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 The PH domain from the Toxoplasma gondii PH-containing protein-1 (TgPH1) serves as an ectopic reporter of phosphatidylinositol 3-phosphate in mammalian cells

2021 ◽  
Author(s):  
Krishna Chintaluri
Keyword(s):  
Toxoplasma Gondii ◽  
Membrane Trafficking ◽  
Mammalian Cells ◽  
Effector Proteins ◽  
Ph Domain ◽  
Fyve Domain ◽  
Px Domain ◽  
Early Endosomes ◽  
Phosphatidylinositol 3

Phosphoinositides (PtdInsPs) lipids recruit effector proteins to membranes to mediate a variety of functions including signal transduction and membrane trafficking. Each PtdInsP binds to a specific set of effectors through characteristic protein domains such as the PH, FYVE and PX domains. Domains with high affinity for a single PtdInsP species are useful as probes to visualize the distribution and dynamics of that PtdInsP. The endolysosomal system is governed by two primary PtdInsPs: phosphatidylinositol-3-phosphate [PtdIns(3)P] and phosphatidylinositol-3,5-bisphosphate [PtdIns(3,5)P2], which are thought to localize and control early endosomes and lysosomes, respectively. While PtdIns(3)P has been analysed with mammalian-derived PX and FYVE domains, PtdIns(3,5)P2 indicators remain controversial. Thus, complementary probes against these PtdInsPs are needed, including those originating from non-mammalian proteins. Here, we characterized in mammalian cells the dynamics of the PH domain from PH-containing protein-1 from the parasite Toxoplasma gondii (TgPH1), which was previously shown to bind PtdIns(3,5)P2 in vitro. However, we show that TgPH1 retains membrane-binding in PIKfyve-inhibited cells, suggesting that TgPH1 is not a viable PtdIns(3,5)P2 marker in mammalian cells. Instead, PtdIns(3)P depletion using pharmacological treatments dissociated TgPH1 from membranes. Indeed, TgPH1 co-localized to EEA1-positive endosomes. In addition, TgPH1 co-localized and behaved similarly to the PX domain of p40phox and tandem FYVE domain of EEA1, which are commonly used as PtdIns(3)P indicators. Collectively, TgPH1 offers a complementary reporter for PtdIns(3)P derived from a non-mammalian protein and that is distinct from commonly employed PX and FYVE domain-based probes.

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 from the Toxoplasma gondii PH-containing protein-1 (TgPH1) serves as an ectopic reporter of phosphatidylinositol 3-phosphate in mammalian cells

2021 ◽  
Author(s):  
Krishna Chintaluri
Keyword(s):  
Toxoplasma Gondii ◽  
Membrane Trafficking ◽  
Mammalian Cells ◽  
Effector Proteins ◽  
Ph Domain ◽  
Fyve Domain ◽  
Px Domain ◽  
Early Endosomes ◽  
Phosphatidylinositol 3

Phosphoinositides (PtdInsPs) lipids recruit effector proteins to membranes to mediate a variety of functions including signal transduction and membrane trafficking. Each PtdInsP binds to a specific set of effectors through characteristic protein domains such as the PH, FYVE and PX domains. Domains with high affinity for a single PtdInsP species are useful as probes to visualize the distribution and dynamics of that PtdInsP. The endolysosomal system is governed by two primary PtdInsPs: phosphatidylinositol-3-phosphate [PtdIns(3)P] and phosphatidylinositol-3,5-bisphosphate [PtdIns(3,5)P2], which are thought to localize and control early endosomes and lysosomes, respectively. While PtdIns(3)P has been analysed with mammalian-derived PX and FYVE domains, PtdIns(3,5)P2 indicators remain controversial. Thus, complementary probes against these PtdInsPs are needed, including those originating from non-mammalian proteins. Here, we characterized in mammalian cells the dynamics of the PH domain from PH-containing protein-1 from the parasite Toxoplasma gondii (TgPH1), which was previously shown to bind PtdIns(3,5)P2 in vitro. However, we show that TgPH1 retains membrane-binding in PIKfyve-inhibited cells, suggesting that TgPH1 is not a viable PtdIns(3,5)P2 marker in mammalian cells. Instead, PtdIns(3)P depletion using pharmacological treatments dissociated TgPH1 from membranes. Indeed, TgPH1 co-localized to EEA1-positive endosomes. In addition, TgPH1 co-localized and behaved similarly to the PX domain of p40phox and tandem FYVE domain of EEA1, which are commonly used as PtdIns(3)P indicators. Collectively, TgPH1 offers a complementary reporter for PtdIns(3)P derived from a non-mammalian protein and that is distinct from commonly employed PX and FYVE domain-based probes.

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.

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.

Pleckstrin homology domains: not just for phosphoinositides

2004 ◽  
Vol 32 (5) ◽  
pp. 707-711 ◽  
Author(s):  
M.A. Lemmon
Keyword(s):  
Subcellular Localization ◽  
Human Genome ◽  
Small Gtpases ◽  
Membrane Targeting ◽  
Ph Domain ◽  
Cellular Membranes ◽  
Pleckstrin Homology ◽  
Ph Domains ◽  
Common Domain ◽  
Homology Domains

PH domains (pleckstrin homology domains) are the 11th most common domain in the human genome and are best known for their ability to target cellular membranes by binding specifically to phosphoinositides. Recent studies in yeast have shown that, in fact, this is a property of only a small fraction of the known PH domains. Most PH domains are not capable of independent membrane targeting, and those capable of doing so (approx. 33%) appear, most often, to require both phosphoinositide and non-phosphoinositide determinants for their subcellular localization. Several recent studies have suggested that small GTPases such as ARF family proteins play a role in defining PH domain localization. Some others have described a signalling role for PH domains in regulating small GTPases, although phosphoinositides may also play a role. These findings herald a change in our perspective of PH domain function, which will be significantly more diverse than previously supposed.

scholarly journals Determinants of the Endosomal Localization of Sorting Nexin 1

2005 ◽  
Vol 16 (4) ◽  
pp. 2049-2057 ◽  
Author(s):  
Qi Zhong ◽  
Martin J. Watson ◽  
Cheri S. Lazar ◽  
Andrea M. Hounslow ◽  
Jonathan P. Waltho ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Early Endosome ◽  
Nmr Structure ◽  
Sorting Nexin ◽  
High Affinity ◽  
Terminal Domain ◽  
Px Domain ◽  
Phox Homology ◽  
Phosphatidylinositol 3

The sorting nexin (SNX) family of proteins is characterized by sequence-related phox homology (PX) domains. A minority of PX domains bind with high affinity to phosphatidylinositol 3-phosphate [PI(3)P], whereas the majority of PX domains exhibit low affinity that is insufficient to target them to vesicles. SNX1 is located on endosomes, but its low affinity PX domain fails to localize in vivo. The NMR structure of the PX domain of SNX1 reveals an overall fold that is similar to high-affinity PX domains. However, the phosphatidylinositol (PI) binding pocket of the SNX1 PX domain is incomplete; regions of the pocket that are well defined in high-affinity PX domains are highly mobile in SNX1. Some of this mobility is lost upon binding PI(3)P. The C-terminal domain of SNX1 is a long helical dimer that localizes to vesicles but not to the early endosome antigen-1–containing vesicles where endogenous SNX1 resides. Thus, the obligate dimerization of SNX1 that is driven by the C-terminal domain creates a high-affinity PI binding species that properly targets the holo protein to endosomes.

scholarly journals Activated RhoA Binds to the Pleckstrin Homology (PH) Domain of PDZ-RhoGEF, a Potential Site for Autoregulation

2010 ◽  
Vol 285 (27) ◽  
pp. 21070-21081 ◽  
Author(s):  
Zhe Chen ◽  
Frank Medina ◽  
Mu-ya Liu ◽  
Celestine Thomas ◽  
Stephen R. Sprang ◽  
...  
Keyword(s):  
Ph Domain ◽  
Pleckstrin Homology ◽  
Potential Site

scholarly journals TSPO: kaleidoscopic 18-kDa amid biochemical pharmacology, control and targeting of mitochondria

2016 ◽  
Vol 473 (2) ◽  
pp. 107-121 ◽  
Author(s):  
Jemma Gatliff ◽  
Michelangelo Campanella
Keyword(s):  
Neurological Diseases ◽  
Anion Channel ◽  
Translocator Protein ◽  
Endocrine Regulation ◽  
Voltage Dependent Anion Channel ◽  
Steroid Synthesis ◽  
Cellular Processes ◽  
Cellular Events ◽  
Voltage Dependent

The 18-kDa translocator protein (TSPO) localizes in the outer mitochondrial membrane (OMM) of cells and is readily up-regulated under various pathological conditions such as cancer, inflammation, mechanical lesions and neurological diseases. Able to bind with high affinity synthetic and endogenous ligands, its core biochemical function resides in the translocation of cholesterol into the mitochondria influencing the subsequent steps of (neuro-)steroid synthesis and systemic endocrine regulation. Over the years, however, TSPO has also been linked to core cellular processes such as apoptosis and autophagy. It interacts and forms complexes with other mitochondrial proteins such as the voltage-dependent anion channel (VDAC) via which signalling and regulatory transduction of these core cellular events may be influenced. Despite nearly 40 years of study, the precise functional role of TSPO beyond cholesterol trafficking remains elusive even though the recent breakthroughs on its high-resolution crystal structure and contribution to quality-control signalling of mitochondria. All this along with a captivating pharmacological profile provides novel opportunities to investigate and understand the significance of this highly conserved protein as well as contribute the development of specific therapeutics as presented and discussed in the present review.

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