scholarly journals Evidence for an Intrinsic Toxicity of Phosphatidylcholine to Sec14p-dependent Protein Transport from the Yeast Golgi Complex

2001 ◽  
Vol 12 (4) ◽  
pp. 1117-1129 ◽  
Author(s):  
Zhigang Xie ◽  
Min Fang ◽  
Vytas A. Bankaitis
Keyword(s):  
Phospholipase D ◽  
Protein Transport ◽  
Choline Uptake ◽  
Secretory Function ◽  
Direct Production ◽  
Methylation Pathway ◽  
The Media ◽  
Golgi Secretory Function ◽  
Phosphatidylinositol Transfer ◽  
Cytidine Diphosphate

Yeast phosphatidylinositol-transfer protein (Sec14p) is essential for Golgi secretory function and cell viability. This requirement of Sec14p is relieved by genetic inactivation of the cytidine diphosphate-choline pathway for phosphatidycholine (PtdCho) biosynthesis. Standard phenotypic analyses indicate that inactivation of the phosphatidylethanolamine (PtdEtn) pathway for PtdCho biosynthesis, however, does not rescue the growth and secretory defects associated with Sec14p deficiency. We now report inhibition of choline uptake from the media reveals an efficient “bypass Sec14p” phenotype associated with PtdEtn-methylation pathway defects. We further show that the bypass Sec14p phenotype associated with PtdEtn-methylation pathway defects resembles other bypass Sec14p mutations in its dependence on phospholipase D activity. Finally, we find that increased dosage of enzymes that catalyze phospholipase D-independent turnover of PtdCho, via mechanisms that do not result in a direct production of phosphatidic acid or diacylglycerol, effect a partial rescue of sec14-1ts-associated growth defects. Taken together, these data support the idea that PtdCho is intrinsically toxic to yeast Golgi secretory function.

scholarly journals A phosphatidylinositol transfer protein controls the phosphatidylcholine content of yeast Golgi membranes

1994 ◽  
Vol 124 (3) ◽  
pp. 273-287 ◽  
Author(s):  
TP McGee ◽  
HB Skinner ◽  
EA Whitters ◽  
SA Henry ◽  
VA Bankaitis
Keyword(s):  
Protein Transport ◽  
Phospholipid Composition ◽  
Membrane Phospholipid ◽  
Secretory Function ◽  
Golgi Membrane ◽  
Transfer Protein ◽  
Golgi Membranes ◽  
Phosphatidylcholine Biosynthesis ◽  
Phosphatidylinositol Transfer

SEC14p is required for protein transport from the yeast Golgi complex. We describe a quantitative analysis of yeast bulk membrane and Golgi membrane phospholipid composition under conditions where Golgi secretory function has been uncoupled from its usual SEC14p requirement. The data demonstrate that SEC14p specifically functions to maintain a reduced phosphatidylcholine content in Golgi membranes and indicate that overproduction of SEC14p markedly reduces the apparent rate of phosphatidylcholine biosynthesis via the CDP-choline pathway in vivo. We suggest that SEC14p serves as a sensor of Golgi membrane phospholipid composition through which the activity of the CDP-choline pathway in Golgi membranes is regulated such that a phosphatidylcholine content that is compatible with the essential secretory function of these membranes is maintained.

scholarly journals TheSchizosaccharomyces pombe spo20+Gene Encoding a Homologue ofSaccharomyces cerevisiaeSec14 Plays an Important Role in Forespore Membrane Formation

2001 ◽  
Vol 12 (4) ◽  
pp. 901-917 ◽  
Author(s):  
Yukiko Nakase ◽  
Taro Nakamura ◽  
Aiko Hirata ◽  
Sheri M. Routt ◽  
Henry B. Skinner ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Secretory Function ◽  
Membrane Structures ◽  
Direct Role ◽  
Haploid Nucleus ◽  
Transfer Protein ◽  
Golgi Secretory Function ◽  
Phosphatidylinositol Transfer

The Schizosaccharomyces pombe spo20-KC104 mutation was originally isolated in a screen for sporulation-deficient mutants, and the spo20-KC104 mutant exhibits temperature-sensitive growth. Herein, we report that S. pombe, spo20+is essential for fission yeast cell viability and is constitutively expressed throughout the life cycle. We also demonstrate that thespo20+gene product is structurally homologous to Saccharomyces cerevisiae Sec14, the major phosphatidylinositol transfer protein of budding yeast. This structural homology translates to a significant degree of functional relatedness because reciprocal complementation experiments demonstrate that each protein is able to fulfill the essential function of the other. Moreover, biochemical experiments show that, like Sec14, Spo20 is a phosphatidylinositol/phosphatidylcholine-transfer protein. That Spo20 is required for Golgi secretory function in vegetative cells is indicated by our demonstration that the spo20-KC104mutant accumulates aberrant Golgi cisternae at restrictive temperatures. However, a second phenotype observed in Spo20-deficient fission yeast is arrest of cell division before completion of cell separation. Consistent with a direct role for Spo20 in controlling cell septation in vegetatively growing cells, localization experiments reveal that Spo20 preferentially localizes to the cell poles and to sites of septation of fission yeast cells. We also report that, when fission yeasts are challenged with nitrogen starvation, Spo20 translocates to the nucleus. This nuclear localization persists during conjugation and meiosis. On completion of meiosis, Spo20 translocates to forespore membranes, and it is the assembly of forespore membranes that is abnormal in spo20-KC104 cells. In such mutants, a considerable fraction of forming prespores fail to encapsulate the haploid nucleus. Our results indicate that Spo20 regulates the formation of specialized membrane structures in addition to its recognized role in regulating Golgi secretory function.

scholarly journals Activation of exocytosis by cross-linking of the IgE receptor is dependent on ADP-ribosylation factor 1-regulated phospholipase D in RBL-2H3 mast cells: evidence that the mechanism of activation is via regulation of phosphatidylinositol 4,5-bisphosphate synthesis

2000 ◽  
Vol 346 (1) ◽  
pp. 63-70 ◽  
Author(s):  
Gemma WAY ◽  
Niamh O'LUANAIGH ◽  
Shamshad COCKCROFT
Keyword(s):  
Mast Cells ◽  
Phospholipase D ◽  
Protein Interactions ◽  
Cross Linking ◽  
Ige Receptor ◽  
Local Synthesis ◽  
Adp Ribosylation ◽  
Phosphatidylinositol Transfer ◽  
Lipid Protein Interactions ◽  
Adp Ribosylation Factor

The physiological stimulus to exocytosis in mast cells is the cross-linking of the high-affinity IgE receptor, FcϵR1, with antigen. We demonstrate a novel function for ADP-ribosylation factor 1 (ARF1) in the regulation of antigen-stimulated secretion using cytosol-depleted RBL-2H3 mast cells for reconstitution of secretory responses. When antigen is used as the stimulus, ARF1 also reconstitutes phospholipase D activation. Using ethanol to divert the phosphatidic acid (the product of phospholipase D activity) to phosphatidylethanol causes inhibition of ARF1-reconstituted secretion. In addition. ARF1 causes an increase in phosphatidylinositol 4,5-bisphosphate (PIP2) levels at the expense of phosphatidylinositol 4-monophosphate. The requirement for PIP2 in exocytosis was confirmed by using phosphatidylinositol transfer protein (PITPα) to increase PIP2 levels. Exocytosis, restored by either ARF1 or PITPα, was inhibited when PIP2 levels were depleted by phospholipase C∆1. We conclude that the function of ARF1 and PITPα is to increase the local synthesis of PIP2, the function of which in exocytosis is likely to be linked to lipid-protein interactions, whereby recruitment of key components of the exocytotic machinery are targeted to the appropriate membrane compartment.

scholarly journals Phospholipase D activity is required for suppression of yeast phosphatidylinositol transfer protein defects

1998 ◽  
Vol 95 (21) ◽  
pp. 12346-12351 ◽  
Author(s):  
Z. Xie ◽  
M. Fang ◽  
M. P. Rivas ◽  
A. J. Faulkner ◽  
P. C. Sternweis ◽  
...  
Keyword(s):  
Phospholipase D ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

scholarly journals Pleiotropic Alterations in Lipid Metabolism in Yeastsac1Mutants: Relationship to “Bypass Sec14p” and Inositol Auxotrophy

1999 ◽  
Vol 10 (7) ◽  
pp. 2235-2250 ◽  
Author(s):  
Marcos P. Rivas ◽  
Brian G. Kearns ◽  
Zhigang Xie ◽  
Shuling Guo ◽  
M. Chandra Sekar ◽  
...  
Keyword(s):  
Phospholipase D ◽  
Phospholipid Metabolism ◽  
Metabolic Phenotype ◽  
Transcriptional Expression ◽  
Pathway Activity ◽  
Transfer Protein ◽  
Phosphatidylcholine Biosynthesis ◽  
Phosphatidylinositol Transfer ◽  
Phosphatidylinositol 4

SacIp dysfunction results in bypass of the requirement for phosphatidylinositol transfer protein (Sec14p) function in yeast Golgi processes. This effect is accompanied by alterations in inositol phospholipid metabolism and inositol auxotrophy. Elucidation of how sac1mutants effect “bypass Sec14p” will provide insights into Sec14p function in vivo. We now report that, in addition to a dramatic accumulation of phosphatidylinositol-4-phosphate,sac1 mutants also exhibit a specific acceleration of phosphatidylcholine biosynthesis via the CDP-choline pathway. This phosphatidylcholine metabolic phenotype is sensitive to the two physiological challenges that abolish bypass Sec14p insac1 strains; i.e. phospholipase D inactivation and expression of bacterial diacylglycerol (DAG) kinase. Moreover, we demonstrate that accumulation of phosphatidylinositol-4-phosphate in sac1mutants is insufficient to effect bypass Sec14p. These data support a model in which phospholipase D activity contributes to generation of DAG that, in turn, effects bypass Sec14p. A significant fate for this DAG is consumption by the CDP-choline pathway. Finally, we determine that CDP-choline pathway activity contributes to the inositol auxotrophy of sac1 strains in a novel manner that does not involve obvious defects in transcriptional expression of theINO1 gene.

Lipid homoeostasis and Golgi secretory function

2006 ◽  
Vol 34 (3) ◽  
pp. 363-366 ◽  
Author(s):  
S. Lev
Keyword(s):  
Lipid Binding ◽  
Membrane Domains ◽  
Functional Identity ◽  
Secretory Function ◽  
Critical Levels ◽  
Exact Role ◽  
Golgi Membranes ◽  
Golgi Secretory Function ◽  
Underlying Mechanisms ◽  
Specific Membrane

The unique lipid composition of the Golgi membranes is critical for maintaining their structural and functional identity, and is regulated by local lipid metabolism, a variety of lipid-binding, -modifying, -sensing and -transfer proteins, and by selective lipid sorting mechanisms. A growing body of evidence suggests that certain lipids, such as phosphoinositides and diacylglycerol, regulate Golgi-mediated transport events. However, their exact role in this process, and the underlying mechanisms that maintain their critical levels in specific membrane domains of the Golgi apparatus, remain poorly understood. Nevertheless, recent advances have revealed key regulators of lipid homoeostasis in the Golgi complex and have demonstrated their role in Golgi secretory function.

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