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.

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.

Molecular determinants in pleckstrin homology domains that allow specific recognition of phosphoinositides

2001 ◽  
Vol 29 (4) ◽  
pp. 377-384 ◽  
Author(s):  
M. A. Lemmon ◽  
K. M. Ferguson
Keyword(s):  
Binding Sites ◽  
Human Proteome ◽  
Specific Interactions ◽  
Structural Studies ◽  
Pleckstrin Homology ◽  
High Affinity ◽  
Specific Recognition ◽  
Molecular Determinants ◽  
Ph Domains ◽  
Homology Domains

More than 250 pleckstrin homology (PH) domains have been identified in the human proteome. All PH domains studied to date appear to bind phosphoinositides, most binding only weakly and non-specifically. Members of a small subclass of PH domains show both high affinity and specificity for particular phosphoinositides, and recent structural studies have provided detailed views of these specific interactions. We discuss the architecture of the specific phosphoinositide-binding sites of PH domains, and how selectivity can be modulated by sequence changes.

scholarly journals Visualization of Phosphoinositides That Bind Pleckstrin Homology Domains: Calcium- and Agonist-induced Dynamic Changes and Relationship to Myo-[3H]inositol-labeled Phosphoinositide Pools

1998 ◽  
Vol 143 (2) ◽  
pp. 501-510 ◽  
Author(s):  
Péter Várnai ◽  
Tamás Balla
Keyword(s):  
Fluorescent Probe ◽  
Fluorescent Protein ◽  
Dynamic Imaging ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Protein Fusion ◽  
Fusion Construct ◽  
Cellular Expression ◽  
Ph Domains ◽  
Green Fluorescent

Phosphatidylinositol 4,5-bisphosphate (PtdIns[4,5]P2) pools that bind pleckstrin homology (PH) domains were visualized by cellular expression of a phospholipase C (PLC)δ PH domain–green fluorescent protein fusion construct and analysis of confocal images in living cells. Plasma membrane localization of the fluorescent probe required the presence of three basic residues within the PLCδ PH domain known to form critical contacts with PtdIns(4,5)P2. Activation of endogenous PLCs by ionophores or by receptor stimulation produced rapid redistribution of the fluorescent signal from the membrane to cytosol, which was reversed after Ca2+ chelation. In both ionomycin- and agonist-stimulated cells, fluorescent probe distribution closely correlated with changes in absolute mass of PtdIns(4,5)P2. Inhibition of PtdIns(4,5)P2 synthesis by quercetin or phenylarsine oxide prevented the relocalization of the fluorescent probe to the membranes after Ca2+ chelation in ionomycin-treated cells or during agonist stimulation. In contrast, the synthesis of the PtdIns(4,5)P2 imaged by the PH domain was not sensitive to concentrations of wortmannin that had been found inhibitory of the synthesis of myo-[3H]inositol– labeled PtdIns(4,5)P2. Identification and dynamic imaging of phosphoinositides that interact with PH domains will further our understanding of the regulation of such proteins by inositol phospholipids.

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 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.

scholarly journals Bacterial Pleckstrin Homology Domains: A Prokaryotic Origin for the PH Domain

2010 ◽  
Vol 396 (1) ◽  
pp. 31-46 ◽  
Author(s):  
Qingping Xu ◽  
Alex Bateman ◽  
Robert D. Finn ◽  
Polat Abdubek ◽  
Tamara Astakhova ◽  
...  
Keyword(s):  
Ph Domain ◽  
Pleckstrin Homology ◽  
Homology Domains

BCR-ABL Lacking the Pleckstrin Homology (PH) Domain of BCR Induces a More Aggressive Leukemia Than P210BCR-ABL in a Murine Model of CML.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2564-2564
Author(s):  
Shadmehr Demehri ◽  
Thomas O’Hare ◽  
Lisa J. Wood ◽  
Marc Loriaux ◽  
Brian J. Druker ◽  
...  
Keyword(s):  
G Proteins ◽  
Critical Role ◽  
In Vivo Studies ◽  
Rank Test ◽  
Ph Domain ◽  
Downstream Signaling ◽  
Ph Domains ◽  
Small G Proteins ◽  
Homology Domains

Abstract Background: The two common types of BCR-ABL fusion protein, p190 and p210, are associated with Ph-positive ALL and CML, respectively. Compared to p190, p210 retains the Dbl-like, CDC24 homology and PH domains. Dbl-like and CDC24 homology domains are thought to serve as a guanidine exchange factor (GEF) for small G proteins, including Rac, CDC42 and RhoA. It has been suggested that the loss of GEF activity may be responsible for the more aggressive phenotype of p190-positive leukemia (McWhirter and Wang, Oncogene1997; 15(14):1625–34). PH domains bind phosphoinositides and have been implicated in targeting proteins to cellular membranes as well as modulating the activity of the adjacent Dbl-like GEFs. It is not known, however, if the PH domain of BCR plays a role in determining disease features. We therefore decided to study the biology of p200BCR-ABL, a naturally occurring variant identified in several CML patients that lacks the PH domain but retains the Dbl-like and CDC24 homology domains. Materials and methods: p190, p210 and p200BCR-ABL cDNAs were cloned into pSRa and MIGR1 for expression in cell lines and primary murine bone marrow (BM) cells, respectively. The activation of signaling pathways downstream of BCR-ABL was studied in Ba/F3 and 32D cells expressing the various constructs. For colony-forming assays, murine BM cells were infected with p210, p200 or p190 retroviral vectors and seeded in methylcellulose in the presence or absence of cytokines (SCF, IL3 & IL6) and colonies were counted on day 12. For in vivo studies, BM cells isolated from 5-fluorouracil treated 5-6 week-old female Balb/c mice were infected with the various retroviral constructs in the presence of cytokines (SCF, IL3 & IL6), and 4.0 x 105 cells were injected retro-orbitally into lethally irradiated recipient mice. Disease latency was determined by monitoring white blood counts at 48h intervals. Moribund animals were sacrificed and studied by FACS and histopathology. Results: As expected, p200 transformed Ba/F3 and 32D cells to IL-3 independence. Immunoblotting of whole cell lysates with a phosphotyrosine antibody revealed no significant differences in global protein phosphorylation between the three variants, but phosphorylation of BCR-ABL was consistently less sensitive to imatinib inhibition in cells expressing p190 compared to p200 and p210. Analysis of downstream signaling showed approximately 15 fold higher levels of phosphorylated STAT6 in Ba/F3 cells expressing p200 and p190 vs. p210, while STAT5 and AKT phophorylation were similar in all variants. In the absence of cytokines, BM cells from Balb/c mice transduced with p200 and p190 generated significantly more CFU-GM than cells transduced with p210 (p<0.001, Student’s t-test). The significance of these findings was tested in vivo in a murine CML model. Mice transplanted with p210-transduced cells survived 16–20 days, significantly longer than mice transplanted with p200 or p190 expressing cells (13–15 days) (p = 0.01, log-rank test). Conclusion: p200 is closely related to p190 but very distinct from p210 with respect to downstream signaling and in vivo transforming potential. These data suggest a critical role for the PH domain of BCR in attenuating the phenotype of the leukemia. We speculate that deletion of the PH domain may disrupt the ability of the Dbl-like and CDC24 homology domains to function as a GEF towards small G proteins.

scholarly journals Signal-dependent membrane targeting by pleckstrin homology (PH) domains

2000 ◽  
Vol 350 (1) ◽  
pp. 1-18 ◽  
Author(s):  
Mark A. LEMMON ◽  
Kathryn M. FERGUSON
Keyword(s):  
Fluorescent Protein ◽  
Cell Signalling ◽  
Physiological Role ◽  
Structural Basis ◽  
Membrane Targeting ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Oligomeric State ◽  
Ph Domains ◽  
Green Fluorescent

Pleckstrin homology (PH) domains are small protein modules of around 120 amino acids found in many proteins involved in cell signalling, cytoskeletal rearrangement and other processes. Although several different protein ligands have been proposed for PH domains, their only clearly demonstrated physiological function to date is to bind membrane phosphoinositides. The PH domain from phospholipase C-δ1 binds specifically to PtdIns(4,5)P2 and its headgroup, and has become a valuable tool for studying cellular PtdIns(4,5)P2 functions. More recent developments have demonstrated that a subset of PH domains recognizes the products of agonist-stimulated phosphoinositide 3-kinases. Fusion of these PH domains to green fluorescent protein has allowed dramatic demonstrations of their independent ability to drive signal-dependent recruitment of their host proteins to the plasma membrane. We discuss the structural basis for this 3-phosphoinoistide recognition and the role that it plays in cellular signalling. PH domains that bind specifically to phosphoinositides comprise only a minority (perhaps 15%) of those known, raising questions as to the physiological role of the remaining 85% of PH domains. Most (if not all) PH domains bind weakly and non-specifically to phosphoinositides. Studies of dynamin-1 have indicated that oligomerization of its PH domain may be important in driving membrane association. We discuss the possibility that membrane targeting by PH domains with low affinity for phosphoinositides could be driven by alteration of their oligomeric state and thus the avidity of their membrane binding.

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