Distinct Roles of Nitrate and Nitrite in Regulation of Expression of the Nitrate Transport Genes in the Moss Physcomitrella patens

Abstract

Five NRT2 genes and three Nar2 genes, encoding putative high-affinity nitrate transporters, and the respective cDNAs were identified and characterized in Physcomitrella patens. The deduced moss NRT2 and NAR2 proteins were more similar to the corresponding proteins of higher plants than to those of the green alga Chlamydomonas reinhardtii. Expression of all the genes was inhibited by ammonium added to the medium. The regulation by ammonium was abolished by an inhibitor of glutamine synthetase, but the effect of this inhibitor was counteracted by an inhibitor of glutamate synthase. Negative correlation was observed between the glutamine content of protonemata and the transcript levels of PpNRT2 and PpNar2. These results indicated that glutamine is the signal for repression of the genes. All the genes except PpNRT2;5 showed transient expression stimulated by nitrate but not by nitrite, peaking at 2-4 h after the medium was deprived of ammonium. When the glutamine synthetase inhibitor was used to inhibit assimilation of the ammonium generated intracellularly from nitrate or nitrite, the second phase of activation of genes became manifest at $8 h after the medium was deprived of ammonium. Surprisingly, both nitrate and nitrite stimulated gene expression at this stage. PpNRT2;5 was distinct from the other genes in that its expression is sharply induced by nitrite, is strictly dependent on nitrite or nitrate, and is much less susceptible to the feedback regulation, retaining a constant level in nitratecontaining medium. These results indicated that P. patens has multiple mechanisms for sensing nitrate and nitrite.

Nitrite Acts as a Transcriptome Signal at Micromolar Concentrations in Arabidopsis Roots

“…In Arabidopsis, the nonsymbiotic hemoglobin gene AtGLB1 is strongly induced after 2 h of 5 mM KNO 2 treatment, which is similar to the response observed with 5 mM KNO 3 (Sakamoto et al, 2004). In Physcomitrella, the NRT gene NRT2;5 is induced by 1 mM nitrite after 4 h, whereas NRT2;1 and NRT2;2 are strongly repressed (Tsujimoto et al, 2007). In Arabidopsis roots, 0.5 to 1.0 mM KNO 2 repressed NRT1.1 and NIA1 mRNA levels after 6 and 24 h of treatment (Loque et al, 2003).…”

Nitrate Assimilation inChlamydomonas

“…Some NRT2 proteins require a second component named NAR2. The requirement for the NAR2 protein for the functionality of high-affinity nitrate transporters (HANT) has been corroborated not only with Chlamydomonas (69) but also with other systems such as the moss Physcomitrella patens (64) and plants (39,41,62). The functionality of NRT2 proteins has been shown only for some of them, as follows: NRT2.1/NAR2 as a bispecific HANNiT, NRT2.2/NAR2 as a HANT, and NRT2.3 as a high-affinity nitrite transporter (14,(47)(48)(49) (Fig.…”

“…The NRT2 genes that encode highaffinity nitrate/nitrite transporters (HANNiT) have also been cloned from mosses, fungi, algae, yeast, and bacteria, and in almost all plant species, they appear as a multigene family (19,20,40,64). Some NRT2 proteins require a second component named NAR2.…”

Two-component high-affinity nitrate transport system in barley: Membrane localization, protein expression in roots and a direct protein-protein interaction

“…Many studies of the molecular basis of nitrate uptake reveal the existence of two gene families, namely the NRT1 and NRT2 families, which potentially encode for LATS and HATS respectively. NRT2 genes are identified in a variety of organisms including fungi, certain yeasts, green algae, and higher plants ( Unkles et al 1991;Quesada et al 1994;Trueman et al 1996;Pérez et al 1997;Quesada et al 1997;Amarasinghe et al 1998;Zhuo et al, 1999;Araki and Hasegawa 2006;Tsujimoto et al 2007). In most species, NRT2 genes are members of a multigene family: for example, seven Arabidopsis genes (AtNRT2.1-AtNRT2.7) and four rice genes (OsNRT2.1-OsNRT2.4) have been found in their genomes (Orsel et al 2002; Araki and Hasegawa 2007), and at least four NRT2 genes (HvNRT2.1-HvNRT2.4) exist in barley (Vidmar 2000a).…”