Jana Patton-Vogt

Duquesne University, Pittsburgh, Pennsylvania, United States

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Publications (19)78.42 Total impact

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    ABSTRACT: Candida albicans contains 4 ORFs (GIT1,2,3,4) predicted to encode proteins involved in the transport of glycerophosphodiester metabolites. Previously we reported that Git1, encoded by ORF 19.34, is responsible for the transport of intact glycerophosphoinositol (GroPIns), but not glycerophosphocholine (GroPCho). Here, we report that a strain lacking both GIT3 (ORF 19.1979) and GIT4 (ORF 19.1980) is unable to transport [(3)H]GroPCho into the cell. In the absence of a GroPCho transporter, C. albicans can utilize GroPCho via a mechanism involving extracellular hydrolysis. Upon reintegration of either GIT3 or GIT4 into the genome, measurable uptake of [(3)H]GroPCho is observed. Transport assays and kinetic analyses indicate that Git3 has the greater transport velocity. We present evidence that GDE1 (ORF 19.3936) codes for an enzyme with glycerophosphodiesterase activity against GroPCho. Homozygous deletion of GDE1 results in a buildup of internal GroPCho that is restored to wild type levels by reintegration of GDE1 into the genome. The transcriptional regulator, Pho4, is shown to regulate the expression of GIT3, GIT4, and GDE1. Finally, Git3 is shown to be required for full virulence in a mouse model of disseminated candidiasis, and Git3 sequence orthologs are present in other pathogenic Candida species. In summary, we have characterized multiple aspects of GroPCho utilization by C. albicans and have demonstrated that GroPCho transport plays a key role in the growth of the organism in the host.
    Journal of Biological Chemistry 10/2013; · 4.65 Impact Factor
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    ABSTRACT: Acetic acid inhibition of yeast fermentation has a negative impact in several industrial processes. As an initial step in the construction of a Saccharomyces cerevisiae strain with increased tolerance for acetic acid, mutations conferring resistance were identified by screening a library of deletion mutants in a multiply auxotrophic genetic background. Of the 23 identified mutations, 11 were then introduced into a prototrophic laboratory strain for further evaluation. Because none of the 11 mutations was found to increase resistance in the prototrophic strain, potential interference by the auxotrophic mutations themselves was investigated. Mutants carrying single auxotrophic mutations were constructed and found to be more sensitive to growth inhibition by acetic acid than an otherwise isogenic prototrophic strain. At a concentration of 80 mM acetic acid at pH 4.8, the initial uptake of uracil, leucine, lysine, histidine, tryptophan, phosphate, and glucose was lower in the prototrophic strain than in a non-acetic acid-treated control. These findings are consistent with two mechanisms by which nutrient uptake may be inhibited. Intracellular adenosine triphosphate (ATP) levels were severely decreased upon acetic acid treatment, which likely slowed ATP-dependent proton symport, the major form of transport in yeast for nutrients other than glucose. In addition, the expression of genes encoding some nutrient transporters was repressed by acetic acid, including HXT1 and HXT3 that encode glucose transporters that operate by facilitated diffusion. These results illustrate how commonly used genetic markers in yeast deletion libraries complicate the effort to isolate strains with increased acetic acid resistance.
    Applied Microbiology and Biotechnology 07/2013; · 3.69 Impact Factor
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    ABSTRACT: A highly sensitive hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method was developed and validated for the quantification of glycerophosphoinositol (GroPIns), glycerophosphocholine (GroPCho), glycerol 3-phosphate (GroP), inositol, and choline in the extracellular medium of Saccharomyces cerevisiae. The media samples were pretreated with a single two-phase liquid extraction. Chromatographic separation was achieved on a Waters Xbridge HILIC (150 mm × 4.6 mm, 5 μm) column under isocratic conditions using a mobile phase composed of acetonitrile/water, 70:30 (v/v) with 10mM ammonium acetate (pH adjusted to 4.5) at a flow-rate of 0.5 mL/min. Using a triple quadrupole tandem mass spectrometer, samples were detected in multiple reaction monitoring (MRM) mode via an electrospray ionization (ESI) source. The calibration curves were linear (r² ≥ 0.995) over the range of 0.5-150 nM, with the lower limit of quantitation validated at 0.5 nM for all analytes. The intra- and inter-day precision (calculated by coefficient of variation, CV%) ranged from 1.24 to 5.88% and 2.46 to 9.77%, respectively, and intra- and inter-day accuracy (calculated by relative error, RE%) was between -8.42 to 8.22% and -9.35 to 6.62%, respectively, at all quality control levels. The extracellular metabolites were stable throughout various storage stability studies. The fully validated method was successfully applied to determine the extracellular levels of phospholipid-related metabolites in S. cerevisiae.
    Journal of chromatography. B, Analytical technologies in the biomedical and life sciences 04/2012; 897:1-9. · 2.78 Impact Factor
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    ABSTRACT: Glycerophosphodiesters are the products of phospholipase-mediated deacylation of phospholipids. In Saccharomyces cerevisiae, a single gene, GIT1, encodes a permease responsible for importing glycerophosphodiesters, such as glycerophosphoinositol and glycerophosphocholine, into the cell. In contrast, the Candida albicans genome contains four open reading frames (ORFs) with a high degree of similarity to S. cerevisiae GIT1 (ScGIT1) Here, we report that C. albicans utilizes glycerophosphoinositol (GroPIns) and glycerophosphocholine (GroPCho) as sources of phosphate at both mildly acidic and physiological pHs. Insertional mutagenesis of C. albicans GIT1 (CaGIT1) (orf19.34), the ORF most similar to ScGit1, abolished the ability of cells to use GroPIns as a phosphate source at acidic pH and to transport [(3)H]GroPIns at acidic and physiological pHs, while reintegration of a GIT1 allele into the genome restored those functions. Several lines of evidence, including the detection of internal [(3)H]GroPIns, indicated that GroPIns is transported intact through CaGit1. GroPIns transport was shown to conform to Michaelis-Menten kinetics, with an apparent K(m) of 28 ± 6 μM. Notably, uptake of label from [(3)H]GroPCho was found to be roughly 50-fold greater than uptake of label from [(3)H]GroPIns and roughly 500-fold greater than the equivalent activity in S. cerevisiae. Insertional mutagenesis of CaGIT1 had no effect on the utilization of GroPCho as a phosphate source or on the uptake of label from [(3)H]GroPCho. Growth under low-phosphate conditions was shown to increase label uptake from both [(3)H]GroPIns and [(3)H]GroPCho. Screening of a transcription factor deletion set identified CaPHO4 as required for the utilization of GroPIns, but not GroPCho, as a phosphate source.
    Eukaryotic Cell 12/2011; 10(12):1618-27. · 3.59 Impact Factor
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    ABSTRACT: Biofilms of Candida albicans include both yeast cells and hyphae. Prior studies indicated that a zap1Δ/Δ mutant, defective in zinc regulator Zap1, has increased accumulation of yeast cells in biofilms. This altered yeast-hypha balance may arise from internal regulatory alterations or from an effect on the production of diffusible quorum-sensing (QS) molecules. Here, we develop biosensor reporter strains that express yeast-specific YWP1-RFP or hypha-specific HWP1-RFP, along with a constitutive TDH3-GFP normalization standard. Seeding these biosensor strains into biofilms allows a biological activity assay of the surrounding biofilm milieu. A zap1Δ/Δ biofilm induces the yeast-specific YWP1-RFP reporter in a wild-type biosensor strain, as determined by both quantitative reverse transcription-PCR (qRT-PCR) gene expression measurements and confocal microscopy. Remediation of the zap1Δ/Δ zinc uptake defect through zinc transporter gene ZRT2 overexpression reverses induction of the yeast-specific YWP1-RFP reporter. Gas chromatography-mass spectrometry (GC-MS) measurements of known organic QS molecules show that the zap1Δ/Δ mutant accumulates significantly less farnesol than wild-type or complemented strains and that ZRT2 overexpression does not affect farnesol accumulation. Farnesol is a well-characterized inhibitor of hypha formation; hence, a reduction in farnesol levels in zap1Δ/Δ biofilms is unexpected. Our findings argue that a Zap1- and zinc-dependent signal affects the yeast-hypha balance and that it is operative in the low-farnesol environment of the zap1Δ/Δ biofilm. In addition, our results indicate that Zap1 is a positive regulator of farnesol accumulation.
    Eukaryotic Cell 09/2011; 10(11):1448-54. · 3.59 Impact Factor
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    ABSTRACT: White lupin (Lupinus albus) is a legume that is very efficient in accessing unavailable phosphorus (Pi). It develops short, densely clustered tertiary lateral roots (cluster/proteoid roots) in response to Pi limitation. In this report, we characterize two glycerophosphodiester phosphodiesterase (GPX-PDE) genes (GPX-PDE1 and GPX-PDE2) from white lupin and propose a role for these two GPX-PDEs in root hair growth and development and in a Pi stress-induced phospholipid degradation pathway in cluster roots. Both GPX-PDE1 and GPX-PDE2 are highly expressed in Pi-deficient cluster roots, particularly in root hairs, epidermal cells, and vascular bundles. Expression of both genes is a function of both Pi availability and photosynthate. GPX-PDE1 Pi deficiency-induced expression is attenuated as photosynthate is deprived, while that of GPX-PDE2 is strikingly enhanced. Yeast complementation assays and in vitro enzyme assays revealed that GPX-PDE1 shows catalytic activity with glycerophosphocholine while GPX-PDE2 shows highest activity with glycerophosphoinositol. Cell-free protein extracts from Pi-deficient cluster roots display GPX-PDE enzyme activity for both glycerophosphocholine and glycerophosphoinositol. Knockdown of expression of GPX-PDE through RNA interference resulted in impaired root hair development and density. We propose that white lupin GPX-PDE1 and GPX-PDE2 are involved in the acclimation to Pi limitation by enhancing glycerophosphodiester degradation and mediating root hair development.
    Plant physiology 04/2011; 156(3):1131-48. · 6.56 Impact Factor
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    ABSTRACT: Saccharomyces cerevisiae produces extracellular glycerophosphoinositol through phospholipase-mediated turnover of phosphatidylinositol and transports glycerophosphoinositol into the cell upon nutrient limitation. A screening identified the RAS GTPase-activating proteins Ira1 and Ira2 as required for utilization of glycerophosphoinositol as the sole phosphate source, but the RAS/cyclic AMP pathway does not appear to be involved in the growth phenotype. Ira1 and Ira2 affect both the production and transport of glycerophosphoinositol.
    Eukaryotic Cell 09/2009; 8(11):1808-11. · 3.59 Impact Factor
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    ABSTRACT: To investigate the contributions of phosphatidylethanolamine to the growth and morphogenesis of the fission yeast Schizosaccharomyces pombe, we have characterized three predicted genes in this organism, designated psd1, psd2, and psd3, encoding phosphatidylserine decarboxylases, which catalyze the conversion of phosphatidylserine to phosphatidylethanolamine in both eukaryotic and prokaryotic organisms. S. pombe mutants carrying deletions in any one or two psd genes are viable in complex rich medium and synthetic defined minimal medium. However, mutants carrying deletions in all three psd genes (psd1-3Delta mutants) grow slowly in rich medium and are inviable in minimal medium, indicating that the psd1 to psd3 gene products share overlapping essential cellular functions. Supplementation of growth media with ethanolamine, which can be converted to phosphatidylethanolamine by the Kennedy pathway, restores growth to psd1-3Delta cells in minimal medium, indicating that phosphatidylethanolamine is essential for S. pombe cell growth. psd1-3Delta cells produce lower levels of phosphatidylethanolamine than wild-type cells, even in medium supplemented with ethanolamine, indicating that the Kennedy pathway can only partially compensate for the loss of phosphatidylserine decarboxylase activity in S. pombe. psd1-3Delta cells appear morphologically indistinguishable from wild-type S. pombe cells in medium supplemented with ethanolamine, but when cultured in nonsupplemented medium, they produce high frequencies of abnormally shaped cells as well as cells exhibiting severe septation defects, including multiple, mispositioned, deformed, and misoriented septa. Our results demonstrate that phosphatidylethanolamine is essential for cell growth and for normal cytokinesis and cellular morphogenesis in S. pombe, and they illustrate the usefulness of this model eukaryote for investigating potentially conserved biological and molecular functions of phosphatidylethanolamine.
    Eukaryotic Cell 04/2009; 8(5):790-9. · 3.59 Impact Factor
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    ABSTRACT: The highly conserved yeast cell wall integrity mitogen-activated protein kinase pathway regulates cellular responses to cell wall and membrane stress. We report that this pathway is activated and essential for viability under growth conditions that alter both the abundance and pattern of synthesis and turnover of membrane phospholipids, particularly phosphatidylinositol and phosphatidylcholine. Mutants defective in this pathway exhibit a choline-sensitive inositol auxotrophy, yet fully derepress INO1 and other Opi1p-regulated genes when grown in the absence of inositol. Under these growth conditions, Mpk1p is transiently activated by phosphorylation and stimulates the transcription of known targets of Mpk1p signaling, including genes regulated by the Rlm1p transcription factor. mpk1Delta cells also exhibit severe defects in lipid metabolism, including an abnormal accumulation of phosphatidylcholine, diacylglycerol, triacylglycerol, and free sterols, as well as aberrant turnover of phosphatidylcholine. Overexpression of the NTE1 phospholipase B gene suppresses the choline-sensitive inositol auxotrophy of mpk1Delta cells, whereas overexpression of other phospholipase genes has no effect on this phenotype. These results indicate that an intact cell wall integrity pathway is required for maintaining proper lipid homeostasis in yeast, especially when cells are grown in the absence of inositol.
    Journal of Biological Chemistry 11/2008; 283(49):34204-17. · 4.65 Impact Factor
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    ABSTRACT: The product of the open reading frame YPL206c, Pgc1p, of the yeast Saccharomyces cerevisiae displays homology to bacterial and mammalian glycerophosphodiester phosphodiesterases. Deletion of PGC1 causes an accumulation of the anionic phospholipid, phosphatidylglycerol (PG), especially under conditions of inositol limitation. This PG accumulation was not caused by increased production of phosphatidyl-glycerol phosphate or by decreased consumption of PG in the formation of cardiolipin, the end product of the pathway. PG accumulation in the pgc1Delta strain was caused rather by inactivation of the PG degradation pathway. Our data demonstrate an existence of a novel regulatory mechanism in the cardiolipin biosynthetic pathway in which Pgc1p is required for the removal of excess PG via a phospholipase C-type degradation mechanism.
    Journal of Biological Chemistry 07/2008; 283(25):17107-15. · 4.65 Impact Factor
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    ABSTRACT: To investigate the contributions of phosphatidylserine to the growth and morphogenesis of the rod-shaped fission yeast Schizosaccharomyces pombe, we have characterized the single gene in this organism, pps1, encoding a predicted phosphatidylserine synthase. S. pombe pps1Delta mutants grow slowly in rich medium and are inviable in synthetic minimal medium. They do not produce detectable phosphatidylserine in vivo and possess negligible in vitro phosphatidylserine synthase activity, indicating that pps1 encodes the major phosphatidylserine synthase activity in S. pombe. Supplementation of growth medium with ethanolamine partially suppresses the growth-defective phenotype of pps1Delta cells, reflecting the likely importance of phosphatidylserine as a precursor for phosphatidylethanolamine in S. pombe. In medium lacking ethanolamine, pps1Delta mutants exhibit striking cell morphology, cytokinesis, actin cytoskeleton, and cell wall remodeling and integrity defects. Overexpression of pps1 likewise leads to defects in cell morphology and cytokinesis, thus implicating phosphatidylserine as a dosage-dependent regulator of these processes. During log-phase growth, green fluorescent protein-Pps1p fusion proteins are concentrated at the cell and nuclear peripheries as well as presumptive endoplasmic reticulum membranes, while in stationary-phase cells, they are redistributed to unusual cytoplasmic structures of unknown origin. Moreover, stationary-phase pps1Delta cultures retain very poor viability relative to wild-type S. pombe cells, even in medium containing ethanolamine, demonstrating a role for phosphatidylserine in the physiological adaptations required for stationary-phase survival. Our findings reveal novel cellular functions for phosphatidylserine and emphasize the usefulness of S. pombe as a model organism for elucidating potentially conserved biological and molecular functions of this phospholipid.
    Eukaryotic Cell 12/2007; 6(11):2092-101. · 3.59 Impact Factor
  • Jana Patton-Vogt
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    ABSTRACT: Phospholipid deacylation results in the formation of glycerophosphodiesters and free fatty acids. In Saccharomyces cerevisiae, four gene products with phospholipase B (deacylating) activity have been characterized (PLB1, PLB2, PLB3, NTE1), and those activities account for most, if not all, of the glycerophosphodiester production observed to date. The glycerophosphodiesters themselves are hydrolyzed into glycerol-3-phosphate and the corresponding alcohol by glycerophosphodiester phosphodiesterases. Although only one glycerophosphodiester phosphodiesterase-encoding gene (GDE1) has been characterized in S. cerevisiae, others certainly exist. Both internal and external glycerophosphodiesters (primarily glycerophosphocholine and glycerophosphoinositol) are formed as a result of phospholipid turnover in S. cerevisiae. A permease encoded by the GIT1 gene imports extracellular glycerophosphodiesters across the plasma membrane, where their hydrolytic products can provide crucial nutrients such as inositol, choline, and phosphate to the cell. The importance of this metabolic pathway in various aspects of S. cerevisiae cell physiology is being explored.
    Biochimica et Biophysica Acta 04/2007; 1771(3):337-42. · 4.66 Impact Factor
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    ABSTRACT: The glycerophosphoinositols are ubiquitous phosphoinositide metabolites involved in the control of several cell functions. They exert their actions both intracellularly and by rapidly equilibrating across the plasma membrane when added to cells, implying the existence of a transporter for their membrane permeation. Such a transporter, GIT1, has been cloned in yeast. By PSI-BLAST analysis, we have identified the Glut2 transporter as a human-genome candidate ortholog of GIT1. This was supported directly through the use of inhibitors, siRNAs and competition studies of specific uptake of GroPIns in HeLa cells over-expressing human Glut2. These data identify Glut2 as a GroPIns transporter in mammals, and define a physiologically relevant cell-permeation mechanism.
    FEBS Letters 01/2007; 580(30):6789-96. · 3.58 Impact Factor
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    Claudia Almaguer, Edward Fisher, Jana Patton-Vogt
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    ABSTRACT: Glycerophosphoinositol (GroPIns) and glycerophosphocholine (GroPCho) are the products of phospholipase-B mediated deacylation of phosphatidylinositol and phosphatidylcholine, respectively. GroPIns and GroPCho are transported across the Saccharomyces cerevisiae plasma membrane into the cell via the transporter encoded by GIT1. Previous studies have shown that GIT1 expression is regulated by inorganic phosphate (P(i)) availability through the transcription factors Pho2p and Pho4p. We now report that posttranscriptional mechanisms also regulate Git1p activity in response to P(i) availability. Mutations that inhibit endocytosis and vacuolar proteolysis inhibit Git1p degradation, indicating that Git1p downregulation involves internalization and subsequent degradation in the vacuole. Similar to the effect seen with P(i), provision of cells with high levels of the Git1p substrates, GroPIns and GroPCho, posttranscriptionally downregulates Git1p activity. Unlike P(i), high levels of GroPCho and GroPIns do not repress GIT1 promoter-driven reporter gene activity. These results indicate that Git1p transport activity is regulated at multiple levels by P(i) availability. In addition, the results indicate that the Git1p substrates (and alternate phosphate sources) GroPIns and GroPCho behave distinctly from P(i) in their ability to affect GIT1 expression.
    Current Genetics 01/2007; 50(6):367-75. · 2.41 Impact Factor
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    ABSTRACT: To study the consequences of depleting the major membrane phospholipid phosphatidylcholine (PC), exponentially growing cells of a yeast cho2opi3 double deletion mutant were transferred from medium containing choline to choline-free medium. Cell growth did not cease until the PC level had dropped below 2% of total phospholipids after four to five generations. Increasing contents of phosphatidylethanolamine (PE) and phosphatidylinositol made up for the loss of PC. During PC depletion, the remaining PC was subject to acyl chain remodeling with monounsaturated species replacing diunsaturated species, as shown by mass spectrometry. The remodeling of PC did not require turnover by the SPO14-encoded phospholipase D. The changes in the PC species profile were found to reflect an overall shift in the cellular acyl chain composition that exhibited a 40% increase in the ratio of C16 over C18 acyl chains, and a 10% increase in the degree of saturation. The shift was stronger in the phospholipid than in the neutral lipid fraction and strongest in the species profile of PE. The shortening and increased saturation of the PE acyl chains were shown to decrease the nonbilayer propensity of PE. The results point to a regulatory mechanism in yeast that maintains intrinsic membrane curvature in an optimal range.
    Molecular Biology of the Cell 03/2006; 17(2):1006-17. · 4.60 Impact Factor
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    ABSTRACT: Glycerophosphocholine is formed via the deacylation of the phospholipid phosphatidylcholine. The protein encoded by Saccharomyces cerevisiae open reading frame YPL110c effects glycerophosphocholine metabolism in vivo, most likely by acting as a glycerophosphocholine phosphodiesterase. Deletion of YPL110c causes an accumulation of glycerophosphocholine in cells prelabeled with [14C]choline. Correspondingly, overexpression of YPL110c results in reduced intracellular glycerophosphocholine in cells prelabeled with [14C]choline. Glycerophospho[3H]choline supplied in the growth medium accumulates to a much greater extent in the intracellular fraction of a YPL110Delta strain than in a wild type strain. Furthermore, glycerophospho[3H]choline accumulation requires the transporter encoded by GIT1, a known glycerophosphoinositol transporter. Growth on glycerophosphocholine as the sole phosphate source requires YPL110c and the Git1p permease. In contrast to glycerophosphocholine, glycerophosphoinositol metabolism is unaffected by deletion of YPL110c. The open reading frame YPL110c has been termed GDE1.
    Journal of Biological Chemistry 11/2005; 280(43):36110-7. · 4.65 Impact Factor
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    ABSTRACT: Git1p mediates the transport of the phospholipid metabolite, glycerophosphoinositol, into Saccharomyces cerevisiae. We report that phosphate limitation and inositol limitation affect GIT1 expression and Git1p transport activity via distinct mechanisms that involve multiple transcription factors. GIT1 transcript levels and Git1p activity are greater in cells starved for phosphate, with or without inositol limitation, than in cells only limited for inositol. Furthermore, the kinetics of GIT1 transcript accumulation and Git1p activity upon transfer of cells to phosphate starvation media are different from those obtained upon transfer of cells to inositol-free media. Pho2p and Pho4p are required for GIT1 expression and for Git1p transport activity under all growth conditions tested. In contrast, Ino2p and Ino4p are required for full GIT1 expression when inositol is limiting, with or without phosphate limitation, but not when only phosphate is limiting. Greatly reduced transport activity was detected in ino2Delta and ino4Delta cells under all growth conditions. A 300-base pair region of the GIT1 promoter containing potential Pho4p binding sites was shown to be required for full GIT1 expression. Git1p appears to act as a H(+)-symporter, and neither inositol nor phosphate effectively compete with glycerophosphoinositol for transport by Git1p. Glycerophosphoinositol was shown previously to support the growth of an inositol auxotroph. Remarkably, we now report that glycerophosphoinositol can act as the sole source of phosphate for the cell, providing functional relevance for the regulation of Git1p transport activity by phosphate.
    Journal of Biological Chemistry 08/2004; 279(30):31937-42. · 4.65 Impact Factor
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    C Almaguer, D Mantella, E Perez, J Patton-Vogt
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    ABSTRACT: Glycerophosphoinositol is produced through deacylation of the essential phospholipid phosphatidylinositol. In Saccharomyces cerevisiae, the glycerophosphoinositol produced is excreted from the cell but is recycled for phosphatidylinositol synthesis when inositol is limiting. To be recycled, glycerophosphoinositol enters the cell through the permease encoded by GIT1. The transport of exogenous glycerophosphoinositol through Git1p is sufficiently robust to support the growth of an inositol auxotroph (ino1Delta). We now report that S. cerevisiae also uses exogenous phosphatidylinositol as an inositol source. Evidence suggests that phosphatidylinositol is deacylated to glycerophosphoinositol extracellularly before being transported across the plasma membrane by Git1p. A genetic screen identified Pho86p, which is required for targeting of the major phosphate transporter (Pho84p) to the plasma membrane, as affecting the utilization of phosphatidylinositol and glycerophosphoinositol. Deletion of PHO86 in an ino1Delta strain resulted in faster growth when either phosphatidylinositol or glycerophosphoinositol was supplied as the sole inositol source. The incorporation of radiolabeled glycerophosphoinositol into an ino1Delta pho86Delta mutant was higher than that into wild-type, ino1Delta, and pho86Delta strains. All strains accumulated the most GIT1 transcript when incubated in media limited for inositol and phosphate in combination. However, the ino1Delta pho86Delta mutant accumulated approximately threefold more GIT1 transcript than did the other strains when incubated in inositol-free media containing either high or low concentrations of P(i). Deletion of PHO4 abolished GIT1 transcription in a wild-type strain. These results indicate that the transport of glycerophosphoinositol by Git1p is regulated by factors affecting both inositol and phosphate availabilities and suggest a regulatory connection between phosphate metabolism and phospholipid metabolism.
    Eukaryotic Cell 09/2003; 2(4):729-36. · 3.59 Impact Factor
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    ABSTRACT: Mutations in the Saccharomyces cerevisiae SNF1 gene affect a number of cellular processes, including the expression of genes involved in carbon source utilization and phospholipid biosynthesis. To identify targets of the Snf1 kinase that modulate expression of INO1, a gene required for an early, rate-limiting step in phospholipid biosynthesis, we performed a genetic selection for suppressors of the inositol auxotrophy of snf1Delta strains. We identified mutations in ACC1 and FAS1, two genes important for fatty acid biosynthesis in yeast; ACC1 encodes acetyl coenzyme A carboxylase (Acc1), and FAS1 encodes the beta subunit of fatty acid synthase. Acc1 was shown previously to be phosphorylated and inactivated by Snf1. Here we show that snf1Delta strains with increased Acc1 activity exhibit decreased INO1 transcription. Strains carrying the ACC1 suppressor mutation have reduced Acc1 activity in vitro and in vivo, as revealed by enzymatic assays and increased sensitivity to the Acc1-specific inhibitor soraphen A. Moreover, a reduction in Acc1 activity, caused by addition of soraphen A, provision of exogenous fatty acid, or conditional expression of ACC1, suppresses the inositol auxotrophy of snf1Delta strains. Together, these findings indicate that the inositol auxotrophy of snf1Delta strains arises in part from elevated Acc1 activity and that a reduction in this activity restores INO1 expression in these strains. These results reveal a Snf1-dependent connection between fatty acid production and phospholipid biosynthesis, identify Acc1 as a Snf1 target important for INO1 transcription, and suggest models in which metabolites that are generated or utilized during fatty acid biosynthesis can significantly influence gene expression in yeast.
    Molecular and Cellular Biology 10/2001; 21(17):5710-22. · 5.37 Impact Factor

Publication Stats

213 Citations
22 Downloads
908 Views
78.42 Total Impact Points

Institutions

  • 2003–2013
    • Duquesne University
      • • The Department of Chemistry and Biochemistry
      • • Biological Sciences
      Pittsburgh, Pennsylvania, United States
  • 2011
    • University of Nebraska at Lincoln
      • Department of Biological Sciences
      Lincoln, Nebraska, United States