Direct Sorting of the Yeast Uracil Permease to the Endosomal System Is Controlled by Uracil Binding and Rsp5p-dependent Ubiquitylation

Blondel, Marie-Odile; Morvan, Joëlle; Dupré-Crochet, Sophie; Urban‐Grimal, Danièle; Haguenauer‐Tsapis, Rosine; Volland, Christiane

doi:10.1091/mbc.e03-04-0202

Cited by 114 publications

(173 citation statements)

References 62 publications

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“…How differential ubiquitination influences the dynamic of IRT1 along the endocytic pathway remains an open question. Although single monoubiquitination appears sufficient for internalization of plasma membrane proteins (10)(11)(12), multiple monoubiquitinations of IRT1 may increase the rate of such a process, as observed for the Fur4 yeast uracil permease (33,45). The presence of pools of IRT1 decorated with different numbers of monoubiquitination residues may also reflect a hierarchy in ubiquitinated residues.…”

Section: Discussionmentioning

confidence: 99%

Monoubiquitin-dependent endocytosis of the IRON-REGULATED TRANSPORTER 1 (IRT1) transporter controls iron uptake in plants

Barberon

Zelazny

Robert

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

416

430

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Plants take up iron from the soil using the IRON-REGULATED TRANSPORTER 1 (IRT1) high-affinity iron transporter at the root surface. Sophisticated regulatory mechanisms allow plants to tightly control the levels of IRT1, ensuring optimal absorption of essential but toxic iron. Here, we demonstrate that overexpression of Arabidopsis thaliana IRT1 leads to constitutive IRT1 protein accumulation, metal overload, and oxidative stress. IRT1 is unexpectedly found in trans-Golgi network/early endosomes of root hair cells, and its levels and localization are unaffected by iron nutrition. Using pharmacological approaches, we show that IRT1 cycles to the plasma membrane to perform iron and metal uptake at the cell surface and is sent to the vacuole for proper turnover. We also prove that IRT1 is monoubiquitinated on several cytosol-exposed residues in vivo and that mutation of two putative monoubiquitination target residues in IRT1 triggers stabilization at the plasma membrane and leads to extreme lethality. Together, these data suggest a model in which monoubiquitin-dependent internalization/sorting and turnover keep the plasma membrane pool of IRT1 low to ensure proper iron uptake and to prevent metal toxicity. More generally, our work demonstrates the existence of monoubiquitin-dependent trafficking to lytic vacuoles in plants and points to proteasome-independent turnover of plasma membrane proteins.ubiquitin | protein dynamic | plant cell biology

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Section: Discussionmentioning

confidence: 99%

Monoubiquitin-dependent endocytosis of the IRON-REGULATED TRANSPORTER 1 (IRT1) transporter controls iron uptake in plants

Barberon

Zelazny

Robert

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

416

430

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“…Among the substrates for Rsp5-mediated ubiquitylation are the copper transporter Ctr1p (Liu et al, 2007), the zinc transporter Zrt1p (Gitan and Eide, 2000), the general amino acid permease Gap1p (Hein et al, 1995), the uracil permease Fur4p (Galan et al, 1996;Blondel et al, 2004), and the tryptophan permease Tat2p (Beck et al, 1999). Rsp5 is encoded by an essential gene, so we investigated the localization of Ftr1-GFP by epifluorescence microscopy in two different rsp5 strains that each contain a point mutation within the Rsp5 catalytic HECT domain (rsp5-1 and rsp5-smm1; Fisk and Yaffe, 1999), and in control cells expressing a wild-type copy of RSP5.…”

Section: Constitutive Endocytic Recycling Of Fet3-ftr1 In Degradativementioning

confidence: 99%

Opposing Activities of the Snx3-Retromer Complex and ESCRT Proteins Mediate Regulated Cargo Sorting at a Common Endosome

Strochlic

Schmiedekamp

Lee

et al. 2008

MBoC

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Endocytosed proteins are either delivered to the lysosome to be degraded or are exported from the endosomal system and delivered to other organelles. Sorting of the Saccharomyces cerevisiae reductive iron transporter, composed of the Fet3 and Ftr1 proteins, in the endosomal system is regulated by available iron; in iron-starved cells, Fet3-Ftr1 is sorted by Snx3/Grd19 and retromer into a recycling pathway that delivers it back to the plasma membrane, but when starved cells are exposed to iron, Fet3-Ftr1 is targeted to the lysosome-like vacuole and is degraded. We report that iron-induced endocytosis of Fet3-Ftr1 is independent of Fet3-Ftr1 ubiquitylation, and after endocytosis, degradation of Fet3-Ftr1 is mediated by the multivesicular body (MVB) sorting pathway. In mutant cells lacking any component of the ESCRT protein-dependent MVB sorting machinery, the Rsp5 ubiquitin ligase, or in wild-type cells expressing Fet3-Ftr1 lacking cytosolic lysyl ubiquitin acceptor sites, Fet3-Ftr1 is constitutively sorted into the recycling pathway independent of iron status. In the presence and absence of iron, Fet3-Ftr1 transits an endosomal compartment where a subunit of the MVB sorting receptor (Vps27), Snx3/Grd19, and retromer proteins colocalize. We propose that this endosome is where Rsp5 ubiquitylates Fet3-Ftr1 and where the recycling and degradative pathways diverge. INTRODUCTIONThe endocytic pathway is initiated with the internalization of protein and lipid cargo from the plasma membrane. After endocytosis, internalized molecules transit through the endosomal system, a network of interconnected organelles that process and sort cargo molecules back to the plasma membrane, to other organelles, or to lysosomes for degradation. In mammalian cells, early endosomes are pleiomorphic organelles composed of vacuolar domains that contain lumenal vesicles and tubular domains that lead into and emanate from the vacuolar portion. This distinct architecture has functional significance. The tubular elements of the organelle are regions where the majority of internalized plasma membrane proteins and lipids are exported and recycled from the endosomal system via bulk flow and cargo-specific processes (Mayor et al., 1993;Maxfield and McGraw, 2004;Bonifacino and Rojas, 2006). Cargo proteins that are not exported are delivered to lysosomes and many of these are degraded. This occurs via an active, signal-mediated event in which these proteins are targeted to the vacuolar region of the endosome where they are incorporated into vesicles that bud from the limiting membrane into the lumen of the organelle (Piper and Katzmann, 2007). During endosome maturation lumenal vesicles accrue until fusion of the late endosome with the lysosome results in degradation of these vesicles and their contents. A central sorting function of the endosomal system is thus to segregate cargo to be exported from the endosome from cargo to be degraded via lysosomal proteolysis.The core components of the recycling (retrograde) and degradative sorting pathways have be...

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“…The uracil permease, Fur4, in the presence of its substrate is able to leave the Golgi apparatus and is sorted to the endosomal system for its degradation. Ubiquitinylation is then required for delivering Fur4 into the vacuolar lumen but is not a dominant signal for Golgi-to-endosome trafficking (32). Some of these permeases are phosphorylated; however, the role of this post-translational modification in direct sorting to the vacuole remains unclear.…”

Section: Discussionmentioning

confidence: 99%

“…Some of these permeases are phosphorylated; however, the role of this post-translational modification in direct sorting to the vacuole remains unclear. For example, Fur4 phosphorylation is required for ubiquitinylating the permease present at the cell surface but is not implicated in vacuolar sorting from the Golgi apparatus (32).…”

Section: Discussionmentioning

confidence: 99%

“…We addressed the question if ubiquitinylation, in addition to dephosphorylation, was also involved in the sorting of Ynt1 from the Golgi apparatus to the vacuole under nitrogen deprivation. In fact, in Saccharomyces cerevisiae, vacuolar sorting directly from the Golgi apparatus of newly synthesized yeast permeases Gap1, Fur4, and Tat2 to the vacuole requires ubiquitinylation (20,32,34). Efficient endocytosis of Ynt1 requires the ubiquitinylation of lysine residues of the central intracellular loop, principally Lys-253 and Lys-270.…”

Section: Under Nitrogen Limitation Ubiquitinylation Is Required To Dmentioning

confidence: 99%

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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

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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.

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Direct Sorting of the Yeast Uracil Permease to the Endosomal System Is Controlled by Uracil Binding and Rsp5p-dependent Ubiquitylation

Cited by 114 publications

References 62 publications

Monoubiquitin-dependent endocytosis of the IRON-REGULATED TRANSPORTER 1 (IRT1) transporter controls iron uptake in plants

Monoubiquitin-dependent endocytosis of the IRON-REGULATED TRANSPORTER 1 (IRT1) transporter controls iron uptake in plants

Opposing Activities of the Snx3-Retromer Complex and ESCRT Proteins Mediate Regulated Cargo Sorting at a Common Endosome

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

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