Phosphatidylinositol 3-Phosphate-interacting Domains in PIKfyve

Sbrissa, Diego; Ikonomov, Ognian C.; Shisheva, Assia

doi:10.1074/jbc.m110194200

Cited by 99 publications

(38 citation statements)

References 39 publications

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“…Substrate binding is a well established mechanism for enzyme targeting. PIKfyve for example, the kinase that produces PI(3,5)P 2 , binds to PI(3)P with its FYVE domain (37). Binding of the product, as also observed in our assays, might provide a mechanism to concentrate an enzyme at a specific place for a subsequent function.…”

Section: Resultssupporting

confidence: 62%

Membrane association of myotubularin-related protein 2 is mediated by a pleckstrin homology-GRAM domain and a coiled-coil dimerization module

Berger¹,

Schaffitzel²,

Berger³

et al. 2003

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

105

128

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Mutations in the myotubularin (MTM)-related protein 2 (MTMR2) gene are responsible for the severe autosomal recessive neuropathy Charcot-Marie-Tooth disease type 4B1. MTMR2 belongs to the MTM family of dual-specific phosphatases that use phosphatidylinositol (PI) 3,5-bisphosphate [PI(3,5)P 2] and PI 3-phosphate [PI(3)P] as their substrate. Because these substrates are localized in the membrane bilayer, membrane targeting of Mtmr2 is an important regulatory mechanism. In hypoosmotically stressed COS cells with increased levels of PI(3,5)P 2, Mtmr2 is bound to the membrane of vacuoles formed under these conditions. Using several mutant forms of Mtmr2, we identified two domains that are necessary for membrane association: (i) A pleckstrin homology-GRAM domain; and (ii) a coiled-coil module. Protein-lipid overlay assays show that the pleckstrin homology-GRAM domain binds to PI(3,5)P 2 and PI(5)P, a substrate and a product of the Mtmr2 enzyme, respectively. We also demonstrate that Mtmr2 forms a dimer and that the C-terminal coiled-coil is responsible for homodimerization, in addition to membrane association. Our data indicate that phosphoinositide-protein interactions, as well as protein-protein interactions, are necessary for the correct regulation of MTMR2.

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Section: Resultssupporting

confidence: 62%

Membrane association of myotubularin-related protein 2 is mediated by a pleckstrin homology-GRAM domain and a coiled-coil dimerization module

Berger¹,

Schaffitzel²,

Berger³

et al. 2003

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

105

128

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“…The FENS-1 protein exhibits a 48 nM affinity for liposomes containing PtdIns(3)P, and similar binding is displayed by its isolated FYVE domain (50). The interaction of PIKfyve with PtdIns(3)P-containing liposomes is shown to be mediated by the FYVE domain and involves an affinity of at least 550 nM (51). The FYVE domains of Drosophila Hrs and Vps27p have affinities of 42 and 75 nM, respectively, for monolayers which contain PtdIns(3)P (37).…”

Section: Resultsmentioning

confidence: 99%

Multivalent Mechanism of Membrane Insertion by the FYVE Domain

Kutateladze

Capelluto

Ferguson

et al. 2004

Journal of Biological Chemistry

109

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Targeting of a wide variety of proteins to membranes involves specific recognition of phospholipid head groups and insertion into lipid bilayers. For example, proteins that contain FYVE domains are recruited to endosomes through interaction with phosphatidylinositol 3-phosphate (PtdIns(3)P). However, the structural mechanism of membrane docking and insertion by this domain remains unclear. Here, the depth and angle of micelle insertion and the lipid binding properties of the FYVE domain of early endosome antigen 1 are estimated by NMR spectroscopy. Spin label probes incorporated into micelles identify a hydrophobic protuberance that inserts into the micelle core and is surrounded by interfacially active polar residues. A novel proxyl PtdIns(3)P derivative is developed to map the position of the phosphoinositide acyl chains, which are found to align with the membrane insertion element. Dual engagement of the FYVE domain with PtdIns(3)P and dodecylphosphocholine micelles yields a 6-fold enhancement of affinity. The additional interaction of phosphatidylserine with a conserved basic site of the protein further amplifies the micelle binding affinity and dramatically alters the angle of insertion. Thus, the FYVE domain is targeted to endosomes through the synergistic action of stereospecific PtdIns(3)P head group ligation, hydrophobic insertion and electrostatic interactions with acidic phospholipids.Cellular processes including signal transduction, vesicular trafficking, and cytoskeletal rearrangement require selective recruitment of proteins to membrane surfaces. Well established mechanisms for localizing cytosolic proteins to membranes include electrostatic interactions through a basic peptide sequence, anchoring by covalently attached acyl chains, and association with the cytoplasmic domains of transmembrane proteins (reviewed in Refs. 1 and 2). (4), PDZ (5), and PTB (6) domains. Although the majority of these domains can interact with several PIs, the FYVE domain is remarkably selective for PtdIns(3)P (7-9). In addition to PI ligation, these domains often insert hydrophobic elements into the membrane bilayer, as has been demonstrated for the C2 (10), ENTH (11), FERM (12), FYVE (13), and PX (14) domains and the vinculin tail (15). Insertion into the membrane can be accompanied by interactions with multiple lipid head groups. For example, the PX domain of the p47 subunit of the NADPH oxidase binds cooperatively to PtdIns(3)P and phosphatidic acid (16), and the vinculin tail co-ligates phosphatidylinositol 4,5-bisphosphate and PtdSer (15). Although it is becoming evident that insertion of proteins into membranes is widespread, the three-dimensional orientations and quantitative binding properties remain challenging to characterize. The most common electron paramagnetic resonance and fluorescence approaches have provided important insights (17-20) but require covalent attachment of paramagnetic groups to various positions of the protein or mutations of residues. The inevitable effects of these modifications on lipi...

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“…Thus, eGFP2xFYVE, a probe comprising two tandem FYVE domains of PIKfyve that binds to PtdIns3P-enriched endosomal membranes with high affinity (12,24), was found to similarly colocalize with internalized dextran (10 min) or with the early endosomal marker EEA1 in both PIKfyve WT/KO and PIKfyve WT/WT MEFs ( Fig. 4C and data not shown).…”

Section: Disruption Of Pikfyve Gene and Generation Of Wt/flox Mice-mentioning

confidence: 87%

“…PIKfyve harbors a PtdIns3P-binding module, i.e. the FYVE finger domain that associates with the PtdIns3P-enriched endosomal membranes, assuring a rapid PIKfyve recruitment to this low abundance substrate of its catalytic activity (12). PIKfyve interacts physically or functionally with multiple partner proteins; the ones involved in PtdIns(3,5)P 2 homeostasis are the best studied (9).…”

mentioning

confidence: 99%

The Phosphoinositide Kinase PIKfyve Is Vital in Early Embryonic Development

Ikonomov¹,

Sbrissa²,

Delvecchio³

et al. 2011

Journal of Biological Chemistry

Self Cite

129

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Gene mutations in the phosphoinositide-metabolizing enzymes are linked to various human diseases. In mammals, PIKfyve synthesizes PtdIns(3,5)P 2 and PtdIns5P lipids that regulate endosomal trafficking and responses to extracellular stimuli. The consequence of pikfyve gene ablation in mammals is unknown. To clarify the importance of PIKfyve and PIKfyve lipid products, in this study, we have characterized the first mouse model with global deletion of the pikfyve gene using the Cre-loxP approach. We report that nearly all PIKfyve KO/KO mutant embryos died before the 32-64-cell stage. Cultured fibroblasts derived from PIKfyve flox/flox embryos and rendered pikfyve-null by Cre recombinase expression displayed severely reduced DNA synthesis, consistent with impaired cell division causing early embryo lethality. The heterozygous PIKfyve WT/KO mice were born at the expected Mendelian ratio and developed into adulthood. PIKfyve WT/KO mice were ostensibly normal by several other in vivo, ex vivo, and in vitro criteria despite the fact that their levels of the PIKfyve protein and in vitro enzymatic activity in cells and tissues were 50 -55% lower than those of wild-type mice. Consistently, steady-state levels of the PIKfyve products PtdIns(3,5)P 2 and PtdIns5P selectively decreased, but this reduction (35-40%) was 10 -15% less than that expected based on PIKfyve protein reduction. The nonlinear decrease of the PIKfyve protein versus PIKfyve lipid products, the potential mechanism(s) discussed herein, may explain how one functional allele in PIKfyve WT/KO mice is able to support the demands for PtdIns(3,5)P 2 /PtdIns5P synthesis during life. Our data also shed light on the known human disorder linked to PIKFYVE mutations.Reversible phosphorylation by kinases and phosphatases at positions 3, 4, and/or 5 of the inositol head group in phosphatidylinositol (PtdIns) 2 generates a family of seven phosphoinositide species (1-6). They all are now found to function as versatile membrane-anchored signals that control diverse and essential cellular processes. Consequently, mutations in the genes encoding the phosphoinositide-metabolizing enzymes are associated with an increasing number of human diseases (1-6). The mammalian enzyme that makes PtdIns(3,5)P 2 from PtdIns3P and PtdIns5P from PtdIns is PIKfyve (7,8). It is an evolutionarily conserved large protein of ϳ230 kDa, a product of a single gene in the animal kingdom, whose function is required for proper performance of the endosomal system and certain signaling pathways (9 -11). PIKfyve harbors a PtdIns3P-binding module, i.e. the FYVE finger domain that associates with the PtdIns3P-enriched endosomal membranes, assuring a rapid PIKfyve recruitment to this low abundance substrate of its catalytic activity (12). PIKfyve interacts physically or functionally with multiple partner proteins; the ones involved in PtdIns(3,5)P 2 homeostasis are the best studied (9). Thus, PIKfyve physically associates with the antagonistic enzyme, i.e. the PtdIns(3,5)P 2 -specific phosphatase Sac3, which ...

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Phosphatidylinositol 3-Phosphate-interacting Domains in PIKfyve

Cited by 99 publications

References 39 publications

Membrane association of myotubularin-related protein 2 is mediated by a pleckstrin homology-GRAM domain and a coiled-coil dimerization module

Membrane association of myotubularin-related protein 2 is mediated by a pleckstrin homology-GRAM domain and a coiled-coil dimerization module

Multivalent Mechanism of Membrane Insertion by the FYVE Domain

The Phosphoinositide Kinase PIKfyve Is Vital in Early Embryonic Development

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