Biomedical subjects

Bong-Hyun Ahn

Publications and source records attributed to Bong-Hyun Ahn.

10 recordsLinked to original sources

PubMed2006

Phospholipase Cgamma1 stimulates transcriptional activation of the matrix metalloproteinase-3 gene via the protein kinase C/Raf/ERK cascade.

The phospholipid hydrolase phospholipase Cgamma1 (PLCgamma1) plays a major role in regulation of cell proliferation, development, and cell motility. Overexpression of PLCgamma1 is associated with tumor development, and it is overexpressed in some tumors. Matrix metalloproteinase-3 (MMP-3) is a protein involved in tumor invasion and metastasis. Here, we demonstrate that overexpression of PLCgamma1 stimulates MMP-3 expression at the transcriptional level via the PKC-mediated Raf/MEK1/ERK signaling cascade. We propose that modulation of PLCgamma1 activity might be of value in controlling the activity of MMPs, which are important regulators of invasion and metastasis in malignant tumors.

Animals↗

PubMed2006

Phospholipase D1 is associated with amyloid precursor protein in Alzheimer's disease.

Amyloid precursor protein (APP) is a widely expressed transmembrane protein of unknown function that is involved in the pathogenesis of Alzheimer's disease (AD). We investigated the involvement of phospholipase D (PLD) in the pathophysiology of AD. We showed dramatic upregulation of PLD1 immunoreactivity in reactive astroglial cells in brain tissue sections from authentic AD patients. Expression and activity of PLD1 were up-regulated in brain tissues from AD patients, especially caveolae membrane fraction, compared with those of control brains. Interestingly, PLD1 physically interacts and colocalizes with APP and caveolin-3. We found that APP was associated with the pleckstrin homology domain of PLD1, and the amyloid region of APP interacted with PLD. Elevated expression of APP stimulated PLD activity in human astroglioma cells. These results suggest that up-regulation of PLD might have a role in the neuronal pathology associated with AD.

Aged↗

PubMed2006

Phospholipase D is activated and phosphorylated by casein kinase-II in human U87 astroglioma cells.

Elevated expression of protein casein kinase II (CKII) stimulated basal phospholipase D (PLD) activity as well as PMA-induced PLD activation in human U87 astroglioma cells. Moreover, CKII-selective inhibitor, emodin and apigenin suppressed PMA-induced PLD activation in a dose-dependent manner as well as basal PLD activity, suggesting the involvement of CKII in the activation of both PLD1 and PLD2. CKII was associated with PLD1 and PLD2 in co-transfection experiments. Furthermore, CKII induced serine/threonine phosphorylation of PLD2 in vivo, and the multiple regions of PLD2 were phosphorylated by CKII in vitro kinase assay using glutathione S-transferase-PLD2 fusion protein fragments. Elevated expression of CKII or PLD increased cell proliferation but pretreatment of cells with 1-butanol suppressed CKII-induced cell proliferation. These results suggest that CKII is involved in proliferation of U87 cells at least in part, through stimulation of PLD activity.

1-Butanol↗

PubMed2006

The mammalian longevity-associated gene product p66shc regulates mitochondrial metabolism.

Previous studies have determined that mice with a homozygous deletion in the adapter protein p66(shc) have an extended life span and that cells derived from these mice exhibit lower levels of reactive oxygen species. Here we demonstrate that a fraction of p66(shc) localizes to the mitochondria and that p66(shc-/-) fibroblasts have altered mitochondrial energetics. In particular, despite similar cytochrome content, under basal conditions, the oxygen consumption of spontaneously immortalized p66(shc-/-) mouse embryonic fibroblasts were lower than similarly maintained wild type cells. Differences in oxygen consumption were particularly evident under chemically uncoupled conditions, demonstrating that p66(shc-/-) cells have a reduction in both their resting and maximal oxidative capacity. We further demonstrate that reconstitution of p66(shc) expression in p66(shc-/-) cells increases oxygen consumption. The observed defect in oxidative capacity seen in p66(shc-/-) cells is partially offset by augmented levels of aerobic glycolysis. This metabolic switch is manifested by p66(shc-/-) cells exhibiting an increase in lactate production and a stricter requirement for extracellular glucose in order to maintain intracellular ATP levels. In addition, using an in vivo NADH photobleaching technique, we demonstrate that mitochondrial NADH metabolism is reduced in p66(shc-/-) cells. These results demonstrate that p66(shc) regulates mitochondrial oxidative capacity and suggest that p66(shc) may extend life span by repartitioning metabolic energy conversion away from oxidative and toward glycolytic pathways.

Adaptor Proteins, Signal Transducing↗

PubMed2004

Phospholipase D isozymes mediate epigallocatechin gallate-induced cyclooxygenase-2 expression in astrocyte cells.

Little is known about the effect of epigallocatechin-3 gallate (EGCG), a major constituent of green tea, on the expression of cyclooxygenase (COX)-2. Here, we studied the role of phospholipase D (PLD) isozymes in EGCG-induced COX-2 expression. Stimulation of human astrocytoma cells (U87) with EGCG induced formation of phosphatidylbutanol, a specific product of PLD activity, and synthesis of COX-2 protein and its product, prostaglandin E(2) (PGE(2)). Pretreatment of cells with 1-butanol, but not 3-butanol, suppressed EGCG-induced COX-2 expression and PGE synthesis. Furthermore, evidence that PLD was involved in EGCG-induced COX-2 expression was provided by the observations that COX-2 expression was stimulated by overexpression of PLD1 or PLD2 isozymes and treatment with phosphatidic acid (PA), and that prevention of PA dephosphorylation by 1-propranolol significantly potentiated COX-2 expression induced by EGCG. EGCG induced activation of p38 mitogen-activated protein kinase (p38 MAPK), and specific inhibition of p38 MAPK dramatically abolished EGCG-induced PLD activation, COX-2 expression, and PGE(2) formation. Moreover, protein kinase C (PKC) inhibition suppressed EGCG-induced p38 MAPK activation, COX-2 expression, and PGE(2) accumulation. The same pathways as those obtained (2)in the astrocytoma cells were active in primary rat astrocytes, suggesting the relevance of the findings. Collectively, our results demonstrate for the first time that PLD isozymes mediate EGCG-induced COX-2 expression through PKC and p38 in immortalized astroglial line and normal astrocyte cells.

1-Butanol↗

PubMed2004

Phospholipase C, protein kinase C, Ca2+/calmodulin-dependent protein kinase II, and redox state are involved in epigallocatechin gallate-induced phospholipase D activation in human astroglioma cells.

We show that epigallocatechin-3 gallate (EGCG), a major component of green tea, stimulates phospholipase D (PLD) activity in U87 human astroglioma cells. EGCG-induced PLD activation was abolished by the phospholipase C (PLC) inhibitor and a lipase inactive PLC-gamma1 mutant, which is dependent on intracellular or extracellular Ca(2+), with the possible involvement of Ca(2+)/calmodulin-dependent protein kinase II (CaM kinase II). EGCG induced translocation of PLC-gamma1 from the cytosol to the membrane and PLC-gamma1 interaction with PLD1. EGCG regulates the activity of PLD by modulating the redox state of the cells, and antioxidants reverse this effect. Moreover, EGCG-induced PLD activation was reduced by PKC inhibitors or down-regulation of PKC. Taken together, these results show that, in human astroglioma cells, EGCG regulates PLD activity via a signaling pathway involving changes in the redox state that stimulates a PLC-gamma1 [Ins(1,4,5)P(3)-Ca(2+)]-CaM kinase II-PLD pathway and a PLC-gamma1 (diacylglycerol)-PKC-PLD pathway.

Antineoplastic Agents↗

PubMed2003

Identification of mutations in protein kinase CKIIbeta subunit that affect its binding to ribosomal protein L41 and homodimerization.

Protein kinase CKII is composed of two catalytic (alpha or alpha') subunits and two regulatory (beta) subunits. The CKIIbeta subunit is thought to mediate the tetramer formation and interact with other target proteins. However, its physiological function remains obscure. In this study, point mutants of CKIIbeta that are defective for the L41 binding were isolated by using the reverse two-hybrid system. A sequence analysis of the point mutants revealed that Asp-26, Met-52, and Met-78 of CKIIbeta are critical for L41 binding; Asn-67 (and/or Lys-139) and Met-52 are important for CKIIbeta homodimerization. Two point mutants, R75 and R83, of CKIIbeta interacted with L5, topoisomerase IIbeta, and CKBBP1/SAG, but not with the wild-type CKIIbeta. This indicates that CKIIbeta homodimerization is not a prerequisite for its binding to target proteins. These CKIIbeta point mutants may be useful in exploring the biochemical physiological functions of CKIIbeta.

Casein Kinase II↗

PubMed2003

Transmodulation between phospholipase D and c-Src enhances cell proliferation.

Phospholipase D (PLD) has been implicated in the signal transduction pathways initiated by several mitogenic protein tyrosine kinases. We demonstrate for the first time that most notably PLD2 and to a lesser extent the PLD1 isoform are tyrosine phosphorylated by c-Src tyrosine kinase via direct association. Moreover, epidermal growth factor induced tyrosine phosphorylation of PLD2 and its interaction with c-Src in A431 cells. Interaction between these proteins is via the pleckstrin homology domain of PLD2 and the catalytic domain of c-Src. Coexpression of PLD1 or PLD2 with c-Src synergistically enhances cellular proliferation compared with expression of either molecule. While PLD activity as a lipid-hydrolyzing enzyme is not affected by c-Src, wild-type PLDs but not catalytically inactive PLD mutants significantly increase c-Src kinase activity, up-regulating c-Src-mediated paxillin phosphorylation and extracellular signal-regulated kinase activity. These results demonstrate the critical role of PLD catalytic activity in the stimulation of Src signaling. In conclusion, we provide the first evidence that c-Src acts as a kinase of PLD and PLD acts as an activator of c-Src. This transmodulation between c-Src and PLD may contribute to the promotion of cellular proliferation via amplification of mitogenic signaling pathways.

Animals↗

PubMed2002

Interaction of elongation factor-1alpha and pleckstrin homology domain of phospholipase C-gamma 1 with activating its activity.

The pleckstrin homology (PH) domain is a small motif for membrane targeting in the signaling molecules. Phospholipase C (PLC)-gamma1 has two putative PH domains, an NH(2)-terminal and a split PH domain. Here we report studies on the interaction of the PH domain of PLC-gamma1 with translational elongation factor (EF)-1alpha, which has been shown to be a phosphatidylinositol 4-kinase activator. By pull-down of cell extract with the glutathione S-transferase (GST) fusion proteins with various domains of PLC-gamma1 followed by peptide sequence analysis, we identified EF-1alpha as a binding partner of a split PH domain of PLC-gamma1. Analysis by site-directed mutagenesis of the PH domain revealed that the beta2-sheet of a split PH domain is critical for the interaction with EF-1alpha. Moreover, Dot-blot assay shows that a split PH domain specifically binds to phosphoinositides including phosphatidylinositol 4-phosphate and phosphatidylinositol 4, 5-bisphosphate (PIP(2)). So the PH domain of PLC-gamma1 binds to both EF-1alpha and PIP(2). The binding affinity of EF-1alpha to the GST.PH domain fusion protein increased in the presence of PIP(2), although PIP(2) does not bind to EF-1alpha directly. This suggests that EF-1alpha may control the binding affinity between the PH domain and PIP(2). PLC-gamma1 is substantially activated in the presence of EF-1alpha with a bell-shaped curve in relation to the molar ratio between them, whereas a double point mutant PLC-gamma1 (Y509A/F510A) that lost its binding affinity to EF-1alpha shows basal level activity. Taken together, our data show that EF-1alpha plays a direct role in phosphoinositide metabolism of cellular signaling by regulating PLC-gamma1 activity via a split PH domain.

1-Phosphatidylinositol 4-Kinase↗

PubMed2002

alpha-Synuclein interacts with phospholipase D isozymes and inhibits pervanadate-induced phospholipase D activation in human embryonic kidney-293 cells.

alpha-Synuclein has been implicated in the pathogenesis of many neurodegenerative diseases, including Parkinson's disease and Alzheimer's disease. Although the function of alpha-synuclein remains largely unknown, recent studies have demonstrated that this protein can interact with phospholipids. To address the role of alpha-synuclein in neurodegenerative disease, we have investigated whether it binds phospholipase D (PLD) and affects PLD activity in human embryonic kidney (HEK)-293 cells overexpressing wild type alpha-synuclein or the mutant forms of alpha-synuclein (A53T, A30P) associated with Parkinson's disease. Tyrosine phosphorylation of alpha-synuclein appears to play a modulatory role in the inhibition of PLD, because mutation of Tyr(125) to Phe slightly increases inhibitory effect of alpha-synuclein on PLD activity. Treatment with pervanadate or phorbol myristate acetate inhibits PLD more in HEK 293 cells overexpressing alpha-synuclein than in control cells. Binding of alpha-synuclein to PLD requires phox and pleckstrin homology domain of PLD and the amphipathic repeat region and non-Abeta component of alpha-synuclein. Although biologically important, co-transfection studies indicate that the interaction of alpha-synuclein with PLD does not influence the tendency of alpha-synuclein to form pathological inclusions. These results suggest that the association of alpha-synuclein with PLD, and modulation of PLD activity, is biologically important, but PLD does not appear to play an essential role in the pathophysiology of alpha-synuclein.

Animals↗