake system of Arabidopsis.
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genetic identification of a gene involved in constitutive high affinity Nitrate transport in higher plants
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Rongchen Wang, Nigel M CrawfordAbstract:Abstract Two mutations have been found in a gene (NRT2) of Arabidopsis thaliana that specifically impair constitutive, high-affinity Nitrate Uptake. These mutants were selected for resistance to 0.1 mM chlorate in the absence of Nitrate. Progency from one of the backcrossed mutants showed no constitutive Uptake of Nitrate below 0.5 mM at pH 7.0 in liquid culture (that is, within 30 min of initial exposure to Nitrate). All other Uptake activities measured (high-affinity phosphate and sulfate Uptake, inducible high-affinity Nitrate Uptake, and constitutive low-affinity Nitrate Uptake) were present or nearly normal in the backcrossed mutant. Electrophysiological analysis of individual root cells showed that the nrt2 mutant showed little response to 0.25 mM of Nitrate, whereas NRT2 wild-type cells showed an initial depolarization followed by recovery. At 10 mM of Nitrate both the mutant and wild-type cells displayed similar, strong electrical responses. These results indicate that NRT2 is a critical and perhaps necessary gene for constitutive, high-affinity Nitrate Uptake in Arabidopsis, but not for inducible, high-affinity nor constitutive, low-affinity Nitrate Uptake. Thus, these systems are genetically distinct.
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the herbicide sensitivity gene chl1 of arabidopsis encodes a Nitrate inducible Nitrate transporter
Cell, 1993Co-Authors: Yifang Tsay, Julian I Schroeder, Kenneth A Feldmann, Nigel M CrawfordAbstract:This paper reports the identification and functional expression of a gene that is involved in Nitrate Uptake in plants, a process essential for the assimilation of Nitrate and the biological removal of Nitrate from the soil solution. The CHL1 gene of Arabidopsis, which when mutated confers resistance to the herbicide chlorate and a decrease in Nitrate Uptake, was isolated and found to encode a protein with 12 putative membrane-spanning segments. Injection of CHL1 mRNA into Xenopus oocytes produces a Nitrate- and pH-dependent membrane depolarization, inward current, and Nitrate Uptake. These data show that the CHL1 gene encodes an electrogenic Nitrate transporter. CHL1 mRNA is found predominantly in roots and displays Nitrate- and pH-dependent regulation.
Yifang Tsay - One of the best experts on this subject based on the ideXlab platform.
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Uptake allocation and signaling of Nitrate
Trends in Plant Science, 2012Co-Authors: Ya-yun Wang, Yifang TsayAbstract:Plants need to acquire nitrogen (N) efficiently from the soil for growth. Nitrate is one of the major N sources for higher plants. Therefore, Nitrate Uptake and allocation are key factors in efficient N utilization. Membrane-bound transporters are required for Nitrate Uptake from the soil and for the inter- and intracellular movement of Nitrate inside the plants. Four gene families, Nitrate transporter 1/peptide transporter (NRT1/PTR), NRT2, chloride channel (CLC), and slow anion channel-associated 1 homolog 3 (SLAC1/SLAH), are involved in Nitrate Uptake, allocation, and storage in higher plants. Recent studies of these transporters or channels have provided new insights into the molecular mechanisms of Nitrate Uptake and allocation. Interestingly, several of these transporters also play versatile roles in Nitrate sensing, plant development, pathogen defense, and/or stress response.
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cloning and functional characterization of an arabidopsis Nitrate transporter gene that encodes a constitutive component of low affinity Uptake
The Plant Cell, 1999Co-Authors: Nienchen Huang, Kunhsiang Liu, Yifang TsayAbstract:The Arabidopsis CHL1 ( AtNRT1 ) gene encodes an inducible component of low-affinity Nitrate Uptake, which necessitates a “two-component” model to account for the constitutive low-affinity Uptake observed in physiological studies. Here, we report the cloning and characterization of a CHL1 homolog, AtNRT1:2 (originally named NTL1 ), with data to indicate that this gene encodes a constitutive component of low-affinity Nitrate Uptake. Transgenic plants expressing antisense AtNRT1:2 exhibited reduced Nitrate-induced membrane depolarization and Nitrate Uptake activities in assays with 10 mM Nitrate. Furthermore, transgenic plants expressing antisense AtNRT1:2 in the chl1-5 background exhibited an enhanced resistance to chlorate (7 mM as opposed to 2 mM for the chl1-5 mutant). Kinetic analysis of AtNRT1:2 -injected Xenopus oocytes yielded a K m for Nitrate of ∼5.9 mM. In contrast to CHL1 , AtNRT1:2 was constitutively expressed before and after Nitrate exposure (it was repressed transiently only when the level of CHL1 mRNA started to increase significantly), and its mRNA was found primarily in root hairs and the epidermis in both young (root tips) and mature regions of roots. We conclude that low-affinity systems of Nitrate Uptake, like high-affinity systems, are composed of inducible and constitutive components and that with their distinct functions, they are part of an elaborate Nitrate Uptake network in Arabidopsis.
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chl1 is a dual affinity Nitrate transporter of arabidopsis involved in multiple phases of Nitrate Uptake
The Plant Cell, 1999Co-Authors: Chiying Huang, Kunhsiang Liu, Yifang TsayAbstract:Higher plants have both high- and low-affinity Nitrate Uptake systems. These systems are generally thought to be genetically distinct. Here, we demonstrate that a well-known low-affinity Nitrate Uptake mutant of Arabidopsis, chl1 , is also defective in high-affinity Nitrate Uptake. Two to 3 hr after Nitrate induction, Uptake activities of various chl1 mutants at 250 μM Nitrate (a high-affinity concentration) were only 18 to 30% of those of wild-type plants. In these mutants, both the inducible phase and the constitutive phase of high-affinity Nitrate Uptake activities were reduced, with the inducible phase being severely reduced. Expressing a CHL1 cDNA driven by the cauliflower mosaic virus 35S promoter in a transgenic chl1 plant effectively recovered the defect in high-affinity Uptake for the constitutive phase but not for the induced phase, which is consistent with the constitutive level of CHL1 expression in the transgenic plant. Kinetic analysis of Nitrate Uptake by CHL1 -injected Xenopus oocytes displayed a biphasic pattern with a Michaelis–Menten K m value of ~50 μM for the high-affinity phase and ~4 mM for the low-affinity phase. These results indicate that in addition to being a low-affinity Nitrate transporter, as previously recognized, CHL1 is also involved in both the inducible and constitutive phases of high-affinity Nitrate Uptake in Arabidopsis.
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chl1 encodes a component of the low affinity Nitrate Uptake system in arabidopsis and shows cell type specific expression in roots
The Plant Cell, 1996Co-Authors: Nienchen Huang, Nigel M Crawford, Chiensung Chiang, Yifang TsayAbstract:The Arabidopsis CHL1 (AtNRT1) gene confers sensitivity to the herbicide chlorate and encodes a Nitrate-regulated Nitrate transporter. However, how CHL1 participates in Nitrate Uptake in plants is not yet clear. In this study, we examined the in vivo function of CHL1 with in vivo Uptake measurements and in situ hybridization experiments. Under most conditions tested, the amount of Nitrate Uptake by a chl1 deletion mutant was found to be significantly less than that of the wild type. This Uptake deficiency was reversed when a CHL1 cDNA clone driven by the cauliflower mosaic virus 35S promoter was expressed in transgenic chl1 plants. Furthermore, tissue-specific expression patterns showed that near the root tip, CHL1 mRNA is found primarily in the epidermis, but further from the root tip, the mRNA is found in the cortex or endodermis. These results are consistent with the involvement of CHL1 in Nitrate Uptake at different stages of root cell development. A functional analysis in Xenopus oocytes indicated that CHL1 is a low-affinity Nitrate transporter with a K(m) value of approximately 8.5 mM for Nitrate. This finding is consistent with the chlorate resistance phenotype of chl1 mutants. However, these results do not fit the current model of a single, constitutive component for the low-affinity Uptake system. To reconcile this discrepancy and the complex Uptake behavior observed, we propose a "two-gene" model for the low-affinity Nitrate Uptake system of Arabidopsis.
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the herbicide sensitivity gene chl1 of arabidopsis encodes a Nitrate inducible Nitrate transporter
Cell, 1993Co-Authors: Yifang Tsay, Julian I Schroeder, Kenneth A Feldmann, Nigel M CrawfordAbstract:This paper reports the identification and functional expression of a gene that is involved in Nitrate Uptake in plants, a process essential for the assimilation of Nitrate and the biological removal of Nitrate from the soil solution. The CHL1 gene of Arabidopsis, which when mutated confers resistance to the herbicide chlorate and a decrease in Nitrate Uptake, was isolated and found to encode a protein with 12 putative membrane-spanning segments. Injection of CHL1 mRNA into Xenopus oocytes produces a Nitrate- and pH-dependent membrane depolarization, inward current, and Nitrate Uptake. These data show that the CHL1 gene encodes an electrogenic Nitrate transporter. CHL1 mRNA is found predominantly in roots and displays Nitrate- and pH-dependent regulation.
Kunhsiang Liu - One of the best experts on this subject based on the ideXlab platform.
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cloning and functional characterization of an arabidopsis Nitrate transporter gene that encodes a constitutive component of low affinity Uptake
The Plant Cell, 1999Co-Authors: Nienchen Huang, Kunhsiang Liu, Yifang TsayAbstract:The Arabidopsis CHL1 ( AtNRT1 ) gene encodes an inducible component of low-affinity Nitrate Uptake, which necessitates a “two-component” model to account for the constitutive low-affinity Uptake observed in physiological studies. Here, we report the cloning and characterization of a CHL1 homolog, AtNRT1:2 (originally named NTL1 ), with data to indicate that this gene encodes a constitutive component of low-affinity Nitrate Uptake. Transgenic plants expressing antisense AtNRT1:2 exhibited reduced Nitrate-induced membrane depolarization and Nitrate Uptake activities in assays with 10 mM Nitrate. Furthermore, transgenic plants expressing antisense AtNRT1:2 in the chl1-5 background exhibited an enhanced resistance to chlorate (7 mM as opposed to 2 mM for the chl1-5 mutant). Kinetic analysis of AtNRT1:2 -injected Xenopus oocytes yielded a K m for Nitrate of ∼5.9 mM. In contrast to CHL1 , AtNRT1:2 was constitutively expressed before and after Nitrate exposure (it was repressed transiently only when the level of CHL1 mRNA started to increase significantly), and its mRNA was found primarily in root hairs and the epidermis in both young (root tips) and mature regions of roots. We conclude that low-affinity systems of Nitrate Uptake, like high-affinity systems, are composed of inducible and constitutive components and that with their distinct functions, they are part of an elaborate Nitrate Uptake network in Arabidopsis.
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chl1 is a dual affinity Nitrate transporter of arabidopsis involved in multiple phases of Nitrate Uptake
The Plant Cell, 1999Co-Authors: Chiying Huang, Kunhsiang Liu, Yifang TsayAbstract:Higher plants have both high- and low-affinity Nitrate Uptake systems. These systems are generally thought to be genetically distinct. Here, we demonstrate that a well-known low-affinity Nitrate Uptake mutant of Arabidopsis, chl1 , is also defective in high-affinity Nitrate Uptake. Two to 3 hr after Nitrate induction, Uptake activities of various chl1 mutants at 250 μM Nitrate (a high-affinity concentration) were only 18 to 30% of those of wild-type plants. In these mutants, both the inducible phase and the constitutive phase of high-affinity Nitrate Uptake activities were reduced, with the inducible phase being severely reduced. Expressing a CHL1 cDNA driven by the cauliflower mosaic virus 35S promoter in a transgenic chl1 plant effectively recovered the defect in high-affinity Uptake for the constitutive phase but not for the induced phase, which is consistent with the constitutive level of CHL1 expression in the transgenic plant. Kinetic analysis of Nitrate Uptake by CHL1 -injected Xenopus oocytes displayed a biphasic pattern with a Michaelis–Menten K m value of ~50 μM for the high-affinity phase and ~4 mM for the low-affinity phase. These results indicate that in addition to being a low-affinity Nitrate transporter, as previously recognized, CHL1 is also involved in both the inducible and constitutive phases of high-affinity Nitrate Uptake in Arabidopsis.
Stuart J Ferguson - One of the best experts on this subject based on the ideXlab platform.
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the paracoccus denitrificans nark like Nitrate and nitrite transporters probing Nitrate Uptake and Nitrate nitrite exchange mechanisms
Molecular Microbiology, 2017Co-Authors: Alan D Goddard, Shilpa Bali, Despoina A I Mavridou, Victor M Luquealmagro, Andrew J Gates, Dolores M Roldan, Simon Newstead, D J Richardson, Stuart J FergusonAbstract:Nitrate and nitrite transport across biological membranes is often facilitated by protein transporters that are members of the major facilitator superfamily. Paracoccus denitrificans contains an unusual arrangement whereby two of these transporters, NarK1 and NarK2, are fused into a single protein, NarK, which delivers Nitrate to the respiratory Nitrate reductase and transfers the product, nitrite, to the periplasm. Our complementation studies, using a mutant lacking the Nitrate/proton symporter NasA from the assimilatory Nitrate reductase pathway, support that NarK1 functions as a Nitrate/proton symporter while NarK2 is a Nitrate/nitrite antiporter. Through the same experimental system, we find that Escherichia coli NarK and NarU can complement deletions in both narK and nasA in P. denitrificans, suggesting that, while these proteins are most likely Nitrate/nitrite antiporters, they can also act in the net Uptake of Nitrate. Finally, we argue that primary sequence analysis and structural modelling do not readily explain why NasA, NarK1 and NarK2, as well as other transporters from this protein family, have such different functions, ranging from net Nitrate Uptake to Nitrate/nitrite exchange.
Rongchen Wang - One of the best experts on this subject based on the ideXlab platform.
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the arabidopsis chl1 protein plays a major role in high affinity Nitrate Uptake
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Rongchen Wang, Dong Liu, Nigel M CrawfordAbstract:The CHL1 (NRT1) gene of Arabidopsis encodes a Nitrate-inducible Nitrate transporter that is thought to be a component of the low-affinity (mechanism II) Nitrate-Uptake system in plants. A search was performed to find high-affinity (mechanism I) Uptake mutants by using chlorate selections on plants containing Tag1 transposable elements. Chlorate-resistant mutants defective in high-affinity Nitrate Uptake were identified, and one had a Tag1 insertion in chl1, which was responsible for the phenotype. Further analysis showed that chl1 mutants have reduced high-affinity Uptake in induced plants and are missing a saturable component of the constitutive, high-affinity Uptake system in addition to reduced low-affinity Uptake. The contribution of CHL1 to constitutive high-affinity Uptake is higher when plants are grown at more acidic pH, conditions that increase the level of CHL1 mRNA. chl1 mutants show reduced membrane depolarization in root epidermal cells in response to low (250 μM) and high (10 mM) concentrations of Nitrate. Low levels of Nitrate (100 μM) induce a rapid increase in CHL1 mRNA. These results show that CHL1 is an important component of both the high-affinity and the low-affinity Nitrate-Uptake systems and indicate that CHL1 may be a dual-affinity Nitrate transporter.
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genetic identification of a gene involved in constitutive high affinity Nitrate transport in higher plants
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Rongchen Wang, Nigel M CrawfordAbstract:Abstract Two mutations have been found in a gene (NRT2) of Arabidopsis thaliana that specifically impair constitutive, high-affinity Nitrate Uptake. These mutants were selected for resistance to 0.1 mM chlorate in the absence of Nitrate. Progency from one of the backcrossed mutants showed no constitutive Uptake of Nitrate below 0.5 mM at pH 7.0 in liquid culture (that is, within 30 min of initial exposure to Nitrate). All other Uptake activities measured (high-affinity phosphate and sulfate Uptake, inducible high-affinity Nitrate Uptake, and constitutive low-affinity Nitrate Uptake) were present or nearly normal in the backcrossed mutant. Electrophysiological analysis of individual root cells showed that the nrt2 mutant showed little response to 0.25 mM of Nitrate, whereas NRT2 wild-type cells showed an initial depolarization followed by recovery. At 10 mM of Nitrate both the mutant and wild-type cells displayed similar, strong electrical responses. These results indicate that NRT2 is a critical and perhaps necessary gene for constitutive, high-affinity Nitrate Uptake in Arabidopsis, but not for inducible, high-affinity nor constitutive, low-affinity Nitrate Uptake. Thus, these systems are genetically distinct.