Phosphatidylserine is involved in the ferrichrome-induced plasma membrane trafficking of Arn1 in Saccharomyces cerevisiae.
ABSTRACT Arn1 is an integral membrane protein that mediates the uptake of ferrichrome, an important nutritional source of iron, in Saccharomyces cerevisiae. In the absence of ferrichrome, Arn1p is sorted directly from the trans-Golgi network to the vacuolar lumen for degradation. In the presence of low levels of ferrichrome, the siderophore binds to a receptor domain on Arn1, triggering the redistribution of Arn1 to the plasma membrane. When extracellular ferrichrome levels are high, Arn1 cycles between the plasma membrane and intracellular vesicles. To further understand the mechanisms of trafficking of Arn1p, we screened 4580 viable yeast deletion mutants for mislocalization of Arn1-GFP using synthetic genetic array technology. We identified over 100 genes required for trans-Golgi network-to-vacuole trafficking of Arn1-GFP and only two genes, SER1 and SER2, required for the ferrichrome-induced plasma membrane trafficking of Arn1-GFP. SER1 and SER2 encode two enzymes of the major serine biosynthetic pathway, and the Arn1 trafficking defect in the ser1Δ strain was corrected with supplemental serine or glycine. Plasma membrane trafficking of Hxt3, a structurally related glucose transporter, was unaffected by SER1 deletion. Serine is required for the synthesis of multiple cellular components, including purines, sphingolipids, and phospholipids, but of these only phosphatidylserine corrected the Arn1 trafficking defects of the ser1Δ strain. Strains with defects in phospholipid synthesis also exhibited alterations in Arn1p trafficking, indicating that the intracellular trafficking of some transporters is dependent on the phospholipid composition of the cellular membranes.
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ABSTRACT: The trans-Golgi network (TGN) is an important cargo sorting station within the cell where newly synthesized proteins are packaged into distinct transport carriers that are targeted to various destinations. To maintain the fidelity of protein transport, elaborate protein sorting machinery is employed to mediate sorting of specific cargo proteins into distinct transport carriers. Protein sorting requires assembly of the cytosolic sorting machinery onto the TGN membrane and capture of cargo proteins. We review the cytosolic and transmembrane sorting machinery that function at the TGN and describe molecular interactions and regulatory mechanisms that enable accurate protein sorting. In addition, we highlight the importance of TGN sorting in physiology and disease. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 30 is October 06, 2014. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.Annual Review of Cell and Developmental Biology 08/2014; DOI:10.1146/annurev-cellbio-100913-013012 · 20.24 Impact Factor
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ABSTRACT: Sac1 is a phosphoinositide phosphatase, which preferentially dephosphorylates phosphatidylinositol 4-phosphate. Mutation of SAC1 causes not only accumulation of phosphoinositides, but also reduction of the phosphatidylserine (PS) level in yeast Saccharomyces cerevisiae. In this study, we characterized the mechanism underlying the PS reduction in SAC1-deleted cells. Incorporation of (32) P into PS was significantly delayed in sac1∆ cells. Such delay was also observed in SAC1- and PS decarboxylase gene-deleted cells, suggesting that the reduction in the PS level is caused by reduction in the rate of biosynthesis of PS. A reduction in the PS level was also observed with repression of STT4 encoding phosphatidylinositol 4-kinase or deletion of VPS34 encoding phophatidylinositol 3-kinase. However, the combination of mutations of SAC1 and STT4 or VPS34 did not restore the reduced PS level, suggesting that both the synthesis and degradation of phosphoinositides are important for maintenance of the PS level. Finally, we observed an abnormal PS distribution in sac1∆ cells when a specific probe for PS was expressed. Collectively, these results suggested that Sac1 is involved in maintenance of a normal rate of biosynthesis and distribution of PS. This article is protected by copyright. All rights reserved.Yeast 04/2014; 31(4). DOI:10.1002/yea.3004 · 1.74 Impact Factor
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ABSTRACT: Di- and tripeptide transporters of the PTR/NRT1 (peptide transporter/nitrate transporter1)-family are localized either at the tonoplast (TP) or plasma membrane (PM). As limited information is available on structural determinants required for targeting of plant membrane proteins, we performed gene shuffling and domain swapping experiments of Arabidopsis PTRs. A 7 amino acid fragment of the hydrophilic N-terminal region of PTR2, PTR4 and PTR6 was required for TP localization and sufficient to redirect not only PM-localized PTR1 or PTR5, but also sucrose transporter SUC2 to the TP. Alanine scanning mutagenesis identified L(11) and I(12) of PTR2 to be essential for TP targeting, while only one acidic amino acid at position 5, 6 or 7 was required, revealing a dileucine (LL or LI) motif with at least one upstream acidic residue. Similar dileucine motifs could be identified in other plant TP transporters, indicating a broader role of this targeting motif in plants. Targeting to the PM required the loop between transmembrane domain 6 and 7 of PTR1 or PTR5. Deletion of either PM or TP targeting signals resulted in retention in internal membranes, indicating that PTR trafficking to these destination membranes requires distinct signals and is in both cases not by default.Traffic 04/2012; 13(8):1090-105. DOI:10.1111/j.1600-0854.2012.01370.x · 4.71 Impact Factor