-
Mary LaLonde,
Hilde Janssens,
Suyong Yun,
Juan Crosby,
Olga Redina,
Virginie Olive, Yelena M Altshuller,
Seok-Yong Choi,
Guangwei Du,
J Peter Gergen,
Michael A Frohman
[show abstract]
[hide abstract]
ABSTRACT: Cellularization of the Drosophila embryo is an unusually synchronous form of cytokinesis in which polarized membrane extension proceeds in part through incorporation of new membrane via fusion of apically-translocated Golgi-derived vesicles.
We describe here involvement of the signaling enzyme Phospholipase D (Pld) in regulation of this developmental step. Functional analysis using gene targeting revealed that cellularization is hindered by the loss of Pld, resulting frequently in early embryonic developmental arrest. Mechanistically, chronic Pld deficiency causes abnormal Golgi structure and secretory vesicle trafficking.
Our results suggest that Pld functions to promote trafficking of Golgi-derived fusion-competent vesicles during cellularization.
BMC Developmental Biology 02/2006; 6:60. · 2.79 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Insulin stimulates glucose uptake in fat and muscle by mobilizing Glut4 glucose transporters from intracellular membrane storage sites to the plasma membrane. This process requires the trafficking of Glut4-containing vesicles toward the cell periphery, docking at exocytic sites, and plasma membrane fusion. We show here that phospholipase D (PLD) production of the lipid phosphatidic acid (PA) is a key event in the fusion process. PLD1 is found on Glut4-containing vesicles, is activated by insulin signaling, and traffics with Glut4 to exocytic sites. Increasing PLD1 activity facilitates glucose uptake, whereas decreasing PLD1 activity is inhibitory. Diminished PA production does not substantially hinder trafficking of the vesicles or their docking at the plasma membrane, but it does impede fusion-mediated extracellular exposure of the transporter. The fusion block caused by RNA interference-mediated PLD1 deficiency is rescued by exogenous provision of a lipid that promotes fusion pore formation and expansion, suggesting that the step regulated by PA is late in the process of vesicle fusion.
Molecular Biology of the Cell 07/2005; 16(6):2614-23. · 4.94 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: 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.
The Journal of Cell Biology 08/2003; 162(2):305-15. · 10.26 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Phospholipase D (PLD), a signal-transducing membrane-associated enzyme, is implicated in diverse processes including apoptosis,
ERK activation, and glucose transport. Prior studies have identified specific PLD activators and repressors that directly
regulate its enzymatic activity. Using two-hybrid screens, we have identified PEA-15 as a PLD interactor that unexpectedly
functions to alter its level of expression. PEA-15 is a widely expressed death effector domain-containing phosphoprotein involved
in signal transduction, apoptosis, ERK activation, and glucose transport. The PLD1-interacting site on PEA-15 consists of
part of the death effector domain domain plus additional C-terminal flanking sequences, whereas the PEA-15-interacting site
on PLD1 overlaps the previously identified RhoA-interacting site. PEA-15 did not affect basal or stimulatedin vitro PLD1 enzymatic activation. However, co-expression of PEA-15 increased levels of PLD1 activity. This increased activation
correlated with higher PLD1 protein expression levels, as marked by faster accumulation and longer persistence of PLD1 when
PEA-15 was present. PEA-15 similarly increased protein expressions level of PLD2 and co-immunoprecipitated with it. These
results suggest that PEA-15 may stabilize PLD or act as a PLD chaperone. The common involvement of PEA-15 and PLD in apoptosis,
ERK activation, and glucose transport additionally suggests functional significance.
Journal of Biological Chemistry 11/2000; 275(45):35224-35232. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Activation of phosphatidylcholine-specific phospholipase D (PLD) has been implicated as a critical step in numerous cellular
pathways, including signal transduction, membrane trafficking, and the regulation of mitosis. We report here the identification
of the first human PLD cDNA, which defines a new and highly conserved gene family. Characterization of recombinant human PLD1
reveals that it is membrane-associated, selective for phosphatidylcholine, stimulated by phosphatidylinositol 4,5-bisphosphate,
activated by the monomeric G-protein ADP-ribosylation factor-1, and inhibited by oleate. PLD1 likely encodes the gene product
responsible for the most widely studied endogenous PLD activity.
Journal of Biological Chemistry 12/1995; 270(50):29640-29643. · 4.77 Impact Factor
-
