scholarly journals Regulation of phospholipase D1 subcellular cycling through coordination of multiple membrane association motifs

2003 ◽  
Vol 162 (2) ◽  
pp. 305-315 ◽  
Author(s):  
Guangwei Du ◽  
Yelena M. Altshuller ◽  
Nicolas Vitale ◽  
Ping Huang ◽  
Sylvette Chasserot-Golaz ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Vesicle ◽  
Membrane Association ◽  
Ph Domain ◽  
Px Domain ◽  
Ph Domains ◽  
Phospholipase D1 ◽  
And Function ◽  
Phox Homology ◽  
Cellular Stimulation

The signaling enzyme phospholipase D1 (PLD1) facilitates membrane vesicle trafficking. Here, we explore how PLD1 subcellular localization is regulated via Phox homology (PX) and pleckstrin homology (PH) domains and a PI4,5P2-binding site critical for its activation. PLD1 localized to perinuclear endosomes and Golgi in COS-7 cells, but on cellular stimulation, translocated to the plasma membrane in an activity-facilitated manner and then returned to the endosomes. The PI4,5P2-interacting site sufficed to mediate outward translocation and association with the plasma membrane. However, in the absence of PX and PH domains, PLD1 was unable to return efficiently to the endosomes. The PX and PH domains appear to facilitate internalization at different steps. The PH domain drives PLD1 entry into lipid rafts, which we show to be a step critical for internalization. In contrast, the PX domain appears to mediate binding to PI5P, a lipid newly recognized to accumulate in endocytosing vesicles. Finally, we show that the PH domain–dependent translocation step, but not the PX domain, is required for PLD1 to function in regulated exocytosis in PC12 cells. We propose that PLD1 localization and function involves regulated and continual cycling through a succession of subcellular sites, mediated by successive combinations of membrane association interactions.

scholarly journals The PH Domain and the Polybasic c Domain of Cytohesin-1 Cooperate specifically in Plasma Membrane Association and Cellular Function

1998 ◽  
Vol 9 (8) ◽  
pp. 1981-1994 ◽  
Author(s):  
Wolfgang Nagel ◽  
Pierre Schilcher ◽  
Lutz Zeitlmann ◽  
Waldemar Kolanus
Keyword(s):  
Plasma Membrane ◽  
Cell Adhesion ◽  
Tyrosine Kinase ◽  
Biological Function ◽  
Common Mechanism ◽  
Membrane Association ◽  
Bruton’S Tyrosine Kinase ◽  
Ph Domain ◽  
Bruton's Tyrosine Kinase ◽  
Ph Domains

Recruitment of intracellular proteins to the plasma membrane is a commonly found requirement for the initiation of signal transduction events. The recently discovered pleckstrin homology (PH) domain, a structurally conserved element found in ∼100 signaling proteins, has been implicated in this function, because some PH domains have been described to be involved in plasma membrane association. Furthermore, several PH domains bind to the phosphoinositides phosphatidylinositol-(4,5)-bisphosphate and phosphatidylinositol-(3,4,5)-trisphosphate in vitro, however, mostly with low affinity. It is unclear how such weak interactions can be responsible for observed membrane binding in vivo as well as the resulting biological phenomena. Here, we investigate the structural and functional requirements for membrane association of cytohesin-1, a recently discovered regulatory protein of T cell adhesion. We demonstrate that both the PH domain and the adjacent carboxyl-terminal polybasic sequence of cytohesin-1 (c domain) are necessary for plasma membrane association and biological function, namely interference with Jurkat cell adhesion to intercellular adhesion molecule 1. Biosensor measurements revealed that phosphatidylinositol-(3,4,5)-trisphosphate binds to the PH domain and c domain together with high affinity (100 nM), whereas the isolated PH domain has a substantially lower affinity (2–3 μM). The cooperativity of both elements appears specific, because a chimeric protein, consisting of the c domain of cytohesin-1 and the PH domain of the β-adrenergic receptor kinase does not associate with membranes, nor does it inhibit adhesion. Moreover, replacement of the c domain of cytohesin-1 with a palmitoylation–isoprenylation motif partially restored the biological function, but the specific targeting to the plasma membrane was not retained. Thus we conclude that two elements of cytohesin-1, the PH domain and the c domain, are required and sufficient for membrane association. This appears to be a common mechanism for plasma membrane targeting of PH domains, because we observed a similar functional cooperativity of the PH domain of Bruton’s tyrosine kinase with the adjacent Bruton’s tyrosine kinase motif, a novel zinc-containing fold.

Phospholipase D1 and potential targets of its hydrolysis product, phosphatidic acid

2003 ◽  
Vol 31 (1) ◽  
pp. 94-97 ◽  
Author(s):  
N.T. Ktistakis ◽  
C. Delon ◽  
M. Manifava ◽  
E. Wood ◽  
I. Ganley ◽  
...  
Keyword(s):  
Phosphatidic Acid ◽  
Hydrolysis Product ◽  
Ph Domain ◽  
Novel Proteins ◽  
Fatty Acylation ◽  
Px Domain ◽  
Phospholipase D1 ◽  
Sphingosine 1 Phosphate ◽  
Phox Homology ◽  
Adp Ribosylation Factor

Phospholipase D (PLD) hydrolyses phosphatidylcholine into phosphatidic acid (PA) and choline. Our work aims to understand the properties of PLD1, and to identify downstream targets of PA. In one set of projects, we have focused on membrane-targeting mechanisms and have proposed a hierarchy of signals that allows PLD1 to localize to intracellular membranes. These signals involve a functional pleckstrin homology (PH) domain and its fatty acylation on two adjacent cysteine residues. A nearby Phox homology (PX) domain may modulate the function of the fatty acylated PH domain. This complex array of signals is probably necessitated by the targeting of PLD1 to multiple endocytic and secretory membranes under basal and signal-dependent conditions. In another set of projects, we have used chemically synthesized PA coupled to a solid support in order to identify proteins that interact with this phospholipid. Several proteins have emerged from this screen as potential targets. Some (e.g. ADP-ribosylation factor, coatomer β subunit) are involved in trafficking and their PA affinity can be understood in terms of their regulated cycling on and off membranes during rounds of transport. Others (sphingosine 1-phosphate kinase and PtdIns4P 5-kinase) are implicated in pathways that also involve PLD activation. Others still are novel proteins (brain-specific neurochondrin) whose affinity for PA may contribute to an understanding of their cellular function.

scholarly journals Regulation of cytokine-independent survival kinase (CISK) by the Phox homology domain and phosphoinositides

2001 ◽  
Vol 154 (4) ◽  
pp. 699-706 ◽  
Author(s):  
Jun Xu ◽  
Dan Liu ◽  
Gordon Gill ◽  
Zhou Songyang
Keyword(s):  
Membrane Trafficking ◽  
Direct Interaction ◽  
Lipid Binding ◽  
Ph Domain ◽  
Growth And Survival ◽  
Binding Domains ◽  
Px Domain ◽  
And Function ◽  
Phox Homology

PKB/Akt and serum and glucocorticoid–regulated kinase (SGK) family kinases are important downstream targets of phosphatidylinositol 3 (PI-3) kinase and have been shown to mediate a variety of cellular processes, including cell growth and survival. Although regulation of Akt can be achieved through several mechanisms, including its phosphoinositide-binding Pleckstrin homology (PH) domain, how SGK kinases are targeted and regulated remains to be elucidated. Unlike Akt, cytokine-independent survival kinase (CISK)/SGK3 contains a Phox homology (PX) domain. PX domains have been implicated in several cellular events involving membrane trafficking. However, their precise function remains unknown. We demonstrate here that the PX domain of CISK interacts with phosphatidylinositol (PtdIns)(3,5)P2, PtdIns(3,4,5)P3, and to a lesser extent PtdIns(4,5)P2. The CISK PX domain is required for targeting CISK to the endosomal compartment. Mutation in the PX domain that abolished its phospholipid binding ability not only disrupted CISK localization, but also resulted in a decrease in CISK activity in vivo. These results suggest that the PX domain regulates CISK localization and function through its direct interaction with phosphoinositides. Therefore, CISK and Akt have evolved to utilize different lipid binding domains to accomplish a similar mechanism of activation in response to PI-3 kinase signaling.

scholarly journals Analysis of the roles of phosphatidylinositol-4,5-bisphosphate and individual subunits in assembly, localization, and function of Saccharomyces cerevisiae target of rapamycin complex 2

2019 ◽  
Vol 30 (12) ◽  
pp. 1555-1574 ◽  
Author(s):  
Maria Nieves Martinez Marshall ◽  
Anita Emmerstorfer-Augustin ◽  
Kristin L. Leskoske ◽  
Lydia H. Zhang ◽  
Biyun Li ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Lipid Metabolism ◽  
Eukaryotic Cell ◽  
Target Of Rapamycin ◽  
Multiprotein Complex ◽  
Ph Domain ◽  
Evolutionarily Conserved ◽  
And Function

Eukaryotic cell survival requires maintenance of plasma membrane (PM) homeostasis in response to environmental insults and changes in lipid metabolism. In yeast, a key regulator of PM homeostasis is target of rapamycin (TOR) complex 2 (TORC2), a multiprotein complex containing the evolutionarily conserved TOR protein kinase isoform Tor2. PM localization is essential for TORC2 function. One core TORC2 subunit (Avo1) and two TORC2-­associated regulators (Slm1 and Slm2) contain pleckstrin homology (PH) domains that exhibit specificity for binding phosphatidylinositol-4,5- bisphosphate (PtdIns4,5P2). To investigate the roles of PtdIns4,5P2 and constituent subunits of TORC2, we used auxin-inducible degradation to systematically eliminate these factors and then examined localization, association, and function of the remaining TORC2 components. We found that PtdIns4,5P2 depletion significantly reduced TORC2 activity, yet did not prevent PM localization or cause disassembly of TORC2. Moreover, truncated Avo1 (lacking its C-terminal PH domain) was still recruited to the PM and supported growth. Even when all three PH-containing proteins were absent, the remaining TORC2 subunits were PM-bound. Revealingly, Avo3 localized to the PM independent of both Avo1 and Tor2, whereas both Tor2 and Avo1 required Avo3 for their PM anchoring. Our findings provide new mechanistic information about TORC2 and pinpoint Avo3 as pivotal for TORC2 PM localization and assembly in vivo.

scholarly journals Systematic simulation of the interactions of Pleckstrin homology domains with membranes

2021 ◽  
Author(s):  
Kyle I.P. Le Huray ◽  
He Wang ◽  
Frank Sobott ◽  
Antreas C Kalli
Keyword(s):  
Crystal Structure ◽  
Membrane Association ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Interaction Site ◽  
The Family ◽  
Family Level ◽  
Ph Domains ◽  
Anionic Lipids ◽  
Phosphatidylinositol Phosphates

Pleckstrin homology (PH) domains can recruit proteins to membranes by recognition of phosphatidylinositol phosphates (PIPs). Here we report the systematic simulation of the interactions of 100 mammalian PH domains with PIP containing model membranes. Comparison with crystal structures of PH domains bound to PIP analogues demonstrates that our method correctly identifies interactions at known canonical and non-canonical sites, while revealing additional functionally important sites for interaction not observed in the crystal structure, such as for P-Rex1 and Akt1. At the family level, we find that the β1 and β2 strands and their connecting loop constitute the primary PIP interaction site for the majority of PH domains, but we highlight interesting exceptional cases. Simultaneous interaction with multiple PIPs and clustering of PIPs induced by PH domain binding are also observed. Our findings support a general paradigm for PH domain membrane association involving multivalent interactions with anionic lipids.

scholarly journals Critical but Distinct Roles for the Pleckstrin Homology and Cysteine-Rich Domains as Positive Modulators of Vav2 Signaling and Transformation

2002 ◽  
Vol 22 (8) ◽  
pp. 2487-2497 ◽  
Author(s):  
Michelle A. Booden ◽  
Sharon L. Campbell ◽  
Channing J. Der
Keyword(s):  
Membrane Association ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Guanine Nucleotide Exchange ◽  
Ph Domains ◽  
Transforming Activity ◽  
Dh Domain ◽  
Exchange Activity

ABSTRACT Vav2, like all Dbl family proteins, possesses tandem Dbl homology (DH) and pleckstrin homology (PH) domains and functions as a guanine nucleotide exchange factor for Rho family GTPases. Whereas the PH domain is a critical positive regulator of DH domain function for a majority of Dbl family proteins, the PH domains of the related Vav and Vav3 proteins are dispensable for DH domain activity. Instead, Vav proteins contain a cysteine-rich domain (CRD) critical for DH domain function. We evaluated the contribution of the PH domain and the CRD to Vav2 guanine nucleotide exchange, signaling, and transforming activity. Unexpectedly, we found that mutations of the PH domain impaired Vav2 signaling, transforming activity, and membrane association. However, these mutations do not influence exchange activity on Rac and only slightly affect exchange on RhoA and Cdc42. We also found that the CRD was critical for the exchange activity in vitro and contributed to Vav2 membrane localization. Finally, we found that phosphoinositol 3-kinase activation synergistically enhanced Vav2 transforming and signaling activity by stimulating exchange activity but not membrane association. In conclusion, the PH domain and CRD are mechanistically distinct, positive modulators of Vav2 DH domain function in vivo.

The Phox homology (PX) domain, a new player in phosphoinositide signalling

2001 ◽  
Vol 360 (3) ◽  
pp. 513-530 ◽  
Author(s):  
Yue XU ◽  
Li-Fong SEET ◽  
Brendon HANSON ◽  
Wanjin HONG
Keyword(s):  
Ph Domain ◽  
Pleckstrin Homology ◽  
Structural Motifs ◽  
Cellular Processes ◽  
Enth Domain ◽  
Fyve Domain ◽  
Cellular Events ◽  
Px Domain ◽  
Phox Homology ◽  
Recent Establishment

Phosphoinositides are key regulators of diverse cellular processes. The pleckstrin homology (PH) domain mediates the action of PtdIns(3,4)P2, PtdIns(4,5)P2 and PtdIns(3,4,5)P3, while the FYVE domain relays the pulse of PtdIns3P. The recent establishment that the Phox homology (PX) domain interacts with PtdIns3P and other phosphoinositides suggests another mechanism by which phosphoinositides can regulate/integrate multiple cellular events via a spectrum of PX domain-containing proteins. Together with the recent discovery that the epsin N-terminal homologue (ENTH) domain interacts with PtdIns(4,5)P2, it is becoming clear that phosphoinositides regulate diverse cellular events through interactions with several distinct structural motifs present in many different proteins.

scholarly journals Phospholipase D1 localises to secretory granules and lysosomes and is plasma-membrane translocated on cellular stimulation

1998 ◽  
Vol 8 (14) ◽  
pp. 835-838 ◽  
Author(s):  
Fraser D. Brown ◽  
Nicola Thompson ◽  
Khalid M. Saqib ◽  
Joanna M. Clark ◽  
Dale Powner ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Secretory Granules ◽  
Phospholipase D1 ◽  
Cellular Stimulation

scholarly journals Pleckstrin homology (PH) domains and phosphoinositides

2007 ◽  
Vol 74 ◽  
pp. 81-93 ◽  
Author(s):  
Mark A. Lemmon
Keyword(s):  
Ph Domain ◽  
Pleckstrin Homology ◽  
High Affinity ◽  
Weak Binding ◽  
Ph Domains ◽  
Common Domain ◽  
Functional Relevance ◽  
Phox Homology ◽  
Adp Ribosylation Factor ◽  
Homology Domains

PH (pleckstrin homology) domains represent the 11th most common domain in the human proteome. They are best known for their ability to bind phosphoinositides with high affinity and specificity, although it is now clear that less than 10% of all PH domains share this property. Cases in which PH domains bind specific phosphoinositides with high affinity are restricted to those phosphoinositides that have a pair of adjacent phosphates in their inositol headgroup. Those that do not [PtdIns3P, PtdIns5P and PtdIns(3,5)P2] are instead recognized by distinct classes of domains including FYVE domains, PX (phox homology) domains, PHD (plant homeodomain) fingers and the recently identified PROPPINs (b-propellers that bind polyphosphoinositides). Of the 90% of PH domains that do not bind strongly and specifically to phosphoinositides, few are well understood. One group of PH domains appears to bind both phosphoinositides (with little specificity) and Arf (ADP-ribosylation factor) family small G-proteins, and are targeted to the Golgi apparatus where both phosphoinositides and the relevant Arfs are both present. Here, the PH domains may function as coincidence detectors. A central challenge in understanding the majority of PH domains is to establish whether the very low affinity phosphoinositide binding reported in many cases has any functional relevance. For PH domains from dynamin and from Dbl family proteins, this weak binding does appear to be functionally important, although its precise mechanistic role is unclear. In many other cases, it is quite likely that alternative binding partners are more relevant, and that the observed PH domain homology represents conservation of structural fold rather than function.

scholarly journals SNX27 mediates PDZ-directed sorting from endosomes to the plasma membrane

2010 ◽  
Vol 190 (4) ◽  
pp. 565-574 ◽  
Author(s):  
Benjamin E.L. Lauffer ◽  
Cristina Melero ◽  
Paul Temkin ◽  
Cai Lei ◽  
Wanjin Hong ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Pdz Domain ◽  
Cis Acting ◽  
Sorting Nexin ◽  
Px Domain ◽  
Early Endosomes ◽  
Discs Large ◽  
Protein Scaffolding ◽  
Phox Homology

Postsynaptic density 95/discs large/zonus occludens-1 (PDZ) domain–interacting motifs, in addition to their well-established roles in protein scaffolding at the cell surface, are proposed to act as cis-acting determinants directing the molecular sorting of transmembrane cargo from endosomes to the plasma membrane. This hypothesis requires the existence of a specific trans-acting PDZ protein that mediates the proposed sorting operation in the endosome membrane. Here, we show that sorting nexin 27 (SNX27) is required for efficient PDZ-directed recycling of the β2-adrenoreceptor (β2AR) from early endosomes. SNX27 mediates this sorting function when expressed at endogenous levels, and its recycling activity requires both PDZ domain–dependent recognition of the β2AR cytoplasmic tail and Phox homology (PX) domain–dependent association with the endosome membrane. These results identify a discrete role of SNX27 in PDZ-directed recycling of a physiologically important signaling receptor, and extend the concept of cargo-specific molecular sorting in the recycling pathway.

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