The role of nitrate reductase in the regulation of the nitrate assimilation pathway in the yeast

Navarro, Francisco J.; Perdomo, Germán; Tejera, Paula; Médina, B.; Machín, Félix; Guillén, Rosa Maria; Lancha, Ana; Siverio, José M.

doi:10.1016/s1567-1356(03)00163-6

Cited by 31 publications

(26 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The role of Npr1 in phosphorylation and cellular localization of Ynt1 raises the question of whether TOR kinases are involved in its post-translational regulation. However, the TOR kinases-inhibitor rapamycin, which mimics the nitrogen deprivation conditions, did not alter either the phosphorylation or Ynt1 stability (results not shown), although the same dose of rapamycin allows the expression of the nitrate reductase gene under repression conditions, such as in glutamine (9). The absence of any effect of rapamycin on Ynt1 regulation is somewhat surprising, considering that in S. cerevisiae Npr1 regulation is under the control of TOR (22) and its role in the phosphorylation of Ynt1, as shown in this work.…”

Section: Discussionmentioning

confidence: 95%

“…These findings led us to think that the Ynt1 regulatory mechanism could be involved in the response to nitrate under very limited nitrogen conditions. We have previously shown in H. polymorpha that nitrate must be transported into the cell to induce the nitrate assimilatory genes, which indicates that nitrate transport systems play an essential role in nitrate assimilation regulation (9). To monitor the induction of these genes when Ynt1 phosphorylation is altered, nitrate reductase gene (YNR1) promoter fused to lacZ was used as reporter.…”

Section: Degradation Of Ynt1mentioning

confidence: 99%

“…In plant, algae, fungi, and yeast, nitrate acts as inducer once it enters the cell (8,9). These facts entail that nitrate transporters play a key role in regulating the influx of nitrate, which is the substrate and the gene inducer of the nitrate assimilation pathway.…”

mentioning

confidence: 99%

“…Post-translational regulation of nitrate transporters in eukaryotes remains somewhat unknown. This kind of regulation would allow fine adjustments of nitrate influx to the cell and, indirectly, the induction of genes involved in nitrate assimilation, at least in those organisms where nitrate is sensed intracellularly, such as H. polymorpha (8,9). Current knowledge about post-transcriptional regulation of nitrate transport derives from Arabidopsis thaliana NRT1;1 (formerly designated CHL1) and the H. polymorpha Ynt1, which represent respectively, two different families of nitrate transporters and different modes of regulation.…”

mentioning

confidence: 99%

See 3 more Smart Citations

Phosphorylation of the Yeast Nitrate Transporter Ynt1 Is Essential for Delivery to the Plasma Membrane during Nitrogen Limitation

Navarro

Martín

Siverio

2008

Journal of Biological Chemistry

View full text Add to dashboard Cite

Ynt1 is the sole high affinity nitrate transporter of the yeast Hansenula polymorpha. It is highly regulated by the nitrogen source, by being down-regulated in response to glutamine by repression of the YNT1 gene and Ynt1 ubiquitinylation, endocytosis, and vacuolar degradation. On the contrary, we show that nitrogen limitation stabilizes Ynt1 levels at the plasma membrane, requiring phosphorylation of the transporter. We determined that Ser-246 in the central intracellular loop plays a key role in the phosphorylation of Ynt1 and that the nitrogen permease reactivator 1 kinase (Npr1) is necessary for Ynt1 phosphorylation. Abolition of phosphorylation led Ynt1 to the vacuole by a pep12-dependent end4-independent pathway, which is also dependent on ubiquitinylation, whereas Ynt1 protein lacking ubiquitinylation sites does not follow this pathway. We found that, under nitrogen limitation, Ynt1 phosphorylation is essential for rapid induction of nitrate assimilation genes. Our results suggest that, under nitrogen limitation, phosphorylation prevents Ynt1 delivery from the secretion route to the vacuole, which, aided by reduced ubiquitinylation, accumulates Ynt1 at the plasma membrane. This mechanism could be part of the response that allows nitrate-assimilatory organisms to cope with nitrogen depletion.

show abstract

Section: Discussionmentioning

confidence: 95%

Section: Degradation Of Ynt1mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Phosphorylation of the Yeast Nitrate Transporter Ynt1 Is Essential for Delivery to the Plasma Membrane during Nitrogen Limitation

Navarro

Martín

Siverio

2008

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The presence or absence of nitrate did not affect these results. 6 Any modification of this procedure was indicated for each experiment. Whenever necessary, media were supplemented with 30 g/ml L-leucine, 20 g/ml uracil, or 100 g/ml Zeocin.…”

Section: Methodsmentioning

confidence: 99%

Npr1 Ser/Thr Protein Kinase Links Nitrogen Source Quality and Carbon Availability with the Yeast Nitrate Transporter (Ynt1) Levels

Martín¹,

González²,

Cabrera³

et al. 2011

Journal of Biological Chemistry

View full text Add to dashboard Cite

Ynt1, the single high affinity nitrate and nitrite transporter of the yeast Hansenula polymorpha, is regulated by the quality of nitrogen sources. Preferred nitrogen sources cause Ynt1 dephosphorylation, ubiquitinylation, endocytosis, and vacuolar degradation. In contrast, under nitrogen limitation Ynt1 is phosphorylated and sorted to the plasma membrane. We show here the involvement of the Ser/Thr kinase HpNpr1 in Ynt1 phosphorylation and regulation of Ynt1 levels in response to nitrogen source quality and the availability of carbon. In ⌬npr1, Ynt1 phosphorylation does not take place, although Ynt1 ubiquitin conjugates increase. As a result, in this strain Ynt1 is sorted to the vacuole, from both plasma membrane and the later biosynthetic pathway in nitrogen-free conditions and nitrate. In contrast, overexpression of NPR1 blocks down-regulation of Ynt1, increasing Ynt1 phosphorylation at Ser-244 and -246 and reducing ubiquitinylation. Furthermore, Npr1 is phosphorylated in response to the preferred nitrogen sources, and indeed it is dephosphorylated in nitrogen-free medium. Under conditions where Npr1 is phosphorylated, Ynt1 is not and vice versa. We show for the first time that carbon starvation leads to Npr1 phosphorylation, whereas Ynt1 is dephosphorylated and degraded in the vacuole. Rapamycin prevents this, indicating a possible role of the target of rapamycin signaling pathway in this process. We concluded that Npr1 plays a key role in adapting Ynt1 levels to the nitrogen quality and availability of a source of carbon.

show abstract

The role of Ynt1 in nitrate and nitrite transport in the yeast Hansenula polymorpha

Machín

Médina

Navarro

et al. 2004

Yeast

View full text Add to dashboard Cite

Ynt1 is the only high-affinity nitrate uptake system in Hansenula polymorpha. Nitrate uptake was directly correlated with the Ynt1 levels and shown to be independent of nitrate reductase (NR) activity levels. Ynt1 failed to transport chlorate and, as a result, strains lacking YNT1 were sensitive to chlorate, as is the wild-type. Nitrite uptake in a wild-type strain was partially inhibited by nitrate to levels shown by a YNT1-disrupted strain in which, in turn, nitrite transport was not inhibited by nitrate. It is concluded that nitrite uptake takes place by two different transport systems: Ynt1 and a nitrite-specific transporter(s). The nitrite-specific transport system was induced by nitrate; consistently, no induction was observed in strains lacking the transcription factor YNA1, which is involved in nitrate and nitrite induction of the nitrate assimilatory structural genes. Ynt1 presents its optimal rate for nitrite uptake at pH 6, while pH 4 was optimal for the specific nitrite uptake system(s). At pH 5.5, the contribution of Ynt1 to high-affinity nitrate and nitrite uptake was around 95% and 60%, respectively. The apparent K m of Ynt1 for nitrate and nitrite is in the µM range, as is the specific nitrite uptake system for nitrite. The analysis of the effect of the reduced nitrogen sources on nitrate assimilation revealed that glutamine inactivates nitrate and nitrite transport, dependent on Ynt1, but not the nitrite-specific system.

show abstract

The role of nitrate reductase in the regulation of the nitrate assimilation pathway in the yeast

Cited by 31 publications

References 42 publications

Phosphorylation of the Yeast Nitrate Transporter Ynt1 Is Essential for Delivery to the Plasma Membrane during Nitrogen Limitation

Phosphorylation of the Yeast Nitrate Transporter Ynt1 Is Essential for Delivery to the Plasma Membrane during Nitrogen Limitation

Npr1 Ser/Thr Protein Kinase Links Nitrogen Source Quality and Carbon Availability with the Yeast Nitrate Transporter (Ynt1) Levels

The role of Ynt1 in nitrate and nitrite transport in the yeast Hansenula polymorpha

Contact Info

Product

Resources

About