JXB Advance Access published online on June 19, 2007
Journal of Experimental Botany, doi:10.1093/jxb/erm106
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
FOCUS PAPER |
Inorganic nitrogen assimilation in Chlamydomonas
Departamento de Bioquímica y Biología Molecular, Universidad de Córdoba, Campus de Rabanales, Edificio Severo Ochoa, Córdoba 14071, Spain
* To whom correspondence should be addressed. E-mail: bb1feree{at}uco.es
Received 3 January 2007; Revised 26 March 2007 Accepted 16 April 2007
 |
Abstract |
|---|
 Top Abstract The structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
Inorganic nitrogen is an essential nutrient for photosynthetic organisms. Its efficient use in nature involves adaptation of the organisms to the availability of the nitrogen supply, to changing environmental conditions, and to the provision of carbon and other nutrients. The unicellular alga Chlamydomonas provides a useful model to identify not only each of the components participating in the assimilative process in a species, but also the regulatory networks modulating their activity. A remarkable fact is the ample array of transporters for inorganic nitrogen compounds operating in this single cell: 13 putative nitrate/nitrite transporters and eight putative ammonium transporters. However, for nitrate, only a few of them participate as the main suppliers of nitrogen for cell growth, and others probably function to adapt nitrogen utilization efficiency to conditions depending not only on the nitrogen source available but also on other nutrients and environmental conditions. This paper summarizes recent findings in Chlamydomonas to provide an integrated perspective.
Key words: Ammonium transport, bicarbonate transport, Chlamydomonas, negative control, nitrate signalling, nitrate transport, nitrite transport, plastidic nitrite transport
|
The structural components participating in inorganic N assimilation in Chlamydomonas |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
Inorganic N assimilation is a key and apparently simple process in mineral nutrition. For nitrate, it consists of two transport and two reduction steps separated into the cytosol (NO3–
NO2–) and the chloroplast (NO2–NH4+). Ammonium is finally incorporated into carbon skeletons by the glutamine synthetase/glutamate synthase pathway (Crawford and Arst, 1993; Crawford, 1995; Daniel-Vedele et al., 1998). Functional genomics approaches and the use of model organisms have allowed the identification of genes and functions for the basic structural components. However, the regulatory circuits are far more complex to decipher since they should have evolved together with the plasticity of the organisms to adapt to changes in nutrient accessibility in addition to their developmental complexity. Thus, new strategies are needed to understand them. The haploid unicellular green alga Chlamydomonas reinhardtii has been shown to be an excellent model system for studies of fundamental biological processes such as the photosynthetic apparatus, microtubule assembly, flagella movement, or mineral nutrition, among others (Rochaix et al., 1998). This has been possible because of the development of excellent biochemical, molecular, genetic, and genomic tools. In addition, the Genome Sequence Project (http://www.chlamy.org/) is promoting the use of this organism in functional and comparative genomics studies.
An interesting analysis is derived from the number and type of genes involved in N assimilation in Chlamydomonas when compared with other eukaryotic photosynthetic model organisms: Arabidopsis, a plant; and Ostreococcus, the smallest unicellular green alga (Derelle et al., 2006; Table 1). The large number of putative transporters for inorganic N in comparison with the number of enzymes of the route is noteworthy. The Chlamydomonas genome contains eight putative ammonium transporters (Amt1) and 13 putative nitrate/nitrite transporters (one Nrt1, six Nrt2, and six Nar1). All these gene families are present in the Ostreococcus genome in much lower numbers, with some difference (two Amt2 are present), and in Arabidopsis in similar numbers but from different families (Nar1 is absent, and Amt2 present).
|
A general picture of nitrate, nitrite, and ammonium assimilation in Chlamydomonas is shown in Fig. 1. Some of the direct gene products have been well characterized, and others come from the Chlamydomonas genome database and will require further studies. Different components of the pathway are highlighted here. (i) The family of Nrt2 genes, comprised of six members, seems to be responsible for the entry of nitrate/nitrite with high affinity into the cell (HAN/NiT). Two members of the family (Nrt2.1 and Nrt2.2) require a second component (Nar2) for functionality (Quesada et al., 1994; Zhou et al., 2000a). (ii) A family of eight AMT1-type ammonium transporters might be related to the transport of ammonium, and representative members could be located at both the plasma and chloroplast membranes (Gonzalez-Ballester et al., 2004). (iii) Two Pma genes encoding H+ATPases are probably needed for generation of the electrochemical gradient for the transport of nutrients (unpublished data from our group). (iv) An Nia1 gene encodes nitrate reductase, requiring six Cnx genes plus an Mcp gene for biosynthesis and transfer of the molybdenum cofactor to the nitrate reductase enzyme and to other molybdoenzymes (Fernandez et al., 1989; Ataya et al., 2003; Fischer et al., 2006). (v) There is a single Nrt1 gene of unknown function. (vi) Six genes from the Nar1 family are putative nitrite and bicarbonate transporters, some of which encode plastidic transporters, and are differentially regulated by carbon and nitrogen (Rexach et al., 2000; Galvan and Fernandez, 2001; Galvan et al., 2002; Mariscal et al., 2006). (vii) There is a single Nii1 gene for the plastidic nitrite reductase (Quesada et al., 1998b). (viii) There are three glutamine synthetase genes for plastidic (two) and cytosolic (one) forms (Chen and Silflow, 1996), two genes for GOGAT (one for the ferredoxin-dependent and another for the NADH-dependent enzyme), and one gene for glutamate dehydrogenase. This plastidic location of complete GS/GOGAT cycles dependent on NADH or ferredoxin points to this organelle as the main site for ammonium incorporation under photosynthetic and non-photosynthetic conditions. Glutamine dehydrogenase has a probable mitochondrial localization and an anaplerotic role in the tricarboxylic acid cycle (Masclaux-Daubresse et al., 2006). (ix) Finally, a gene for a plastidic malate dehydrogenase (NMdh) and another for mitochondrial alternative oxidase (Aox1) are clustered with the main genes for nitrate assimilation and provide a direct molecular link between NADH and ATP pools for nitrate assimilation (Quesada et al., 2000; Galvan et al., 2002). A regulatory gene, Nit2, tightly and positively controls the expression of many genes for nitrate assimilation that are clustered on chromosome IX (Quesada and Fernandez, 1994; Galvan et al., 2006).
|
|
Nitrate and nitrite have specific transport systems |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
The Chlamydomonas genome contains three different gene families (Nrt1, Nrt2, and Nar1) encoding putative nitrate/nitrite transporters. The existence of an NRT1 transporter is suggested only from the genome database, and a molecular/functional characterization of this system is needed. However, it is remarkable that Nrt1 genes are widely distributed and abundant in plant genomes, although only a few (four in Arabidopsis) are related to nitrate transport (Williams and Miller, 2001). The AtNrt1.1 gene is the best studied gene from this family and is involved in regulating multiple functions. (i) AtNRT1.1 appears to be a dual affinity nitrate transporter that shows high affinity (HANTS) or low affinity (LANTS) depending on its phosphorylation state. The phosphorylated form (HANTS) is triggered under limited external NO3–, adapting the functional properties of the transporter to the resource level in the root environment (Wang et al., 1998; Liu and Tsay, 2003). (ii) AtNRT1.1 has a crucial role in early phases of the development of young organs (root tips, emerging lateral roots, and nascent leaves) (Guo et al., 2001). (iii) AtNRT1.1 contributes to drought susceptibility because of its role in stomatal opening (Guo et al., 2003). Two model microalgae (Ostreococcus and Chlamydomonas) contain a single Nrt1 gene, and it will be interesting in the future to determine the role(s) of this putative transporter in algae.
NRT2 transporters are also widely distributed and constitute a family comprised of six and seven members in Chlamydomonas and Arabidopsis, respectively. In Arabidopsis, these genes are differentially expressed in plant tissues with a pattern that can depend on the external N supply (Orsel et al., 2002; Okamoto et al., 2003). AtNrt2.1 is the major inducible HANT and contributes to normal plant development in low nitrate (Filleur et al., 2001; Little et al., 2005; Li et al., 2007).
NRT2 proteins have typical carrier-type structures with 12 putative transmembrane (TM) domains; however, some of them require for functionality a second component consisting of a TM domain protein (NAR2). Thus, NRT2 transporters are classified into single- (NRT2) or two-component (NRT2/NAR2) systems (Galvan et al., 1996; Zhou et al., 2000a; Tong et al., 2005; Okamoto et al., 2006; Orsel et al., 2006).
Comparative studies of Chlamydomonas strains containing or lacking particular transport genes have resulted in the following information. To date, four different transport systems have been identified as being involved in nitrate/nitrite transport into cells, all with high affinity but with different ion specificity and regulation (Table 2). Transport system I is bispecific for nitrate and nitrite (HANT/HANiT) and requires Nrt2.1 and Nar2. The nitrate-specific transport system II (HANT) requires Nrt2.2 and Nar2 (Galvan et al., 1996; Zhou et al., 2000a). System III, which is Nar2 independent, shows high affinity for nitrite (HANiT) but low affinity for nitrate, and probably corresponds to NRT2;3 (Quesada et al., 1998a; Rexach et al., 1999). Finally, system IV is bispecific for nitrate and nitrite (HANT/HANiT) and is proposed to be encoded by a member of the Nrt2 family in Chlamydomonas (Rexach et al., 1999; Navarro et al., 2000). These four transporters show a differential regulation by ammonium, CO2, and chloride. Systems I, II, and III operate optimally at high CO2 and are inhibited by ammonium but not by chloride. In contrast, system IV operates efficiently at low CO2 and is inhibited by chloride but not by ammonium (Galvan et al., 1996; Rexach et al., 1999). These transport systems I, II, and III are induced by nitrate and repressed by ammonium, and system IV is constitutively expressed. Interestingly, there are several facts that should be pointed out.
|
- (i) Systems I and II not only have a high affinity for nitrate, especially system I, but also have the highest capacity for mediating efficient growth (Table 2). Mutants lacking these two systems can only grow on nitrate media at a very low rate, even though other genes Nrt2 (Nrt2.3–Nrt2.6) and Nar2 exist. These Chlamydomonas HANT mutants are distinguished as pale green colonies in nitrate media, and can be transformed with plasmids having either Nrt2.1+Nar2 or Nrt2.2+Nar2 genes, giving rise to transformants selectable as dark green colonies.
- (ii) NAR2 is only required for these two HANTs (I and II), but not for other HAN/NiT systems such as systems III and IV, indicating that transporters of the NRT2 family can function either as two-component HATS, as also shown in plants (Quesada et al., 1994; Zhou et al., 2000a; Tong et al., 2005; Okamoto et al., 2006; Orsel et al., 2006), or just as a one-component system as also shown in Aspergillus NRTA (CRNA) with a different structural organization from the plant-type NRT2 (Unkles et al., 1991; Rexach et al., 1999; Navarro et al., 2000; Zhou et al., 2000b; Derelle et al., 2006).
- (iii) Though nitrate is the main form of oxidized nitrogen in natural environments, nitrite being very scarce (Crawford, 1995; Glass et al., 2002), the nitrite anion is efficiently assimilated in Chlamydomonas. These four HANTs show a very definite selectivity towards nitrate or nitrite. The HANT systems I and IV are bispecific for both anions, whereas systems II and III are more selective for nitrate and nitrite, respectively (Table 2).
- (ii) NAR2 is only required for these two HANTs (I and II), but not for other HAN/NiT systems such as systems III and IV, indicating that transporters of the NRT2 family can function either as two-component HATS, as also shown in plants (Quesada et al., 1994; Zhou et al., 2000a; Tong et al., 2005; Okamoto et al., 2006; Orsel et al., 2006), or just as a one-component system as also shown in Aspergillus NRTA (CRNA) with a different structural organization from the plant-type NRT2 (Unkles et al., 1991; Rexach et al., 1999; Navarro et al., 2000; Zhou et al., 2000b; Derelle et al., 2006).
|
Nitrite transport to the chloroplast is a mediated process and connected to bicarbonate transport |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
A third family of transporters for nitrate assimilation in Chlamydomonas corresponds to NAR1 proteins. The Nar1 family consists of six genes (Galvan et al., 2002; Mariscal et al., 2006) that encode proteins homologous to bacterial putative formate/nitrite transporters (FNT) (Peakman et al., 1990; Suppmann and Sawers, 1994; Rexach et al., 2000; Galvan et al., 2006). FNT proteins seem to be restricted to bacteria and protist organisms, and are absent in plants. This is especially interesting in some yeasts such as Saccharomyces cerevisiae, incapable of assimilating nitrate, suggesting that these NAR1 proteins might fulfil important influx or efflux transport activities across the membranes for nitrite or other structurally related monovalent anions (formate, bicarbonate, etc.).
Studies in Chlamydomonas support that NAR1.1 mediates nitrite transport to the chloroplast and makes nitrate assimilation efficient according to carbon availability (Rexach et al., 2000; Galvan et al., 2002). This indicates a cross-regulation point for carbon–nitrogen mediated by a transporter. However, this important function in plants is still to be identified.
Characterization of the whole Nar1 gene family and their deduced proteins has shown the following facts (Table 3).
|
- (i) NAR1.1, NAR1.2, and NAR1.5 are plastidic proteins containing putative chloroplast transit peptides, whereas NAR1.3, NAR1.4, and NAR1.6 are possibly plasma membrane proteins (Mariscal et al., 2006). Interestingly, phylogenetic analysis locates NAR1.1, NAR1.2, and NAR1.5 in a common branch separated from NAR1.3, NAR1.4, and NAR1.6 that group together.
- (ii) Two model proteins are deduced with either a short (NAR1.1, NAR1.2, NAR1.4, and NAR1.5) or a long (NAR1.3 and NAR1.6) C-terminus (Mariscal et al., 2006), though the physiological significance of this is unknown.
- (iii) Nar1 transcripts are differentially regulated by C and N sources and the general C and N regulatory genes: Nar1.1 and Nar1.6 by nitrogen and Nit2, the regulatory gene for nitrate assimilation (Rexach et al., 2000; Mariscal et al., 2004; Mariscal et al., 2006), and Nar1.2 by carbon and ccm1, the central regulatory gene for carbon assimilation (Miura et al., 2004; Mariscal et al., 2006). The other genes, Nar1.3, Nar1.4, and Nar1.5, show an expression pattern independent of Nit2 and ccm1 but slightly modulated by the nitrogen or carbon source.
- (iv) The functionality of NAR1 proteins as nitrite and bicarbonate transporters was successfully obtained by expression of NAR1.2 in Xenopus oocytes (Mariscal et al., 2006). These findings and those on NAR1.1 (Mariscal et al., 2004) support that nitrite and bicarbonate transport to the chloroplast are interlinked processes.
- (ii) Two model proteins are deduced with either a short (NAR1.1, NAR1.2, NAR1.4, and NAR1.5) or a long (NAR1.3 and NAR1.6) C-terminus (Mariscal et al., 2006), though the physiological significance of this is unknown.
|
Ammonium transport |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
Ammonium is the preferred nitrogen source for Chlamydomonas and negatively signals nitrate assimilation genes (Fernandez et al., 1998). Since ammonium is mostly incorporated into carbon skeletons in the chloroplast (Fig. 1), transporters are required at both plasma and chloroplast membrane levels to ensure appropriate ammonium assimilation. This green alga has the largest AMT1 family of ammonium transporters so far described, consisting of eight members (Gonzalez-Ballester et al., 2004). There are 10 putative Amt genes in rice, from four families, three each encoding OsAMT1, OsAMT2, and OsAMT3, and one encoding OsAMT4 (Suenaga et al., 2003). These AMT proteins belong to a superfamily having an additional component, the Rh protein, whose best known member is the human Rh blood group factor abundant in red blood cell membranes (Avent and Reid, 2000). Evidence has been provided that the Chlamydomonas Rh1 protein is a gas channel for CO2 (Soupene et al., 2002, 2004). The structure of the AmtB protein from Escherichia coli was determined at 1.35 Å resolution, showing that NH4+ is bound to ligands and deprotonated to NH3 gas that is finally transported though the protein channel (Khademi et al., 2004). The eight CrAMT1s have been phylogenetically analysed and shown to group into three subclasses: AMT1.1, AMT1.3, and AMT1.5 group together (subclass I) close to AMT1.2, AMT1.4, and AMT1.6 (subclass II), with AMT1.7 and AMT1.8 grouping in a separate branch (subclass III) (Gonzalez-Ballester et al., 2004). The putative localization of some CrAMTs is indicated in Fig. 1. It is also interesting to note that CrAmt1.5 and CrAmt1.7 show alternative splicing to two different transcripts whose significance will have to be determined (Gonzalez-Ballester et al., 2004; Kim et al., 2005).
CrAmt1 genes have differential expression patterns in response to the nitrogen source, although carbon does not seem to affect this. CrAmt1.1 and CrAmt1.2 are strongly induced in the absence of any nitrogen source, suggesting a role in high affinity transport (Gonzalez-Ballester et al., 2004). Expression of CrAmt1.4 is also enhanced in media containing a poor nitrogen source such as arginine (Kim et al., 2005). The negative effect of nitrate on CrAmt1.1 appears to be 2-fold; one effect has been related to its reduction to ammonium, and another to itself mediated by the regulatory gene Nit2, specific for nitrate assimilation. Thus, NIT2 was proposed to have a dual role on gene expression: the well-known positive one on nitrate assimilation (Fernandez et al., 1998) and a novel negative one on Amt1;1 regulation (Gonzalez-Ballester et al., 2004).
Methylammonium is a non-metabolizable ammonium analogue (Franco et al., 1984) that has been used to isolate methylammonium-resistant mutants and to study ammonium transport processes (Franco et al., 1988). Two activities were determined varying in their specificity and capacity for ammonium transport. However, the picture now appears to be much more complex. Sixteen spontaneous methylammonium-resistant mutants have been isolated and characterized in Chlamydomonas (Kim et al., 2005). They were affected in terms of their methylammonium accumulation capacity and uptake. Most mutations conferring resistance to methylammonium are in Amt1.4 (also named Amt4), and a high proportion of spontaneous mutations are caused by transposon-related events (Kim et al., 2006). The properties of the amt4 mutants are different from those of rh1 RNAi lines, supporting that the physiological substrates for AMT and RH proteins are NH3 and CO2, respectively (Kim et al., 2005).
|
How are nitrate and ammonium sensed? |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
One of the most intriguing and interesting questions of the nitrate pathway relates to the sensing of positive and negative signals modulating nitrate assimilation. Nitrate is not only an essential nutrient that activates the expression of genes involved in its assimilation pathway, but is also a signalling molecule (Crawford, 1995; Crawford and Glass, 1998; Daniel-Vedele et al., 1998; Fernandez et al., 1998; Forde, 2000) modulating metabolic carbon allocation (Scheible et al., 1997a), alternative pathways of the respiratory chain (Escobar et al., 2006), root–shoot balance (Scheible et al., 1997b), seed dormancy (Alboresi et al., 2005), and root development (Zhang et al., 2000; Walch-Liu et al., 2006) in plants and gametogenesis in Chlamydomonas (Pozuelo et al., 2000). On the other hand, ammonium, or glutamine, provides a negative signal, which leads to repression of nitrate transporters and enzymes of the pathway (Crawford and Arst, 1993; Crawford, 1995; Quesada et al., 1997; Fernandez et al., 1998; Vidmar et al., 2000).
In Chlamydomonas, it was shown that nitrate signalling on the Nia1 gene promoter occurs intracellularly and depends on the activity of nitrate transporters (Llamas et al., 2002). Other genes involved in nitrate/nitrite assimilation (Nii1, Nrt2;1, Nrt2;3, and Nar1) were also found to be optimally signalled by the activity of the HANT system I (NRT2;1, NAR2) (Llamas et al., 2002; Rexach et al., 2002). In Arabidopsis, it has been shown that NRT1.1-mediated repression of NRT2.1/NAR2.1 (NRT3.1) at high nitrate is relieved at low nitrate supply in the presence of ammonium; it has been proposed that NRT1.1-mediated regulation of NRT2.1/NAR2.1 is a mechanism able to satisfy a specific nitrate demand of the plant in relation to the various specific roles that nitrate plays, in addition to being an N source (Krouk et al., 2006).
The complex network of transporters in the nitrate assimilation pathway seems to be required to ensure efficient nitrogen incorporation for growth and differentiation under different environmental conditions. However, the precise role of each transporter in the improvement of nitrate use efficiency is not still understood.
In contrast to the well-characterized fungal systems (Crawford and Arst, 1993; Marzluf, 1997), no regulatory genes have been clearly connected to the nitrate assimilation pathway in plants (Daniel-Vedele et al., 1998); only in Chlamydomonas has a regulatory gene Nit2 been shown to be essential in the positive control of genes for the nitrate pathway (Fernandez and Matagne, 1986; Fernandez et al., 1998; Quesada et al., 1998a). Nit2 was cloned (Schnell and Lefebvre, 1993) and is presently being characterized both structurally and functionally (Camargo, 2006).
Concerning the negative control, different negative regulatory loci (Nrg1-4, Far1) identified by insertional mutagenesis have been reported to mediate ammonium repression with partial phenotypes for the nitrate transport and reduction steps (Prieto et al., 1996; Zhang and Lefebvre, 1997; Perez-Alegre et al., 2005). In plants, no genes mediating positive effects of nitrate and negative effects of ammonium in the nitrate pathway have been reported. It has previously been proposed that in plants several genes might participate, so that deficiencies in each of them would lead to partial phenotypes not strong enough for the detection of the lack of induction/repression effects in a mutant (Galvan et al., 2002). In fact, as has recently been found, ammonium-negative effects appear complex, mediated by different genes, and dependent on ammonium concentration (de Montaigu, 2006).
To identify regulatory elements of the pathway, a major effort has been made to target Chlamydomonas genes by constructing an ordered insertional mutant library. Insertional mutagenesis was performed in a strain bearing a ‘nitrate-pathway sensor gene’ for positive and negative regulation, the arylsulfatase reporter gene under the control of the Nia1 gene promoter (Gonzalez-Ballester et al., 2005). This procedure of insertions in the Chlamydomonas genome generally causes deletions of DNA fragments from 5 kb to 57 kb (Kindle, 1998; Pazour and Witman, 2000; Cenkci et al., 2003) that estimated a good coverage of the genome, making the collection representative for identifying non-essential genes related to the nitrate pathway. One hundred and forty-five mutants were isolated, with phenotypes of nitrate insensitivity, ammonium insensitivity, and overexpression in nitrate, that were capable or incapable of growing on nitrate medium (Gonzalez-Ballester et al., 2005). It has been shown that: (i) Nit2 is the key gene for positive nitrate signalling, encoding a transcriptional factor with structural characteristics different from the regulatory genes reported in fungi (Camargo, 2006); (ii) several genes and signalling cascades participate in the negative signalling by ammonium by means of complex pathways dependent on the ammonium concentration (de Montaigu, 2006); and (iii) in addition to Nit2, other genes modulate the positive signalling by nitrate.
|
Future perspectives |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
The nitrate pathway involves at least four cell compartments (cytosol, nucleus, chloroplast, and mitochondria) participating in expression, transport, reduction, and signalling steps. They are required to ensure the proper synthesis of enzymes and transporters, and the supply of ATP and reducing power needed for nitrate transport and reduction (Quesada et al., 2000). There are still some challenges that need to be addressed for a proper understanding of the pathway in photosynthetic eukaryotes. These include the precise role of each of the numerous transporters participating in providing not only the appropriate nutrients but also the proper signal for modulating the complete pathway. Nitrogen assimilation in photosynthetic eukaryotes has to be encompassed within that of other nutrients, where carbon plays a major role. Regulatory genes for positive and negative signalling of the nitrate pathway are starting to be identified, and their study in the organism as a whole will probably constitute a relevant task in the coming years.
|
Acknowledgements |
|---|
This work was supported by ‘Ministerio de Educación y Ciencia’, Spain (Grant BFU2005-07521), and Junta de Andalucía, Spain (PAI, CVI-0128).
|
Abbreviations |
|---|
HANT, high affinity nitrate transport; HANiT, high affinity nitrite transport; HAN/NiT, high affinity nitrate and nitrite transport; LANT, low affinity nitrate transport.
|
References |
|---|
|
TopAbstractThe structural components...Nitrate and nitrite have...Nitrite transport to the...Ammonium transportHow are nitrate and...Future perspectivesReferences |
|---|
Alboresi A, Gestin C, Leydecker MT, Bedu M, Meyer C, Truong HN. Nitrate, a signal relieving seed dormancy in Arabidopsis. Plant, Cell and Environment (2005) 28:500–512.[CrossRef][Medline]
Ataya FS, Witte CP, Galvan A, Igeno MI, Fernandez E. Mcp1 encodes the molybdenum cofactor carrier protein in Chlamydomonas reinhardtii and participates in protection, binding, and storage functions of the cofactor. Journal of Biological Chemistry (2003) 278:10885–10890.
Avent ND, Reid ME. The Rh blood group system: a review. Blood (2000) 95:375–387.
Camargo A. Regulación positiva de la asimilación de nitrato en algas y plantas. In: PhD dissertation (2006) Universidad de Cordoba: Spain.
Cenkci B, Petersen JL, Small GD. REX1, a novel gene required for DNA repair. Journal of Biological Chemistry (2003) 278:22574–22577.
Chen Q, Silflow CD. Isolation and characterization of glutamine synthetase genes in Chlamydomonas reinhardtii. Plant Physiology (1996) 112:987–996.[Abstract]
Crawford NM. Nitrate: nutrient and signal for plant growth. The Plant Cell (1995) 7:859–868.[CrossRef][Web of Science][Medline]
Crawford NM, Glass AD. Molecular and physiological aspects of nitrate uptake in plants. Trends in Plant Science (1998) 3:389–395.[CrossRef][Web of Science]
Crawford NM, Arst HN Jr. The molecular genetics of nitrate assimilation in fungi and plants. Annual Review of Genetics (1993) 27:115–146.[CrossRef][Web of Science][Medline]
Daniel-Vedele F, Filleur S, Caboche M. Nitrate transport: a key step in nitrate assimilation. Current Opinion in Plant Biology (1998) 1:235–239.[CrossRef][Web of Science][Medline]
de Montaigu A. Regulación negativa de la asimilación de nitrato en Chlamydomonas reinhardtii. PhD dissertation (2006) Universidad de Cordoba (Spain).
Derelle E, Ferraz C, Rombauts S, et al. Genome analysis of the smallest free-living eukaryote Ostreococcus tauri unveils many unique features. Proceedings of the National Academy of Sciences, USA (2006) 103:11647–11652.
Escobar MA, Geisler DA, Rasmusson AG. Reorganization of the alternative pathways of the Arabidopsis respiratory chain by nitrogen supply: opposing effects of ammonium and nitrate. The Plant Journal (2006) 45:775–788.[CrossRef][Web of Science][Medline]
Fernandez E, Matagne RF. In vivo complementation analysis of nitrate reductase-deficient mutants in Chlamydomonas reinhardtii. Current Genetics (1986) 10:397–403.[CrossRef][Web of Science][Medline]
Fernandez E, Schnell RA, Ranum LP, Hussey SC, Silflow CD, Lefebvre PA. Isolation and characterization of the nitrate reductase structural gene of Chlamydomonas reinhardtii. Proceedings of the National Academy of Sciences, USA (1989) 86:6449–6453.
Fernandez E, Galvan A, Quesada A. Nitrogen assimilation and its regulation. In: Molecular biology of Chlamydomonas: chloroplast and mitochondria—Rochaix JD, Goldschmidt-Clermont M, Merchant S, eds. (1998) Dordrecht, The Netherlands: Kluwer Academic Publishers. 637–659.
Filleur S, Dorbe MF, Cerezo M, Orsel M, Granier F, Gojon A, Daniel-Vedele F. An Arabidopsis T-DNA mutant affected in Nrt2 genes is impaired in nitrate uptake. FEBS Letters (2001) 489:220–224.[CrossRef][Web of Science][Medline]
Fischer K, Llamas A, Tejada-Jimenez M, Schrader N, Kuper J, Ataya FS, Galvan A, Mendel RR, Fernandez E, Schwarz G. Function and structure of the molybdenum cofactor carrier protein from Chlamydomonas reinhardtii. Journal of Biological Chemistry (2006) 281:30186–30194.
Forde BG. Nitrate transporters in plants: structure, function and regulation. Biochimica et Biophysica Acta (2000) 1465:219–235.[Medline]
Franco AR, Cardenas J, Fernandez E. Ammonium (methylammonium) is the co-repressor of nitrate reductase in Chlamydomonas reinhardtii. FEBS Letters (1984) 176:453–456.[CrossRef][Web of Science]
Franco AR, Cardenas J, Fernandez E. Two different carriers transport both ammonium and methylammonium in Chlamydomonas reinhardtii. Journal of Biological Chemistry (1988) 263:14039–14043.
Galvan A, Fernandez E. Eukaryotic nitrate and nitrite transporters. Cellular and Molecular Life Sciences (2001) 58:225–233.[CrossRef][Web of Science][Medline]
Galvan A, Mariscal V, Gonzalez-Ballester D, Fernandez E. The green alga Chlamydomonas as a tool to study the nitrate assimilation pathway in plants. In: Model plants and crop improvement—Varshney RK, Koebner RMD, eds. (2006) Boca Raton, FL: CRC Press. 125–158.
Galvan A, Quesada A, Fernandez E. Nitrate and nitrate are transported by different specific transport systems and by a bispecific transporter in Chlamydomonas reinhardtii. Journal of Biological Chemistry (1996) 271:2088–2092.
Galvan A, Rexach J, Mariscal V, Fernandez E. Nitrite transport to the chloroplast in Chlamydomonas reinhardtii: molecular evidence for a regulated process. Journal of Experimental Botany (2002) 53:845–853.
Glass AD, Britto DT, Kaiser BN, et al. The regulation of nitrate and ammonium transport systems in plants. Journal of Experimental Botany (2002) 53:855–864.
Gonzalez-Ballester D, Camargo A, Fernandez E. Ammonium transporter genes in Chlamydomonas: the nitrate-specific regulatory gene Nit2 is involved in Amt1;1 expression. Plant Molecular Biology (2004) 56:863–878.[CrossRef][Web of Science][Medline]
Gonzalez-Ballester D, de Montaigu A, Higuera JJ, Galvan A, Fernandez E. Functional genomics of the regulation of the nitrate assimilation pathway in Chlamydomonas. Plant Physiology (2005) 137:522–533.
Guo FQ, Wang R, Chen M, Crawford NM. The Arabidopsis dual-affinity nitrate transporter gene AtNRT1.1 (CHL1) is activated and functions in nascent organ development during vegetative and reproductive growth. The Plant Cell (2001) 13:1761–1777.
Guo FQ, Young J, Crawford NM. The nitrate transporter AtNRT1.1 (CHL1) functions in stomatal opening and contributes to drought susceptibility in Arabidopsis. The Plant Cell (2003) 15:107–117.
Khademi S, O'Connell J III, Remis J, Robles-Colmenares Y, Miercke LJ, Stroud RM. Mechanism of ammonia transport by Amt/MEP/Rh: structure of AmtB at 1.35 Å. Science (2004) 305:1587–1594.
Kim KS, Feild E, King N, Yaoi T, Kustu S, Inwood W. Spontaneous mutations in the ammonium transport gene AMT4 of Chlamydomonas reinhardtii. Genetics (2005) 170:631–644.
Kim KS, Kustu S, Inwood W. Natural history of transposition in the green alga Chlamydomonas reinhardtii: use of the AMT4 locus as an experimental system. Genetics (2006) 173:2005–2019.
Kindle KL. Nuclear transformation: technology and applications. In: The molecular biology of chloroplasts and mitochondria in Chlamydomonas—Rochaix JD, Goldschmidt-Clermont M, Merchant S, eds. (1998) Dordrecht, The Netherlands: Kluwer Academic Publishers. 41–61.
Krouk G, Tillard P, Gojon A. Regulation of the high-affinity NO3– uptake system by NRT1.1-mediated NO3– demand signaling in Arabidopsis. Plant Physiology (2006) 142:1075–1086.
Li W, Wang Y, Okamoto M, Crawford NM, Siddiqi MY, Glass AD. Dissection of the AtNRT2.1:AtNRT2.2 inducible high-affinity nitrate transporter gene cluster. Plant Physiology (2007) 143:425–433.
Little DY, Rao H, Oliva S, Daniel-Vedele F, Krapp A, Malamy JE. The putative high-affinity nitrate transporter NRT2.1 represses lateral root initiation in response to nutritional cues. Proceedings of the National Academy of Sciences, USA (2005) 102:13693–13698.
Liu KH, Tsay YF. Switching between the two action modes of the dual-affinity nitrate transporter CHL1 by phosphorylation. EMBO Journal (2003) 22:1005–1013.[CrossRef][Web of Science][Medline]
Llamas A, Igeno MI, Galvan A, Fernandez E. Nitrate signalling on the nitrate reductase gene promoter depends directly on the activity of the nitrate transport systems in Chlamydomonas. The Plant Journal (2002) 30:261–271.[CrossRef][Web of Science][Medline]
Mariscal V, Moulin P, Orsel M, Miller AJ, Fernandez E, Galvan A. Differential regulation of the Chlamydomonas Nar1 gene family by carbon and nitrogen. Protist (2006) 157:421–433.[Medline]
Mariscal M, Rexach J, Fernandez E, Galvan A. The plastidic nitrite transporter NAR1.1 improves nitrate use efficiency for growth in Chlamydomonas. Plant, Cell and Environment (2004) 27:1321–1328.[CrossRef]
Marzluf GA. Molecular genetics of sulfur assimilation in filamentous fungi and yeast. Annual Review of Microbiology (1997) 51:73–96.[CrossRef][Web of Science][Medline]
Masclaux-Daubresse C, Reisdorf-Cren M, Pageau K, Lelandais M, Grandjean O, Kronenberger J, Valadier MH, Feraud M, Jouglet T, Suzuki A. Glutamine synthetase–glutamate synthase pathway and glutamate dehydrogenase play distinct roles in the sink–source nitrogen cycle in tobacco. Plant Physiology (2006) 140:444–456.
Miura K, Yamano T, Yoshioka S, et al. Expression profiling-based identification of CO2-responsive genes regulated by CCM1 controlling a carbon-concentrating mechanism in Chlamydomonas reinhardtii. Plant Physiology (2004) 135:1595–1607.
Navarro MT, Guerra E, Fernandez E, Galvan A. Nitrite reductase mutants as an approach to understanding nitrate assimilation in Chlamydomonas reinhardtii. Plant Physiology (2000) 122:283–290.
Okamoto M, Kumar A, Li W, Wang Y, Siddiqi MY, Crawford NM, Glass AD. High-affinity nitrate transport in roots of Arabidopsis depends on expression of the NAR2-like gene AtNRT3.1. Plant Physiology (2006) 140:1036–1046.
Okamoto M, Vidmar JJ, Glass AD. Regulation of NRT1 and NRT2 gene families of Arabidopsis thaliana: responses to nitrate provision. Plant and Cell Physiology (2003) 44:304–317.
Orsel M, Chopin F, Leleu O, Smith SJ, Krapp A, Daniel-Vedele F, Miller AJ. Characterization of a two-component high-affinity nitrate uptake system in Arabidopsis. Physiology and protein–protein interaction. Plant Physiology (2006) 142:1304–1317.
Orsel M, Krapp A, Daniel-Vedele F. Analysis of the NRT2 nitrate transporter family in Arabidopsis. Structure and gene expression. Plant Physiology (2002) 129:886–896.
Pazour GJ, Witman GB. Forward and reverse genetic analysis of microtubule motors in Chlamydomonas. Methods (2000) 22:285–298.[CrossRef][Web of Science][Medline]
Peakman T, Crouzet J, Mayaux JF, Busby S, Mohan S, Harborne N, Wootton J, Nicolson R, Cole J. Nucleotide sequence, organisation and structural analysis of the products of genes in the nirB–cysG region of the Escherichia coli chromosome. European Journal of Biochemistry (1990) 191:315–323.[Web of Science][Medline]
Perez-Alegre M, Dubus A, Fernandez E. REM1, a new type of long terminal repeat retrotransposon in Chlamydomonas reinhardtii. Molecular and Cellular Biology (2005) 25:10628–10638.
Pozuelo M, Merchan F, Macias MI, Beck CF, Galvan A, Fernandez E. The negative effect of nitrate on gametogenesis is independent of nitrate assimilation in Chlamydomonas reinhardtii. Planta (2000) 211:287–292.[CrossRef][Web of Science][Medline]
Prieto R, Dubus A, Galvan A, Fernandez E. Isolation and characterization of two new negative regulatory mutants for nitrate assimilation in Chlamydomonas reinhardtii obtained by insertional mutagenesis. Molecular and General Genetics (1996) 251:461–471.
Quesada A, Fernandez E. Expression of nitrate assimilation related genes in Chlamydomonas reinhardtii. Plant Molecular Biology (1994) 24:185–194.[CrossRef][Web of Science][Medline]
Quesada A, Galvan A, Fernandez E. Identification of nitrate transporter genes in Chlamydomonas reinhardtii. The Plant Journal (1994) 5:407–419.[CrossRef][Web of Science][Medline]
Quesada A, Gomez I, Fernandez E. Clustering of the nitrite reductase gene and a light-regulated gene with nitrate assimilation loci in Chlamydomonas reinhardtii. Planta (1998b) 206:259–265.[CrossRef][Web of Science][Medline]
Quesada A, Gomez I, Fernandez E. Involvement of chloroplast and mitochondria redox valves in nitrate assimilation. Trends in Plant Science (2000) 5:463–464.[CrossRef][Web of Science][Medline]
Quesada A, Hidalgo J, Fernandez E. Three Nrt2 genes are differentially regulated in Chlamydomonas reinhardtii. Molecular and General Genetics (1998a) 258:373–377.[CrossRef][Web of Science]
Quesada A, Krapp A, Trueman LJ, Daniel-Vedele F, Fernandez E, Forde BG, Caboche M. PCR-identification of a Nicotiana plumbaginifolia cDNA homologous to the high-affinity nitrate transporters of the crnA family. Plant Molecular Biology (1997) 34:265–274.[CrossRef][Web of Science][Medline]
Rexach J, Fernandez E, Galvan A. The Chlamydomonas reinhardtii Nar1 gene encodes a chloroplast membrane protein involved in nitrite transport. The Plant Cell (2000) 12:1441–1453.
Rexach J, Llamas A, Fernandez E, Galvan A. The activity of the high-affinity nitrate transport system I (NRT2;1, NAR2) is responsible for the efficient signalling of nitrate assimilation genes in Chlamydomonas reinhardtii. Planta (2002) 215:606–611.[CrossRef][Web of Science][Medline]
Rexach J, Montero B, Fernandez E, Galvan A. Differential regulation of the high affinity nitrite transport systems III and IV in Chlamydomonas reinhardtii. Journal of Biological Chemistry (1999) 274:27801–27806.
Rochaix JD, Goldschmidt-Clermont M, Merchant S. The molecular biology of chloroplasts and mitochondria in Chlamydomonas (1998) Dordrecht, The Netherlands: Kluwer Academic Publishers.
Scheible WR, Gonzalez-Fontes A, Lauerer M, Muller-Rober B, Caboche M, Stitt M. Nitrate acts as a signal to induce organic acid metabolism and represses starch metabolism in tobacco. The Plant Cell (1997a) 9:783–798.[Abstract]
Scheible WR, Gonzalez-Fontes A, Morcuende R, Lauerer M, Geiger M, Glaab J, Gojon A, Schulze ED, Stitt M. Tobacco mutants with a decreased number of functional Nia genes compensate by modifying the diurnal regulation of transcription, post-translational modification and turnover of nitrate reductase. Planta (1997b) 203:304–319.[CrossRef][Web of Science][Medline]
Schnell RA, Lefebvre PA. Isolation of the Chlamydomonas regulatory gene NIT2 by transposon tagging. Genetics (1993) 134:737–747.[Abstract]
Soupene E, Inwood W, Kustu S. Lack of the Rhesus protein Rh1 impairs growth of the green alga Chlamydomonas reinhardtii at high CO2. Proceedings of the National Academy of Sciences, USA (2004) 101:7787–7792.
Soupene E, King N, Feild E, Liu P, Niyogi KK, Huang CH, Kustu S. Rhesus expression in a green alga is regulated by CO2. Proceedings of the National Academy of Sciences, USA (2002) 99:7769–7773.
Suenaga A, Moriya K, Sonoda Y, Ikeda A, Von Wiren, Hayakawa T, Yamaguchi J, Yamaya T. Constitutive expression of a novel-type ammonium transporter OsAMT2 in rice plants. Plant and Cell Physiology (2003) 44:206–211.
Suppmann B, Sawers G. Isolation and characterization of hypophosphite resistant mutants of Escherichia coli: identification of the FocA protein, encoded by the pfl operon, as a putative formate transporter. Molecular Microbiology (1994) 11:965–998.[Web of Science][Medline]
Tong Y, Zhou JJ, Li Z, Miller AJ. A two-component high-affinity nitrate uptake system in barley. The Plant Journal (2005) 41:442–450.[Web of Science][Medline]
Unkles SE, Hawker KL, Grieve C, Campbell EI, Montague P, Kinghorn JR. CrnA encodes a nitrate transporter in Aspergillus nidulans. Proceedings of the National Academy of Sciences, USA (1991) 88:204–208.
Vidmar JJ, Zhuo D, Siddiqi MY, Schjoerring JK, Touraine B, Glass AD. Regulation of high-affinity nitrate transporter genes and high-affinity nitrate influx by nitrogen pools in roots of barley. Plant Physiology (2000) 123:307–318.
Walch-Liu P, Ivanov II, Filleur S, Gan Y, Remans T, Forde BG. Nitrogen regulation of root branching. Annals of Botany (2006) 97:875–881.
Wang R, Liu D, Crawford NM. The Arabidopsis CHL1 protein plays a major role in high-affinity nitrate uptake. Proceedings of the National Academy of Sciences, USA (1998) 95:15134–15139.
Williams L, Miller A. Transporters responsible for the uptake and partitioning of nitrogenous solutes. Annual Review of Plant Physiology and Plant Molecular Biology (2001) 52:659–688.[CrossRef][Web of Science][Medline]
Zhang D, Lefebvre PA. FAR1, a negative regulatory locus required for the repression of the nitrate reductase gene in Chlamydomonas reinhardtii. Genetics (1997) 146:121–133.[Abstract]
Zhang H, Forde BG. Regulation of Arabidopsis root development by nitrate availability. Journal of Experimental Botany (2000) 51:51–59.
Zhou JJ, Fernandez E, Galvan A, Miller AJ. A high affinity nitrate transport system from Chlamydomonas requires two gene products. FEBS Letters (2000a) 466:225–227.[CrossRef][Web of Science][Medline]
Zhou JJ, Trueman LJ, Boorer KJ, Theodoulou FL, Forde BG, Miller AJ. A high affinity fungal nitrate carrier with two transport mechanisms. Journal of Biological Chemistry (2000b) 275:39894–39899.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  F. J. Navarro, Y. Martin, and J. M. Siverio Phosphorylation of the Yeast Nitrate Transporter Ynt1 Is Essential for Delivery to the Plasma Membrane during Nitrogen Limitation J. Biol. Chem., November 7, 2008; 283(45): 31208 - 31217. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Fernandez and A. Galvan Nitrate Assimilation in Chlamydomonas Eukaryot. Cell, April 1, 2008; 7(4): 555 - 559. [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||