scholarly journals Phosphatidylinositol-4-Kinase Type II α Is a Component of Adaptor Protein-3-derived Vesicles

2005 ◽  
Vol 16 (8) ◽  
pp. 3692-3704 ◽  
Author(s):  
Gloria Salazar ◽  
Branch Craige ◽  
Bruce H. Wainer ◽  
Jun Guo ◽  
Pietro De Camilli ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Pc12 Cells ◽  
Membrane Fraction ◽  
Functional Relationship ◽  
Adaptor Protein ◽  
Mossy Fibers ◽  
Nerve Terminals ◽  
Type Ii ◽  
Phosphatidylinositol 4 ◽  
Lysosomal Proteins

A membrane fraction enriched in vesicles containing the adaptor protein (AP) -3 cargo zinc transporter 3 was generated from PC12 cells and was used to identify new components of these organelles by mass spectrometry. Proteins prominently represented in the fraction included AP-3 subunits, synaptic vesicle proteins, and lysosomal proteins known to be sorted in an AP-3-dependent way or to interact genetically with AP-3. A protein enriched in this fraction was phosphatidylinositol-4-kinase type IIα (PI4KIIα). Biochemical, pharmacological, and morphological analyses supported the presence of PI4KIIα in AP-3-positive organelles. Furthermore, the subcellular localization of PI4KIIα was altered in cells from AP-3-deficient mocha mutant mice. The PI4KIIα normally present both in perinuclear and peripheral organelles was substantially decreased in the peripheral membranes of AP-3-deficient mocha fibroblasts. In addition, as is the case for other proteins sorted in an AP-3-dependent way, PI4KIIα content was strongly reduced in nerve terminals of mocha hippocampal mossy fibers. The functional relationship between AP-3 and PI4KIIα was further explored by PI4KIIα knockdown experiments. Reduction of the cellular content of PI4KIIα strongly decreased the punctate distribution of AP-3 observed in PC12 cells. These results indicate that PI4KIIα is present on AP-3 organelles where it regulates AP-3 function.

scholarly journals Localization of a highly active pool of type II phosphatidylinositol 4-kinase in a p97/valosin-containing-protein-rich fraction of the endoplasmic reticulum

2003 ◽  
Vol 373 (1) ◽  
pp. 57-63 ◽  
Author(s):  
Mark G. WAUGH ◽  
Shane MINOGUE ◽  
J. Simon ANDERSON ◽  
Adam BALINGER ◽  
Deena BLUMENKRANTZ ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Fraction ◽  
Unknown Origin ◽  
Type Ii ◽  
A431 Cells ◽  
Aaa Atpase ◽  
Highly Active ◽  
Active Pool ◽  
Phosphatidylinositol 4 ◽  
Protein Rich

Different phosphoinositides are synthesized in cell membranes in order to perform a variety of functions. One of the most abundant of these lipids is phosphatidylinositol (PI) 4-phosphate (PI4P), which is formed in human eukaryotes by type II and type III phosphatidylinositol 4-kinase (PI4K II and III) activities. PI4K II activity occurs in many different subcellular membranes, although no detailed analysis of the distribution of this activity has been reported. Using density gradient ultracentrifugation, we have previously found that in A431 cells the predominant PI4K activity arises from a type IIα enzyme that is localized to a buoyant membrane fraction of unknown origin [Waugh, Lawson, Tan and Hsuan (1998) J. Biol. Chem. 273, 17115–17121]. We show here that these buoyant membranes contain an activated form of PI4K IIα that can be separated from the bulk of the PI4K IIα protein in A431 and COS-7 cells. Proteomic analysis revealed that the buoyant membrane fraction contains numerous endoplasmic reticulum (ER)-marker proteins, although it was separated from the bulk of the ER, ER–Golgi intermediate compartment, transitional ER, Golgi and other major subcellular membranes. Furthermore, the majority of the cytoplasmic valosin-containing protein (VCP), an AAA+ATPase implicated in various ER, transitional ER, Golgi and nuclear functions, was almost completely localized to the same buoyant membrane fraction. Co-localization of VCP and PI4K activity was confirmed by co-immunoprecipitation. These results suggest the previously unsuspected existence of an ER-related domain in which the bulk of the cellular PI4P synthesis and VCP are localized.

scholarly journals BLOC-1 Complex Deficiency Alters the Targeting of Adaptor Protein Complex-3 Cargoes

2006 ◽  
Vol 17 (9) ◽  
pp. 4014-4026 ◽  
Author(s):  
G. Salazar ◽  
B. Craige ◽  
M. L. Styers ◽  
K. A. Newell-Litwa ◽  
M. M. Doucette ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Protein Complexes ◽  
Adaptor Protein ◽  
Cellular Mechanism ◽  
Type Ii ◽  
Mouse Mutants ◽  
Late Endosomes ◽  
Complex Deficiency ◽  
Lysosome Related Organelles ◽  
Phosphatidylinositol 4

Mutational analyses have revealed many genes that are required for proper biogenesis of lysosomes and lysosome-related organelles. The proteins encoded by these genes assemble into five distinct complexes (AP-3, BLOC-1-3, and HOPS) that either sort membrane proteins or interact with SNAREs. Several of these seemingly distinct complexes cause similar phenotypic defects when they are rendered defective by mutation, but the underlying cellular mechanism is not understood. Here, we show that the BLOC-1 complex resides on microvesicles that also contain AP-3 subunits and membrane proteins that are known AP-3 cargoes. Mouse mutants that cause BLOC-1 or AP-3 deficiencies affected the targeting of LAMP1, phosphatidylinositol-4-kinase type II alpha, and VAMP7-TI. VAMP7-TI is an R-SNARE involved in vesicle fusion with late endosomes/lysosomes, and its cellular levels were selectively decreased in cells that were either AP-3- or BLOC-1–deficient. Furthermore, BLOC-1 deficiency selectively altered the subcellular distribution of VAMP7-TI cognate SNAREs. These results indicate that the BLOC-1 and AP-3 protein complexes affect the targeting of SNARE and non-SNARE AP-3 cargoes and suggest a function of the BLOC-1 complex in membrane protein sorting.

scholarly journals Purification and characterization of human erythrocyte phosphatidylinositol 4-kinase. Phosphatidylinositol 4-kinase and phosphatidylinositol 3-monophosphate 4-kinase are distinct enzymes

1992 ◽  
Vol 284 (1) ◽  
pp. 39-45 ◽  
Author(s):  
A Graziani ◽  
L E Ling ◽  
G Endemann ◽  
C L Carpenter ◽  
L C Cantley
Keyword(s):  
Monoclonal Antibody ◽  
Human Erythrocyte ◽  
Membrane Fraction ◽  
Kinase Activity ◽  
Bovine Brain ◽  
Erythrocyte Membranes ◽  
Major Protein ◽  
Type Ii ◽  
Effective Inhibitor ◽  
Phosphatidylinositol 4

PtdIns 4-kinase has been purified 83,000-fold from human erythrocyte membranes. The major protein detected by SDS/PAGE is of molecular mass 56 kDa, and enzymic activity can be renatured from this band of the gel. The characteristics of this enzyme are similar to other type II PtdIns kinases previously described: PtdIns presented in Triton X-100 micelles is preferred as a substrate over PtdIns vesicles, the enzyme possesses a relatively low Km for ATP (20 microM), and adenosine is an effective inhibitor. A monoclonal antibody raised against bovine brain type II PtdIns 4-kinase is an effective inhibitor of the purified enzyme. PtdIns(4,5)P2 inhibits by approx. 50% when added in equimolar amounts with PtdIns; PtdIns4P has little effect on activity. A PtdIns3P 4-kinase activity has also been detected in erythrocyte lysates. Approximately two-thirds of this activity is in the cytosolic fraction and one-third in the membrane fraction. No PtdIns3P 4-kinase activity could be detected in the purified type II PtdIns 4-kinase preparation, nor could this activity be detected in a bovine brain type III PtdIns 4-kinase preparation. The monoclonal antibody that inhibits the type II PtdIns 4-kinase does not affect the PtdIns3P 4-kinase activity in the membrane fraction. The cytosolic PtdIns3P 4-kinase can be efficiently recovered from a 60%-satd.-(NH4)2SO4 precipitate that is virtually free of PtdIns 4-kinase activity. We conclude that PtdIns3P 4-kinase is a new enzyme distinct from previously characterized PtdIns 4-kinases, and that this enzyme prefers PtdIns3P over PtdIns as a substrate.

scholarly journals Biphasic regulation of type II phosphatidylinositol-4 kinase by sphingosine: Cross talk between glycero- and sphingolipids in the kidney

2014 ◽  
Vol 1838 (3) ◽  
pp. 1003-1009 ◽  
Author(s):  
Thiago Lemos ◽  
Karine S. Verdoorn ◽  
Luciana Nogaroli ◽  
Thiago Britto-Borges ◽  
Thaís A. Bonilha ◽  
...  
Keyword(s):  
Cross Talk ◽  
Type Ii ◽  
Phosphatidylinositol 4

Retinoschisis and Norrie Disease: A missing link

2021 ◽  
Author(s):  
Rahini Rajendran ◽  
Sudha Dhandayuthapani ◽  
Subbulakshmi Chidambaram ◽  
Hemavathy Nagarajan ◽  
Umashankar Vetrivel ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Functional Relationship ◽  
Direct Interaction ◽  
Specific Protein ◽  
Knock Out ◽  
Norrie Disease ◽  
Protein Protein Interaction ◽  
Protein Interactome ◽  
Molecular Relationships

Abstract Objective: Retinoschisis and Norrie disease are X-linked recessive retinal disorders caused by mutations in RS1 and NDP genes respectively. Both are likely to be monogenic and no locus heterogeneity has been reported. However, there are reports showing overlapping features of Norrie disease and retinoschisis in a NDP knock-out mouse model and also the involvement of both the genes in retinoschisis patients. Yet, the exact molecular relationships between the two disorders have still not been understood. The study investigated the association between retinoschisin (RS1) and norrin (NDP) using in vitro and in silico approaches. Specific protein-protein interaction between RS1 and NDP was analyzed in human retina by co-immunoprecipitation assay and MALDI-TOF mass spectrometry. STRING database was used to explore the functional relationship. Result: Co-immunoprecipitation demonstrated lack of a direct interaction between RS1 and NDP and was further substantiated by mass spectrometry. However, STRING revealed a potential indirect functional association between the two proteins. Progressively, our analyses indicate that FZD4 protein interactome via PLIN2 as well as the MAP kinase signaling pathway to be a likely link bridging the functional relationship between retinoschisis and Norrie disease.

The functional relationship scan in tandem mass spectrometry

1988 ◽  
Vol 23 (8) ◽  
pp. 579-584 ◽  
Author(s):  
M. Vincenti ◽  
J. C. Schwartz ◽  
R. G. Cooks ◽  
A. P. Wade ◽  
C. G. Enke
Keyword(s):  
Mass Spectrometry ◽  
Tandem Mass Spectrometry ◽  
Functional Relationship ◽  
Tandem Mass

Phosphatidylinositol 4-Kinase Type II Alpha

2018 ◽  
pp. 3934-3939
Author(s):  
Yassmeen Radif ◽  
Mark G. Waugh
Keyword(s):  
Type Ii ◽  
Phosphatidylinositol 4

Nuclear localization of phosphatidylinositol 4-kinase β

2002 ◽  
Vol 115 (8) ◽  
pp. 1769-1775 ◽  
Author(s):  
Petra de Graaf ◽  
Elsa E. Klapisz ◽  
Thomas K. F. Schulz ◽  
Alfons F. M. Cremers ◽  
Arie J. Verkleij ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Kinase Activity ◽  
Insoluble Fraction ◽  
Type Ii ◽  
3T3 Cells ◽  
Type Iii ◽  
Pore Complexes ◽  
Phosphatidylinositol 4 ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

Whereas most phosphatidylinositol 4-kinase (PtdIns 4-kinase) activity is localized in the cytoplasm, PtdIns 4-kinase activity has also been detected in membranedepleted nuclei of rat liver and mouse NIH 3T3 cells. Here we have characterized the PtdIns 4-kinase that is present in nuclei from NIH 3T3 cells. Both type II and type III PtdIns 4-kinase activity were observed in the detergent-insoluble fraction of NIH 3T3 cells. Dissection of this fraction into cytoplasmic actin filaments and nuclear lamina-pore complexes revealed that the actin filament fraction contains solely type II PtdIns 4-kinase,whereas lamina-pore complexes contain type III PtdIns 4-kinase activity. Using specific antibodies, the nuclear PtdIns 4-kinase was identified as PtdIns 4-kinase β. Inhibition of nuclear export by leptomycin B resulted in an accumulation of PtdIns 4-kinase β in the nucleus. These data demonstrate that PtdIns 4-kinase β is present in the nuclei of NIH 3T3 fibroblasts,suggesting a specific function for this kinase in nuclear processes.

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