scholarly journals Assembly of a Fab1 Phosphoinositide Kinase Signaling Complex Requires the Fig4 Phosphoinositide Phosphatase

2008 ◽  
Vol 19 (10) ◽  
pp. 4273-4286 ◽  
Author(s):  
Roberto J. Botelho ◽  
Jem A. Efe ◽  
David Teis ◽  
Scott D. Emr
Keyword(s):  
Protein Complex ◽  
Functional Unit ◽  
Dual Function ◽  
Dual Roles ◽  
Lipid Kinase ◽  
Unknown Mechanism ◽  
Signaling Complex ◽  
Lipid Phosphatase ◽  
Phosphoinositide Phosphatase ◽  
Phosphoinositide Kinase

Phosphatidylinositol-3,5-bisphosphate [PtdIns(3,5)P2] regulates several vacuolar functions, including acidification, morphology, and membrane traffic. The lipid kinase Fab1 converts phosphatidylinositol-3-phosphate [PtdIns(3)P] to PtdIns(3,5)P2. PtdIns(3,5)P2levels are controlled by the adaptor-like protein Vac14 and the Fig4 PtdIns(3,5)P2-specific 5-phosphatase. Interestingly, Vac14 and Fig4 serve a dual function: they are both implicated in the synthesis and turnover of PtdIns(3,5)P2by an unknown mechanism. We now show that Fab1, through its chaperonin-like domain, binds to Vac14 and Fig4 and forms a vacuole-associated signaling complex. The Fab1 complex is tethered to the vacuole via an interaction between the FYVE domain in Fab1 and PtdIns(3)P on the vacuole. Moreover, Vac14 and Fig4 bind to each other directly and are mutually dependent for interaction with the Fab1 kinase. Our observations identify a protein complex that incorporates the antagonizing Fab1 lipid kinase and Fig4 lipid phosphatase into a common functional unit. We propose a model explaining the dual roles of Vac14 and Fig4 in the synthesis and turnover of PtdIns(3,5)P2.

scholarly journals Roles for a lipid phosphatase in the activation of its opposing lipid kinase

2020 ◽  
Vol 31 (17) ◽  
pp. 1835-1845
Author(s):  
Bethany S. Strunk ◽  
Noah Steinfeld ◽  
Sora Lee ◽  
Natsuko Jin ◽  
Cecilia Muñoz-Rivera ◽  
...  
Keyword(s):  
Protein Complex ◽  
Dynamic Regulation ◽  
Lipid Kinase ◽  
Lipid Phosphatase ◽  
Phosphoinositide Phosphatase ◽  
Turn Over

The phosphoinositide phosphatase Fig4 is predicted to turn over the signaling lipid PI3,5P2. It is shown that a major role of Fig4 is to elevate PI3,5P2 via dynamic regulation of the protein complex that activates its opposing lipid kinase, Fab1.

scholarly journals Screening Assay for Small-Molecule Inhibitors of Synaptojanin 1, a Synaptic Phosphoinositide Phosphatase

2013 ◽  
Vol 19 (4) ◽  
pp. 585-594 ◽  
Author(s):  
Laura Beth J. McIntire ◽  
Kyu-In Lee ◽  
Belle Chang-Ileto ◽  
Gilbert Di Paolo ◽  
Tae-Wan Kim
Keyword(s):  
Amyloid Β ◽  
Receptor Trafficking ◽  
Water Soluble ◽  
Amyloid Β Peptide ◽  
Screening Assay ◽  
Vesicle Recycling ◽  
Lipid Phosphatase ◽  
Phosphoinositide Phosphatase ◽  
Synaptojanin 1 ◽  
Behavioral Impairments

Elevation of amyloid β-peptide (Aβ) is critically associated with Alzheimer disease (AD) pathogenesis. Aβ-induced synaptic abnormalities, including altered receptor trafficking and synapse loss, have been linked to cognitive deficits in AD. Recent work implicates a lipid critical for neuronal function, phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2], in Aβ-induced synaptic and behavioral impairments. Synaptojanin 1 (Synj1), a lipid phosphatase mediating the breakdown of PI(4,5)P2, has been shown to play a role in synaptic vesicle recycling and receptor trafficking in neurons. Heterozygous deletion of Synj1 protected neurons from Aβ-induced synaptic loss and restored learning and memory in a mouse model of AD. Thus, inhibition of Synj1 may ameliorate Aβ-associated impairments, suggesting Synj1 as a potential therapeutic target. To this end, we developed a screening assay for Synj1 based on detection of inorganic phosphate liberation from a water-soluble, short-chain PI(4,5)P2. The assay displayed saturable kinetics and detected Synj1’s substrate preference for PI(4,5)P2 over PI(3,4,5)P3. The assay will enable identification of novel Synj1 inhibitors that have potential utility as chemical probes to dissect the cellular role of Synj1 as well as potential to prevent or reverse AD-associated synaptic abnormalities.

Comparative phytohormone profiles, lipid kinase and lipid phosphatase activities in barley aleurone, coleoptile, and root tissues

2012 ◽  
Vol 58 ◽  
pp. 83-88 ◽  
Author(s):  
Maria V. Meringer ◽  
Ana L. Villasuso ◽  
Susana J. Pasquaré ◽  
Norma M. Giusto ◽  
Estela E. Machado ◽  
...  
Keyword(s):  
Lipid Kinase ◽  
Phosphatase Activities ◽  
Lipid Phosphatase ◽  
Barley Aleurone ◽  
Root Tissues

scholarly journals Dual function of Rpn5 in two PCI complexes, the 26S proteasome and COP9 signalosome

2011 ◽  
Vol 22 (7) ◽  
pp. 911-920 ◽  
Author(s):  
Zanlin Yu ◽  
Oded Kleifeld ◽  
Avigail Lande-Atir ◽  
Maisa Bsoul ◽  
Maya Kleiman ◽  
...  
Keyword(s):  
Translation Initiation Factor ◽  
26S Proteasome ◽  
Initiation Factor ◽  
Cop9 Signalosome ◽  
Dual Function ◽  
Dual Roles ◽  
Eukaryotic Translation ◽  
Eukaryotic Translation Initiation ◽  
Bona Fide ◽  
Translation Initiation Factor 3

Subunit composition and architectural structure of the 26S proteasome lid is strictly conserved between all eukaryotes. This eight-subunit complex bears high similarity to the eukaryotic translation initiation factor 3 and to the COP9 signalosome (CSN), which together define the proteasome CSN/COP9/initiation factor (PCI) troika. In some unicellular eukaryotes, the latter two complexes lack key subunits, encouraging questions about the conservation of their structural design. Here we demonstrate that, in Saccharomyces cerevisiae, Rpn5 plays dual roles by stabilizing proteasome and CSN structures independently. Proteasome and CSN complexes are easily dissected, with Rpn5 the only subunit in common. Together with Rpn5, we identified a total of six bona fide subunits at roughly stoichiometric ratios in isolated, affinity-purified CSN. Moreover, the copy of Rpn5 associated with the CSN is required for enzymatic hydrolysis of Rub1/Nedd8 conjugated to cullins. We propose that multitasking by a single subunit, Rpn5 in this case, allows it to function in different complexes simultaneously. These observations demonstrate that functional substitution of subunits by paralogues is feasible, implying that the canonical composition of the three PCI complexes in S. cerevisiae is more robust than hitherto appreciated.

Investigating lipid signalling: it's all about finding the right PI

2008 ◽  
Vol 413 (1) ◽  
pp. e5-e6 ◽  
Author(s):  
Erik Nielsen
Keyword(s):  
Signal Transduction ◽  
Signal Transduction Pathways ◽  
Lipid Kinase ◽  
Phosphatase Activities ◽  
Cellular Signal Transduction ◽  
Lipid Signalling ◽  
Lipid Phosphatase ◽  
Biochemical Journal ◽  
Cellular Signal ◽  
The Right

Phosphoinositides are well-known components of cellular signal transduction pathways and, more recently, have been shown to play important roles in organelle identity and targeting determinants for various cytosolic proteins. Conversion of PtdIns into its various phosphorylated derivatives, such as PtdIns4P and PtdIns(4,5)P2, is accomplished by a series of distinct lipid kinase and lipid phosphatase activities that are localized to specific subcellular membranes. As a result, production of distinct PtdIns forms is thought to be largely dependent on the access of these enzymes to their PtdIns or PtdInsP substrates. Interestingly, an investigation of two different PIS (PtdIns synthase) isoforms by Lofke et al. in this issue of the Biochemical Journal now indicates that the ability of PtdIns to be converted into downstream PtdInsPs may depend upon the PIS isoform from which it was synthesized.

scholarly journals The Dual Roles of Zinc Sulfate in Mitigating Peach Gummosis

Plant Disease ◽  
2016 ◽  
Vol 100 (2) ◽  
pp. 345-351 ◽  
Author(s):  
Zhi Li ◽  
Yanchun Fan ◽  
Lei Gao ◽  
Xiu Cao ◽  
Junli Ye ◽  
...  
Keyword(s):  
Zinc Sulfate ◽  
Molecular Data ◽  
Lesion Size ◽  
Zinc Transporters ◽  
Dual Function ◽  
Cell Wall Degradation ◽  
Growth Morphology ◽  
Direct Inhibition ◽  
Dual Roles

Peach gummosis, caused by Lasiodiplodia theobromae, is one of the most prevalent diseases that affects peach production. In this study, we investigated the effect of zinc sulfate on inoculated peach shoots, as well as on the growth, morphology, and pathogenicity of L. theobromae in vitro, in the laboratory. Zinc deficiency was detected in diseased peach shoots by micronutrient analysis (Cu, Mn, and Zn) and confirmed by the measurement of transcript levels of zinc transporters (ZIP4, HAM4, and ZAT). The zinc was transferred from the diseased peach shoots to the peach gum. Applying zinc sulfate to the diseased peach shoots reduced the severity of peach gummosis, showing significantly reduced lesion size and gum weight, as well as downregulation of cell wall degradation-related gene (PG and PME) compared with the control. Zinc sulfate also specifically controlled peach gummosis under L. theobromae phytotoxin stress and induced the expression of defense-related genes (PR4, CHI, PAL, PGIP, and GNS3). In addition, in vitro mycelial growth of L. theobromae was significantly inhibited by zinc sulfate compared with the control. Zinc sulfate caused abnormal hyphae at 25 mM and swelling hyphal tips at 50 mM. Exposure of L. theobromae to zinc sulfate for 20 min inhibited the ability of the pathogen to cause peach gummosis. Our physiological and molecular data demonstrated that zinc sulfate has a dual function by reducing susceptibility in the host and by direct inhibition of the pathogen.

scholarly journals Functional Characterization Of The VAC14 Self-Interaction Domain

2021 ◽  
Author(s):  
Tamadher A. Alghamdi
Keyword(s):  
Functional Characterization ◽  
Regulatory Protein ◽  
Adaptor Protein ◽  
Interaction Domain ◽  
Lipid Kinase ◽  
Lipid Phosphatase ◽  
C Terminus ◽  
Regulatory Complex ◽  
Steady State Levels ◽  
And Function

PtdIns(3,5)P2 is a low-abundance signaling lipid present at < 0.1 % of total PtdIns lipids in yeasts and mammals. Reduced levels of PtdIns(3,5)P2 contributes to neurodegenerative disorders in humans and vacuolar defects in yeasts. Steady-state levels of PtdIns(3,5)P2 are dependent on both its rate of synthesis and turnover. In yeast, PtdIns(3,5)P2 is produced on the vacuole membrane by phosphorylation of PtdIns(3)P at the 5 position of its inositol ring by the Fab1 lipid kinase. Cells lacking Fab1 make no PtdIns(3,5)P2 and exhibit defects in vacuole morphology and function. The lipid phosphatase Fig4 counteracts Fab1 activity by turnover of PtdIns(3,5)P2 into PtdIns(3)P. Vac14 is a regulatory protein implicated in the synthesis and turnover of PtdIns(3,5)P2. It acts as an adaptor protein that controls both of Fab1 and Fig4 proteins. In addition, Vac14 exists as a multimer that allows for self-interaction. However, multimerization state of Vac14 as well as the domain responsible for self-interaction remained unknown. This study aimed to identify the self-interaction domain to elucidate its role in the assembly of the regulatory complex of PtdIns(3,5)P2. The observations seen in this study suggested that Vac14 self-interacts via multiple conserved motifs in the C-terminus, which are crucial for interaction with Fab1 and Fig4, and the normal morphology of yeast vacuoles.

scholarly journals Phosphorylation-Dependent Assembly of a 14-3-3 Mediated Signaling Complex During Red Blood Cell Invasion by Plasmodium falciparum Merozoites

2020 ◽  
Author(s):  
Kunal R. More ◽  
Inderjeet Kaur ◽  
Quentin Giai Gianetto ◽  
Brandon M. Invergo ◽  
Thibault Chaze ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Protein Kinase ◽  
Blood Cell ◽  
Signaling Pathways ◽  
Red Blood Cell ◽  
Protein Complex ◽  
Merozoite Invasion ◽  
Signaling Complex ◽  
Novel Approach ◽  
Low K

AbstractRed blood cell (RBC) invasion by Plasmodium merozoites requires multiple steps that are regulated by signaling pathways. Exposure of P. falciparum merozoites to the physiological signal of low K+, as found in blood plasma, leads to a rise in cytosolic Ca2+, which mediates microneme secretion, motility, and invasion. We have used global phosphoproteomic analysis of merozoites to identify signaling pathways that are activated during invasion. Using quantitative phosphoproteomics we found 394 protein phosphorylation site changes in merozoites subjected to different ionic environments (high K+/ low K+) out of which 143 were Ca2+-dependent. These included a number of signaling proteins such as catalytic and regulatory subunits of protein kinase A (PfPKAc and PfPKAr) and calcium-dependent protein kinase 1 (PfCDPK1). Proteins of the 14-3-3 family interact with phosphorylated target proteins to assemble signaling complexes. Here, using co-immunoprecipitation and gel filtration chromatography, we demonstrate that Pf14-3-3I binds phosphorylated PfPKAr and PfCDPK1 to mediate the assembly of a multi-protein complex in P. falciparum merozoites. A phospho-peptide, P1, based on the Ca2+ dependent phosphosites of PKAr, binds Pf14-3-3I and disrupts assembly of the Pf14-3-3I-mediated multi-protein complex. Disruption of the multi-protein complex with P1 inhibits microneme secretion and RBC invasion. This study thus identifies a novel signaling complex that plays a key role in merozoite invasion of RBCs. Disruption of this signaling complex could serve as a novel approach to inhibit blood stage growth of malaria parasites.ImportanceInvasion of red blood cells (RBCs) by Plasmodium falciparum merozoites is a complex process that is regulated by intricate signaling pathways. Here, we have used phosphoproteomic profiling to identify the key proteins involved in signaling events during invasion. We found changes in the phosphorylation of various merozoite proteins including multiple kinases previously implicated in the process of invasion. We also found that a phosphorylation dependent multi-protein complex including signaling kinases assembles during the process of invasion. Disruption of this multi-protein complex impairs merozoite invasion of RBCs providing a novel approach for the development of inhibitors to block the growth of blood stage malaria parasites.

scholarly journals Assembly of the PtdIns 4-kinase Stt4 complex at the plasma membrane requires Ypp1 and Efr3

2008 ◽  
Vol 183 (6) ◽  
pp. 1061-1074 ◽  
Author(s):  
Dan Baird ◽  
Chris Stefan ◽  
Anjon Audhya ◽  
Sabine Weys ◽  
Scott D. Emr
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Membrane Protein ◽  
Essential Gene ◽  
Additional Component ◽  
Lipid Kinase ◽  
Phosphoinositide Kinase ◽  
Multiple Copies ◽  
The Stability ◽  
Phosphatidylinositol 4

The phosphoinositide phosphatidylinositol 4-phosphate (PtdIns4P) is an essential signaling lipid that regulates secretion and polarization of the actin cytoskeleton. In Saccharomyces cerevisiae, the PtdIns 4-kinase Stt4 catalyzes the synthesis of PtdIns4P at the plasma membrane (PM). In this paper, we identify and characterize two novel regulatory components of the Stt4 kinase complex, Ypp1 and Efr3. The essential gene YPP1 encodes a conserved protein that colocalizes with Stt4 at cortical punctate structures and regulates the stability of this lipid kinase. Accordingly, Ypp1 interacts with distinct regions on Stt4 that are necessary for the assembly and recruitment of multiple copies of the kinase into phosphoinositide kinase (PIK) patches. We identify the membrane protein Efr3 as an additional component of Stt4 PIK patches. Efr3 is essential for assembly of both Ypp1 and Stt4 at PIK patches. We conclude that Ypp1 and Efr3 are required for the formation and architecture of Stt4 PIK patches and ultimately PM-based PtdIns4P signaling.

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