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Michinori Matsuo

Publications and source records attributed to Michinori Matsuo.

At least 19 recordsLinked to original sources
PubMed2007

Modulation of drug-stimulated ATPase activity of human MDR1/P-glycoprotein by cholesterol.

MDR1 (multidrug resistance 1)/P-glycoprotein is an ATP-driven transporter which excretes a wide variety of structurally unrelated hydrophobic compounds from cells. It is suggested that drugs bind to MDR1 directly from the lipid bilayer and that cholesterol in the bilayer also interacts with MDR1. However, the effects of cholesterol on drug-MDR1 interactions are still unclear. To examine these effects, human MDR1 was expressed in insect cells and purified. The purified MDR1 protein was reconstituted in proteoliposomes containing various concentrations of cholesterol and enzymatic parameters of drug-stimulated ATPase were compared. Cholesterol directly binds to purified MDR1 in a detergent soluble form and the effects of cholesterol on drug-stimulated ATPase activity differ from one drug to another. The effects of cholesterol on K(m) values of drug-stimulated ATPase activity were strongly correlated with the molecular mass of that drug. Cholesterol increases the binding affinity of small drugs (molecular mass <500 Da), but does not affect that of drugs with a molecular mass of between 800 and 900 Da, and suppresses that of valinomycin (molecular mass >1000 Da). V(max) values for rhodamine B and paclitaxel are also increased by cholesterol, suggesting that cholesterol affects turnover as well as drug binding. Paclitaxel-stimulated ATPase activity of MDR1 is enhanced in the presence of stigmasterol, sitosterol and campesterol, as well as cholesterol, but not ergosterol. These results suggest that the drug-binding site of MDR1 may best fit drugs with a molecular mass of between 800 and 900 Da, and that cholesterol may support the recognition of smaller drugs by adjusting the drug-binding site and play an important role in the function of MDR1.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2006

Efflux of sphingomyelin, cholesterol, and phosphatidylcholine by ABCG1.

Cholesterol and phospholipids are essential to the body, but an excess of cholesterol or lipids is toxic and a risk factor for arteriosclerosis. ABCG1, one of the half-type ABC proteins, is thought to be involved in cholesterol homeostasis. To explore the role of ABCG1 in cholesterol homeostasis, we examined its subcellular localization and function. ABCG1 and ABCG1-K120M, a WalkerA lysine mutant, were localized to the plasma membrane in HEK293 cells stably expressing ABCG1 and formed a homodimer. A stable transformant expressing ABCG1 exhibited efflux of cholesterol and choline phospholipids in the presence of BSA, and the cholesterol efflux was enhanced by the presence of HDL, whereas cells expressing ABCG1-K120M did not, suggesting that ATP binding and/or hydrolysis is required for the efflux. Mass and TLC analyses revealed that ABCG1 and ABCA1 secrete several species of sphingomyelin (SM) and phosphatidylcholine (PC), and SMs were preferentially secreted by ABCG1, whereas PCs were preferentially secreted by ABCA1. These results suggest that ABCA1 and ABCG1 mediate the lipid efflux in different mechanisms, in which different species of phospholipids are secreted, and function coordinately in the removal of cholesterol and phospholipids from peripheral cells.

ATP Binding Cassette Transporter, Subfamily G, Mem↗
PubMed2006

Purification and ATPase activity of human ABCA1.

ATP-binding cassette protein A1 (ABCA1) plays a major role in cholesterol homeostasis and high density lipoprotein metabolism. Apolipoprotein A-I binds to ABCA1 and cellular cholesterol and phospholipids, mainly phosphatidylcholine, are loaded onto apoA-I to form pre-beta high density lipoprotein (HDL). It is proposed that ABCA1 translocates phospholipids and cholesterol directly or indirectly to form pre-beta HDL. To explore the mechanism of ABCA1-mediated pre-beta HDL formation, we expressed human ABCA1 in insect Sf9 cells and purified it. Trypsin limited-digestion of purified ABCA1 in the detergent-soluble form suggested that it retained conformation similar to ABCA1 expressed in the membranes of human fibroblast WI-38 cells. Purified ABCA1 showed robust ATPase activity when reconstituted in liposomes made of synthetic phosphatidylcholine. ABCA1 showed lower ATPase activity when reconstituted in liposomes containing phosphatidylserine, phosphatidylethanolamine, or phosphatidylglycerol and also showed weak specificity in acyl chain species. ATPase activity was reduced by the addition of cholesterol and decreased by 25% in the presence of 20% cholesterol. Beta-sitosterol and campesterol showed similar inhibitory effects but stigmasterol did not, suggesting structure-specific interaction between ABCA1 and sterols. Glibenclamide suppressed ABCA1 ATPase, suggesting that it inhibits apoA-I-dependent cellular cholesterol efflux by suppressing ABCA1 ATPase activity. These results suggest that the ATPase activity of ABCA1 is stimulated preferentially by phospholipids with choline head groups, phosphatidylcholine and sphingomyelin. This study with purified human ABCA1 provides the first biochemical basis of the mechanism for HDL formation mediated by ABCA1.

ATP Binding Cassette Transporter 1↗
PubMed2005

KATP channel interaction with adenine nucleotides.

ATP-sensitive potassium (K(ATP)) channels are regulated by adenine nucleotides to convert changes in cellular metabolic levels into membrane excitability. Hence, elucidation of interaction of SUR and Kir6.x with adenine nucleotides is an important issue to understand the molecular mechanisms underlying the metabolic regulation of the K(ATP) channels. We analyzed direct interactions with adenine nucleotides of each subunit of K(ATP) channels. Kir6.2 binds adenine nucleotides in a Mg(2+)-independent manner. SUR has two NBFs which are not equivalent: NBF1 is a Mg(2+)-independent high affinity nucleotide binding site, whereas NBF2 is a Mg-dependent low affinity site. Although SUR has ATPase activity at NBF2, it is not used to transport substrates against the concentration gradient unlike other ABC proteins. The ATPase cycle at NBF2 serves as a sensor of cellular metabolism. This may explain the low ATP hydrolysis rate compared to other ABC proteins. Based on studies of photoaffinity labeling, a model of K(ATP) channel regulation is proposed, in which K(ATP) channel activity is regulated by SUR via monitoring the intracellular MgADP concentration. K(ATP) channel activation is expected to be induced by the cooperative interaction of ATP binding at NBF1 and MgADP binding at NBF2.

Adenine↗
PubMed2005

ABC proteins: key molecules for lipid homeostasis.

Forty-nine ABC protein genes exist on human chromosomes. Eukaryotic ABC proteins were originally recognized as drug efflux pumps involved in the multidrug resistance of cancer cells. However, it is now realized that one of their major physiological roles is cellular lipid transport and homeostasis, and their dysfunction is often associated with human diseases. ABCA1 and ABCA7 mediate the apolipoprotein-dependent formation of a high-density lipoprotein-cholesterol complex. ABCA3 is indispensable for pulmonary surfactant secretion. ABCG5 and ABCG8 are involved in the secretion of plant sterols and cholesterol into bile. However, the primary substrates and mechanism of action of these ABC proteins have not been precisely defined. In this review article, we first describe the general structure and functions of eukaryotic ABC proteins. The current model of ABCA1 functionality is then explained based on studies on a topological model, subcellular localization, apoA-I dependence of HDL formation, functional defects of Tangier disease mutants, and ATP hydrolysis of purified ABCA1. ABCA1 is supposed to function as a transporter of lipids as well as a receptor for apoA-I. ABCA3 is likely involved in accumulating phospholipids and cholesterol in lamellar bodies and in generating multivesicular structures.

ATP-Binding Cassette Transporters↗
PubMed2005

Detection of ABCA7-positive cells in salivary glands from patients with Sjögren's syndrome.

ABCA7 is a member of the subfamily A of adenosine triphosphate-binding cassette (ABC) proteins, and highly homologous to ABCA1, which mediates the release of cellular cholesterol and phospholipid to form high-density lipoprotein. ABCA1 and ABCA7 contain two large extracellular domains, ECD1 and 2, which are thought to be important for their functions. Interestingly, part of ECD1 of ABCA7 is deposited as an autoantigen of Sjögren's syndrome. To determine the relationship between ABCA7 and Sjögren's syndrome, an immunohistochemical study was conducted with salivary gland biopsy samples from patients with Sjögren's syndrome. ECD1 of human ABCA7 (amino acids 45-549) was expressed in Escherichia coli as a protein fused with glutathione-S-transferase and a monoclonal antibody, KM3095, was generated. KM3095-immunoreactive cells were observed in salivary glands from 10 of 18 patients with Sjögren's syndrome. Immunostaining of serial sections with the plasma cell marker NCL-PC indicated that most of the plasma cells infiltrating salivary glands of patients with Sjögren's syndrome were KM3095-immunoreactive. Although the pathological or biological significance is not clear, it will be intriguing to further examine the relationship between ABCA7 and Sjögren's syndrome.

ATP-Binding Cassette Transporters↗
PubMed2005

Purification and cDNA cloning of chloroplastic monodehydroascorbate reductase from spinach.

The chloroplastic isoform of monodehydroascorbate (MDA) radical reductase was purified from spinach chloroplasts and leaves. The cDNA of chloroplastic MDA reductase was cloned, and its deduced amino acid sequence, consisting of 497 residues, showed high homology with those of putative organellar MDA reductases deduced from cDNAs of several plants. The amino acid sequence of the amino terminal of the purified enzyme suggested that the chloroplastic enzyme has a transit peptide consisting of 53 residues. A southern blot analysis suggested the occurrence of a gene encoding another isoform homologous to the chloroplastic isoform in spinach. The recombinant enzyme was highly expressed in Eschericia coli using the cDNA, and purified to a homogeneous state with high specific activity. The enzyme properties of the chloroplastic isoform are presented in comparison with those of the cytosolic form.

Amino Acid Sequence↗
PubMed2004

Human ABCA3, a product of a responsible gene for abca3 for fatal surfactant deficiency in newborns, exhibits unique ATP hydrolysis activity and generates intracellular multilamellar vesicles.

ABCA3 is highly expressed at the membrane of lamellar bodies in alveolar type II cells, in which pulmonary surfactant is stored. ABCA3 gene mutations cause fatal surfactant deficiency in newborns. We established HEK293 cells stably expressing human ABCA3 and analyzed the function. Exogenously expressed ABCA3 is glycosylated and localized at the intracellular vesicle membrane. ABCA3 is efficiently photoaffinity labeled by 8-azido-[alpha(32)P]ATP, but not by 8-azido-[gamma(32)P]ATP, when the membrane fraction is incubated in the presence of orthovanadate. Photoaffinity labeling of ABCA3 shows unique metal ion-dependence and is largely reduced by membrane pretreatment with 5% methyl-beta-cyclodextrin, which depletes cholesterol. Electron micrographs show that HEK293/hABCA3 cells contain multivesicular, lamellar body-like structures, which do not exist in HEK293 host cells. Some fuzzy components such as lipids accumulate in the vesicles. These results suggest that ABCA3 shows ATPase activity, which is induced by lipids, and may be involved in the biogenesis of lamellar body-like structures.

ATP-Binding Cassette Transporters↗
PubMed2004

Microanalysis for MDR1 ATPase by high-performance liquid chromatography with a titanium dioxide column.

MDR1 is clinically important because it is involved in multidrug resistance of cancer cells and affects the pharmacokinetics of various drugs. Because MDR1 harnesses adenosine 5'-triphosphate (ATP) hydrolysis for transporting drugs, examining the effect on ATPase activity is imperative for understanding the interactions between drugs and MDR1. However, conventional assay systems for ATPase activity are not sensitive enough for screening drugs using purified MDR1. Here we report a novel method to measure ATPase activity of MDR1 using high-performance liquid chromatography equipped with a titanium dioxide column. The amount of adenosine 5'-diphosphate (ADP) produced by the ATPase reaction was determined within 2 min with a titanium dioxide column (4.6 mm ID x 100 mm). The relationship between ADP amount and chromatogram peak area was linear from 5 pmol to 10 nmol. This method made it possible to reduce the amount of purified MDR1 required for a reaction to 0.5 ng, about 1/20th of the conventional colorimetric inorganic phosphate detection assay. This method is sensitive enough to detect any subtle changes in ATPase activity of MDR1 induced by drugs and can be applied to measure ATPase activity of any protein.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2004

Alpha1-syntrophin modulates turnover of ABCA1.

ABCA1 (ATP-binding cassette transporter A1) mediates the release of cellular cholesterol and phospholipid to form high density lipoprotein. Functions of ABCA1 are highly regulated at the transcriptional and post-transcriptional levels, and the synthesized ABCA1 protein turns over rapidly with a half-life of 1-2 h. To examine whether the functions of ABCA1 are modulated by associated proteins, a yeast two-hybrid library was screened with the C-terminal 120 amino acids of ABCA1. Two PDZ (PSD95-Discs large-ZO1) proteins, alpha1-syntrophin and Lin7, were found to interact with ABCA1. Immunoprecipitation revealed that alpha1-syntrophin interacted with ABCA1 strongly and that the interaction was via the C-terminal three amino acids SYV of ABCA1. Co-expression of alpha1-syntrophin in human embryonic kidney 293 cells retarded degradation of ABCA1 and made the half-life of ABCA1 five times longer than in the cells not expressing alpha1-syntrophin. This effect is not common among PDZ-containing proteins interacting with ABCA1, because Lin7, which was also found to interact with the C terminus region of ABCA1, did not have a significant effect on the half-life of ABCA1. Co-expression of alpha1-syntrophin significantly increased the apoA-I-mediated release of cholesterol. ABCA1 was co-immunoprecipitated with alpha1-syntrophin from mouse brain. These results suggest that alpha1-syntrophin is involved in intracellular signaling, which determines the stability of ABCA1 and modulates cellular cholesterol release.

ATP Binding Cassette Transporter 1↗
PubMed2004

ATP hydrolysis-dependent multidrug efflux transporter: MDR1/P-glycoprotein.

P-glycoprotein/MDR1 was the first member of the ATP-binding cassette (ABC) transporter superfamily to be identified in a eukaryote. In eukaryotes, ABC proteins can be classified into three major groups based on function: transporters, regulators, and channels. MDR1/P-glycoprotein is a prominent member of eukaryotic export-type ABC proteins. MDR1/P-glycoprotein extrudes a very wide array of structurally dissimilar compounds, all lipophilic and ranging in mass from approximately 300 to 2000 Da, including cytotoxic drugs that act on different intracellular targets, steroid hormones, peptide antibiotics, immunosuppressive agents, calcium channel blockers, and others. Nucleotide binding and hydrolysis by MDR1/P-glycoprotein is tightly coupled with its function, substrate transport. ATP binding and hydrolysis were extensively analyzed with the purified MDR1/P-glycoprotein. The vanadate-induced nucleotide trapping method was also applied to study the hydrolysis of ATP by MDR1/P-glycoprotein. When MDR1 hydrolyzes ATP in the presence of excess orthovanadate, an analog of inorganic phosphate, it forms a metastable complex after hydrolysis. Using this method, MDR1/P-glycoprotein can be specifically photoaffinity-labeled in the membrane, if 8-azido-[alpha(32)P]ATP is used as ATP. Visualization of the structure, as well as the biochemical data, is needed to fully understand how MDR1/P-glycoprotein recognizes such a variety of compounds and how it carries its substrates across the membrane using the energy from ATP hydrolysis. To do so, large amounts of pure and stable proteins are required. Heterologous expression systems, which have been used to express P-glycoprotein, are also described.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2003

Posttranscriptional regulation of human ABCA7 and its function for the apoA-I-dependent lipid release.

ABCA7 is expressed predominantly in myelo-lymphatic tissues or reticuloendothelial cells. Physiological role and function of this protein are not fully understood. We isolated the full-length cDNA (type I) and a splicing variant cDNA (type II) of human ABCA7, and developed monoclonal antibodies against extracellular domain (ECD)1 of ABCA7. RT-PCR experiments suggested that human ABCA7 gene produced the type II mRNA in a tissue-specific manner. Immunostaining revealed that the type I ABCA7, expressed in HEK293 cells, was localized to the plasma membrane and ECD1 was exposed to the extracellular space as was the case for ABCA1. HEK293 cells expressing type I ABCA7 showed apoA-I-dependent cholesterol and phospholipid release. In contrast, type II ABCA7 appeared to be localized mainly in endoplasmic reticulum and did not show apoA-I-dependent cholesterol and phospholipid release. Alternative splicing could be involved in the post-transcriptional regulation of the expression and function of human ABCA7.

ATP-Binding Cassette Transporters↗
PubMed2003

Human ABCA7 supports apolipoprotein-mediated release of cellular cholesterol and phospholipid to generate high density lipoprotein.

Apolipoprotein-mediated release of cellular cholesterol and phospholipids was induced in HEK293 cells by expressing human ATP-binding cassette transporter A7 (ABCA7) and ABC transporter A1 (ABCA1) proteins, whether transient or stable, to generate cholesterol-rich high density lipoprotein (HDL). Green fluorescent protein (GFP) attached at their C termini did not influence the lipid release reactions. Transfected ABCA7-GFP induced apolipoprotein-mediated assembly of cholesterol-containing HDL also in L929 cells, which otherwise generate only cholesterol-deficient HDL with their endogenous ABCA1. Time-dependent release of cholesterol and phospholipid by apolipoprotein A (apoA)-I was parallel both with ABCA1 and with ABCA7 when highly expressed in HEK293 cells, but dose-dependent profiles of lipid release on apoA-I and apoA-II were somewhat different between ABCA1 and ABCA7. Analyses of the stable clones with ABCA1-GFP (293/2c) and ABCA7-GFP (293/6c) by using the same vector indicated some differences in regulation of their activities by protein kinase modulators. Dibutyryl cyclic AMP increased ABCA1-GFP and the release of cholesterol and phospholipid in 293/2c but increased neither ABCA7-GFP nor the lipid release in 293/6c. Expression of ABCA1-GFP- and apoA-I-mediated lipid release were enhanced in parallel by phorbol 12-myristate 13-acetate (PMA) in 293/2c cells. In contrast, the same treatment of 293/6c increased ABCA7-GFP, but apoA-I-mediated lipid release was significantly suppressed. Despite these different responses to PMA, all of the effects of PMA were reversed by a specific protein kinase C inhibitor Gö6976, suggesting that the changes were in fact due to protein kinase C activation. A thiol protease inhibitor, N-acetyl-Leu-Leu-norleucinal, increased the protein levels of ABCA1-GFP in 293/2c and ABCA7-GFP in 293/6c, indicating their common degradation pathway. The data indicated that human ABCA7 would compensate the function of ABCA1 for release of cell cholesterol in a certain condition(s), but post-transcriptional regulation of their activity is different.

ATP Binding Cassette Transporter 1↗
PubMed2002

Effects of mutations of ABCA1 in the first extracellular domain on subcellular trafficking and ATP binding/hydrolysis.

ABCA1 mediates release of cellular cholesterol and phospholipid to form high density lipoprotein (HDL). The three different mutants in the first extracellular domain of human ABCA1 associated with Tangier disease, R587W, W590S, and Q597R, were examined for their subcellular localization and function by using ABCA1-GFP fusion protein stably expressed in HEK293 cells. ABCA1-GFP expressed in HEK293 was fully functional for apoA-I-mediated HDL assembly. Immunostaining and confocal microscopic analyses demonstrated that ABCA1-GFP was mainly localized to the plasma membrane (PM) but also substantially in intracellular compartments. All three mutant ABCA1-GFPs showed no or little apoA-I-mediated HDL assembly. R587W and Q597R were associated with impaired processing of oligosaccharide from high mannose type to complex type and failed to be localized to the PM, whereas W590S did not show such dysfunctions. Vanadate-induced nucleotide trapping was examined to elucidate the mechanism for the dysfunction in the W590S mutant. Photoaffinity labeling of W590S with 8-azido-[alpha-(32)P]ATP was stimulated by adding ortho-vanadate in the presence of Mn(2+) as much as in the presence of wild-type ABCA1. These results suggest that the defect of HDL assembly in R587W and Q597R is due to the impaired localization to the PM, whereas W590S has a functional defect other than the initial ATP binding and hydrolysis.

ATP Binding Cassette Transporter 1↗
PubMed2002

Mutations in the linker domain of NBD2 of SUR inhibit transduction but not nucleotide binding.

ATP-sensitive potassium (K(ATP)) channels are composed of an ATP-binding cassette (ABC) protein (SUR1, SUR2A or SUR2B) and an inwardly rectifying K(+) channel (Kir6.1 or Kir6.2). Like other ABC proteins, the nucleotide binding domains (NBDs) of SUR contain a highly conserved "signature sequence" (the linker, LSGGQ) whose function is unclear. Mutation of the conserved serine to arginine in the linker of NBD1 (S1R) or NBD2 (S2R) did not alter the ability of ATP or ADP (100 microM) to displace 8-azido-[(32)P]ATP binding to SUR1, or abolish ATP hydrolysis at NBD2. We co-expressed Kir6.2 with wild-type or mutant SUR in Xenopus oocytes and recorded the resulting currents in inside-out macropatches. The S1R mutation in SUR1, SUR2A or SUR2B reduced K(ATP) current activation by 100 microM MgADP, whereas the S2R mutation in SUR1 or SUR2B (but not SUR2A) abolished MgADP activation completely. The linker mutations also reduced (S1R) or abolished (S2R) MgATP-dependent activation of Kir6.2-R50G co-expressed with SUR1 or SUR2B. These results suggest that the linker serines are not required for nucleotide binding but may be involved in transducing nucleotide binding into channel activation.

ATP-Binding Cassette Transporters↗
PubMed2002

ATP binding/hydrolysis by and phosphorylation of peroxisomal ATP-binding cassette proteins PMP70 (ABCD3) and adrenoleukodystrophy protein (ABCD1).

The 70-kDa peroxisomal membrane protein (PMP70) and adrenoleukodystrophy protein (ALDP), half-size ATP-binding cassette transporters, are involved in metabolic transport of long and very long chain fatty acids into peroxisomes. We examined the interaction of peroxisomal ATP-binding cassette transporters with ATP using rat liver peroxisomes. PMP70 was photoaffinity-labeled at similar efficiencies with 8-azido-[alpha-32P]ATP and 8-azido-[gamma-32P]ATP when peroxisomes were incubated with these nucleotides at 37 degrees C in the absence Mg2+ and exposed to UV light without removing unbound nucleotides. The photoaffinity-labeled PMP70 and ALDP were co-immunoprecipitated together with other peroxisomal proteins, which also showed tight ATP binding properties. Addition of Mg2+ reduced the photoaffinity labeling of PMP70 with 8-azido-[gamma-32P]ATP by 70%, whereas it reduced photoaffinity labeling with 8-azido-[alpha-32P]ATP by only 20%. However, two-thirds of nucleotide (probably ADP) was dissociated during removal of unbound nucleotides. These results suggest that ATP binds to PMP70 tightly in the absence of Mg2+, the bound ATP is hydrolyzed to ADP in the presence of Mg2+, and the produced ADP is dissociated from PMP70, which allows ATP hydrolysis turnover. Properties of photoaffinity labeling of ALDP were essentially similar to those of PMP70. Vanadate-induced nucleotide trapping in PMP70 and ALDP was not observed. PMP70 and ALDP were also phosphorylated at a tyrosine residue(s). ATP binding/hydrolysis by and phosphorylation of PMP70 and ALDP are involved in the regulation of fatty acid transport into peroxisomes.

ATP Binding Cassette Transporter, Subfamily D, Mem↗