Research Interests: Biochemistry, Chemistry, Filtration, Medicine, Cytoskeleton, and 15 moreSignal Transduction, Animals, Neurons, Time Factors, Guanosine Triphosphate, Tubulin, G protein, Quntitative Thin Layer Chromatography, Protein Binding, Gtp Binding Proteins, Nucleotide, Transducin, Biochemistry and cell biology, GTP, and Medical biochemistry and metabolomics
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Research Interests: Biochemistry, Biology, Medicine, Cytoskeleton, Signal Transduction, and 14 moreAnimals, Male, Microtubule Dynamics, Monoclonal Antibodies, Rats, Guanosine Triphosphate, Tubulin, G protein, Gtp Binding Proteins, Oligonucleotides, Biochemistry and cell biology, Adenylyl Cyclase, GTP, and Medical biochemistry and metabolomics
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Research Interests: Biochemistry, Computer Science, Biophysics, Chemistry, Electron Microscopy, and 15 moreKinetics, Transmission Electron Microscopy, Medicine, Brain, Magnesium, Animals, Microtubule Dynamics, Microtubules, Citation, Guanosine Triphosphate, Swine, Nucleotide, Biochemistry and cell biology, GTP, and Medical biochemistry and metabolomics
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Research Interests: Chemistry, Chromatography, Neurochemistry, Medicine, Animals, and 13 moreMicrotubule Dynamics, Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis, High Pressure Liquid Chromatography, Guanosine Triphosphate, Swine, Tubulin, Nucleoside Diphosphate Kinase, EGTA, Molecular weight, Brain Chemistry, Neurosciences, Elution, and GTP
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The active site of the exchangeable nucleotide-binding site of tubulin was studied by using diastereoisomers A (Sp) and B (Rp) of guanosine 5'-O-(2-thiotriphosphate) (GTP beta S) where the phosphorus atom to which sulfur is... more
The active site of the exchangeable nucleotide-binding site of tubulin was studied by using diastereoisomers A (Sp) and B (Rp) of guanosine 5'-O-(2-thiotriphosphate) (GTP beta S) where the phosphorus atom to which sulfur is attached is chiral. Turbidimetric measurements were used to follow kinetics, and electron microscopy was used to evaluate polymeric forms. Both isomers at 0.5 mM promoted the assembly of tubulin in buffer containing 0.1 M 2-(N-morpholino)ethanesulfonic acid, 30% glycerol, 3 mM MgCl2, and 1 mM EGTA, pH 6.6, 23-37 degrees C. GTP beta S(A) promoted assembly into microtubules, although a few bundles were also found by electron microscopy. However, GTP beta S(B) induced assembly of tubulin into bundles of sheets and microtubules. As expected, 0.5 mM GTP induced tubulin to assemble into microtubules, thin sheets, and a few bundles. Both GTP and GTP beta S(A) were hydrolyzed in the tubulin polymers. However, more than 95% of the bound GTP beta S(B) was not hydrolyzed. Higher concentrations of GTP beta S(B), i.e., 1 mM, also induced bundles of sheets and microtubules, with 86% of the thionucleotide bound as the triphosphate. The GTP beta S(B)-induced polymers were considerably more cold stable than the GTB beta S(A)-induced microtubules, which were more cold stable than GTP-induced polymers. Mg(II) (2-5 mM) had minimal effects on the structures induced by GTP beta S(A) or -(B) isomers in the tubulin assembly system. However, at 1 mM Mg(II), no assembly was found with GTP beta S(A) and tubulin.(ABSTRACT TRUNCATED AT 250 WORDS)
Research Interests: Biochemistry, Chemistry, Electron Microscopy, Kinetics, Medicine, and 15 moreMagnesium, Animals, Stereochemistry, Ligand Binding, Guanosine Triphosphate, Swine, Hydrolysis, Binding Site, Nucleotide, Biochemistry and cell biology, binding sites, GTP, Medical biochemistry and metabolomics, In Vitro Techniques, and stereoisomerism
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Research Interests: Signal Processing, Biology, Cell Biology, Medicine, Cytoskeleton, and 15 moreCell Division, Signal Transduction, Brain, Sheep, Animals, Microtubule Dynamics, Microtubules, Biochemical, Tubulin, G protein, Protein Quaternary Structure, Cell Surface Markers, Biochemistry and cell biology, heterotrimeric G-protein, and Medical biochemistry and metabolomics
Neurodegeneration, a progressive loss of nerve cells (neurons), occurs in many neurological disorders including Alzheimer’s disease, Parkinson’s disease, Schizophrenia and drug addiction. Disruption of Microtubules (MTs), a major... more
Neurodegeneration, a progressive loss of nerve cells (neurons), occurs in many neurological disorders including Alzheimer’s disease, Parkinson’s disease, Schizophrenia and drug addiction. Disruption of Microtubules (MTs), a major component of cytoskeleton and aggregation of proteins associated with them is the hallmark of neurodegeneration. Gβγ, an important component of G protein signaling has been shown to induce neurite outgrowth of PC12 cells by interacting with microtubules. The goal of the present study is to understand whether interfering with Gβγ-MT mediated pathway causes neurodegeneration. Because prenylation of γ subunits is important for the interaction of Gβγ with MTs, we used inhibitors (L-23 and L-28) for PMPMEase (polyisoprenylated methylated protein methyl esterase), an enzyme involved in the prenylation pathway to conduct the study. PC12 cells were treated with NGF over the course of three days, followed by overnight treatment with L-28 or L-23. Confocal microscopy was used to analyze the results. We found that more than 70% of PC12 cells exhibit neurite formation in the presence of NGF. Neurite formation was not affected significantly in the presence of PMSF or L-23 (100mM). L-28 (10μM), on the other hand significantly reduced neurite formation as well as MTs and Gβγ labeling. In addition, severe cellular degeneration was observed (more than 60% areas in the slides). The result suggests that inhibitors of PMPMEase could be used as valuable tools to study the mechanism of neurodegeneration and design strategies to develop effective drugs against drug addiction and other neurodegenerative disorders
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Research Interests: Microbiology, Parasitology, Zoology, Tropical Medicine, Fluorescent Dyes and Reagents, and 22 moreCytoskeleton, Network, Secretory Pathway, Endocytosis, Parasite, Animals, Endoplasmic Reticulum, Microtubule Dynamics, Actin, Microtubules, Actin Cytoskeleton, Nuclear envelope, Fluorescent Antibody Technique, Energy Production, Tubulin, For, Veterinary Sciences, Protozoan Proteins, Quaternary Ammonium Compounds, Boron Compounds, Ceramides, and Clathrin
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Research Interests: Microbiology, Parasitology, Zoology, Tropical Medicine, Fluorescent Dyes and Reagents, and 22 moreCytoskeleton, Network, Secretory Pathway, Endocytosis, Parasite, Animals, Endoplasmic Reticulum, Microtubule Dynamics, Actin, Microtubules, Actin Cytoskeleton, Nuclear envelope, Fluorescent Antibody Technique, Energy Production, Tubulin, For, Veterinary Sciences, Protozoan Proteins, Quaternary Ammonium Compounds, Boron Compounds, Ceramides, and Clathrin
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Giardia lamblia, a protozoan parasite, is a major cause of waterborne infection, worldwide. While the trophozoite form of this parasite induces pathological symptoms in the gut, the cyst form transmits the infection. Since Giardia is a... more
Giardia lamblia, a protozoan parasite, is a major cause of waterborne infection, worldwide. While the trophozoite form of this parasite induces pathological symptoms in the gut, the cyst form transmits the infection. Since Giardia is a noninvasive parasite, the actual mechanism by which it causes disease remains elusive. We have previously reported that Giardia assembles cholesterol and GM1 glycosphingolipid-enriched lipid rafts (LRs) that participate in encystation and cyst production. To further delineate the role of LRs in pathogenesis, we isolated LRs from Giardia and subjected them to proteomic analysis. Various cellular proteins including potential virulence factors—e.g., giardins, variant surface proteins, arginine deaminases, elongation factors, ornithine carbomyltransferases, and high cysteine-rich membrane proteins—were found to be present in LRs. Since Giardia secretes virulence factors encapsulated in extracellular vesicles (EVs) that induce proinflammatory responses in ...
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The βγ subunit of heterotrimeric G proteins, a key molecule in the G protein-coupled receptors (GPCRs) signaling pathway, has been shown to be an important factor in the modulation of the microtubule cytoskeleton. Gβγ has been shown to... more
The βγ subunit of heterotrimeric G proteins, a key molecule in the G protein-coupled receptors (GPCRs) signaling pathway, has been shown to be an important factor in the modulation of the microtubule cytoskeleton. Gβγ has been shown to bind to tubulin, stimulate microtubule assembly, and promote neurite outgrowth of PC12 cells. In this study, we demonstrate that in addition to microtubules, Gβγ also interacts with actin filaments, and this interaction increases during NGF-induced neuronal differentiation of PC12 cells. We further demonstrate that the Gβγ-actin interaction occurs independently of microtubules as nocodazole, a well-known microtubule depolymerizing agent did not inhibit Gβγ-actin complex formation in PC12 cells. A confocal microscopic analysis of NGF-treated PC12 cells revealed that Gβγ co-localizes with both actin and microtubule cytoskeleton along neurites, with specific co-localization of Gβγ with actin at the distal end of these neuronal processes. Furthermore, we show that Gβγ interacts with the actin cytoskeleton in primary hippocampal and cerebellar rat neurons. Our results indicate that Gβγ serves as an important modulator of the neuronal cytoskeleton by interacting with both microtubules and actin filaments, and is likely to participate in various aspects of neuronal differentiation including axon and growth cone formation.
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Aberrant organization of microtubules (MTs) is known to be a feature of neurodegeneration, which occurs in many neurological disorder including Alzheimer’s disease, Parkinson’s disease, neuropsychiatric illness such as schizophrenia, and... more
Aberrant organization of microtubules (MTs) is known to be a feature of neurodegeneration, which occurs in many neurological disorder including Alzheimer’s disease, Parkinson’s disease, neuropsychiatric illness such as schizophrenia, and drug addiction. Previously, Gβγ, an important component of G protein-coupled signaling pathways, has been shown to regulate neurite outgrowth by interacting with MTs. The aim of the present study is to understand the Gβγ regulation of MT assembly and its relation to neurodegeneration, using GRK-ct peptide (consists of the carboxy terminus of G protein-coupled-receptor kinase) which is known to inhibit Gβγ-dependent signaling by binding to and sequestering Gβγ. PC12 cells were used to conduct the study because they respond to nerve growth factor (NGF) with growth arrest and exhibit a typical phenotype of neuronal cells sending out neurites. PC12 cells were treated with NGF over the course of three days followed by treatment with GRK-ct peptide for 1h, and the result was evaluated by subcellular fractionation, and confocal microscopy. Confocal microscopy revealed that the GRK-ct peptide has a very dramatic effect on morphology and survival of NGF-differentiated PC12 cells, causing cellular aggregation, neurite and MT disruption, and severe degeneration. Interestingly, the levels of MAP2, a cytoskeletal protein that stabilizes MTs, dramatically decreased in the presence of GRK-ct peptide. The results indicate that the inhibition of Gβγ-dependent signaling induce cytoskeletal alterations and neurodegeneration. Future investigation will address whether Gβγ signaling could be targeted for developing novel drugs against drug addiction and other neurodegenerative disorders
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The early-divergent protozoan Giardia lamblia, which is a major cause of waterborne enteric disease worldwide, was shown to possess limited lipid synthesis ability and to depend upon preformed lipid molecules for energy production and... more
The early-divergent protozoan Giardia lamblia, which is a major cause of waterborne enteric disease worldwide, was shown to possess limited lipid synthesis ability and to depend upon preformed lipid molecules for energy production and membrane biosynthesis. Therefore, questions regarding how Giardia imports and utilizes exogenous lipids are important. Using fluorescent lipids and fatty acids as reporter molecules, we show that anti-actin and anti-microtubule agents affect the uptake, intracellular movement and recycling of fluorescent lipids in Giardia, which indicates that lipid transport in this protozoan parasite, as in higher eukaryotes, is dependent upon cytoskeleton filaments. We propose that actin/microtubule-based lipid and vesicular transport, which is operative in Giardia, could be an ancient cellular process that most probably evolved before the emergence of the mitochondrion and other characteristic eukaryotic organelles.
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Research Interests: Biology, Cell Biology, Medicine, Synaptic Plasticity, Cytoskeleton, and 15 moreCell Division, Signal Transduction, Cell Differentiation, Animals, Microtubules, Neurons, Lysine, Acetylation, Tubulin, G protein, Amino Acid Sequence, C, Biochemistry and cell biology, centrosome, and Medical biochemistry and metabolomics
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Background: In invasive breast cancers, cancerous cells spread outside the ducts of the breast and metastasize to lung and other tissues. Although the rapid arachidonic acid (AA) metabolism and concomitant increase of eicosanoid molecules... more
Background: In invasive breast cancers, cancerous cells spread outside the ducts of the breast and metastasize to lung and other tissues. Although the rapid arachidonic acid (AA) metabolism and concomitant increase of eicosanoid molecules are involved in the proliferation and invasion of breast cancer cells, the exact mechanism by which AA metabolites regulate these phenomena are not well understood. Here, we show that leukotriene B4 (LTB4), one of the metabolites of AA, which is produced by the action of 5-lipoxygenase (5-LOX), causes the proliferation and metastatic migration of breast cancer cells. Material and Methods: MDA-MB-231 (invasive breast cancer cells) and MCF7 (non-invasive breast cancer cells) were used in this study. Briefly, cells were treated with AA (100 μM) and nordihydroguaiarectin acid (NDGA, 10 μM), and the secreted eicosanoids were characterized by HPLC. Immunofluorescence microscopy was performed to elucidate the expression and intracellular localization of 5...