المؤلفون: Kunihisa Kobayashi, Rongying Li, Kazuhiro Oka, Lawrence Chan, Hye-Jeong Lee, Tomoya Terashima, Hui-Chen Lu, M. Cecilia Ljungberg, Keiko Sakai, Merry Sullivan, Oliver Tiebel

المصدر: Brain Research. 1276:11-21

مصطلحات موضوعية: Apolipoprotein E, Very Low-Density Lipoprotein Receptor, Article, Mice, Exon, Apolipoproteins E, Cerebellum, Animals, Humans, RNA, Messenger, Reelin, Molecular Biology, Cells, Cultured, Cerebral Cortex, Mice, Knockout, Neurons, Messenger RNA, biology, General Neuroscience, Alternative splicing, Brain, Gene Expression Regulation, Developmental, Ligand (biochemistry), Molecular biology, Mice, Inbred C57BL, Alternative Splicing, Reelin Protein, Cholesterol, Lipoproteins, IDL, Receptors, LDL, Astrocytes, RNA splicing, biology.protein, Female, Neurology (clinical), Developmental Biology

الوصف: Very-low-density lipoprotein receptor (VLDLR) is a multi ligand apolipoprotein E (apoE) receptor and is involved in brain development through Reelin signaling. Different forms of VLDLR can be generated by alternative splicing. VLDLR-I contains all exons. VLDLR-II lacks an O-linked sugar domain encoded by exon 16, while VLDLR-III lacks the third complement-type repeat in the ligand binding domain encoded by exon 4. We quantitatively compared lipoprotein binding to human VLDLR variants and analyzed their mRNA expression in both human cerebellum and mouse brain. VLDLR-III exhibited the highest capacity in binding to apoE enriched β-VLDL in vitro and was more effective in removing apoE containing lipoproteins from the circulation than other variants in vivo. In human cerebellum, the major species was VLDLR-II, while the second most abundant species was a newly identified VLDLR-IV which lacks both exon 4 and 16. VLDLR-I was present at low levels. In adult mice, exon 4 skipping varied between 30 and 47% in different brain regions, while exon 16 skipping ranged by 51–76%. Significantly higher levels of VLDLR proteins were found in mouse cerebellum and cerebral cortex than other regions. The deletions of exon 4 and exon 16 frequently occurred in primary neurons, indicating that newly identified variant VLDLR-IV is abundant in neurons. In contrast, VLDLR mRNA lacking exon 4 was not detectable in primary astrocytes. Such cell type-specific splicing patterns were found in both mouse cerebellum and cerebral cortex. These results suggest that a VLDLR variant lacking the third complement-type repeat is generated by neuron-specific alternative splicing. Such differential splicing may result in different lipid uptake in neurons and astrocytes.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::96d4520f21e8a57673b245b00fbf61e9Test
https://doi.org/10.1016/j.brainres.2009.04.030Test

المؤلفون: Kazuhiro Oka, Ta-Yuan Chang, Merry Sullivan, Catherine C.Y. Chang, Lawrence Chan, Patricia J. Uelmen

المصدر: Journal of Biological Chemistry. 270:26192-26201

مصطلحات موضوعية: Regulation of gene expression, Messenger RNA, SOAT1, Biochemistry, cDNA library, Chinese hamster ovary cell, Complementary DNA, Sterol O-acyltransferase, Cell Biology, Northern blot, Biology, Molecular Biology

الوصف: Acyl-coenzyme A:cholesterol acyltransferase (ACAT) catalyzes the esterification of cholesterol with long chain fatty acids and is believed to play an important part in the development of atherosclerotic lesions. To facilitate the study of ACAT's role in this process, we have used the human ACAT K1 clone previously described (Chang, C. C. Y., Huh, H. Y., Cadigan, K. M., and Chang, T. Y.(1993) J. Biol. Chem. 268, 20747-20755) to isolate mouse ACAT cDNA from a liver cDNA library. The 3.7-kilobase cDNA clone isolated contains a 1620-base pair open reading frame which encodes a protein of 540 amino acids. The predicted mouse ACAT protein is 87% identical to the protein product of human ACAT K1 and shares many of the same secondary structural features, including two transmembrane domains, a leucine heptad motif consistent with dimer or multimer formation, and five regions homologous to the “signature sequences” found in other enzymes that catalyze acyl adenylation followed by acyl thioester formation and acyl transfer. Using the cDNA as a hybridization probe, we mapped the gene encoding mouse ACAT to chromosome 1 in a region syntenic to human chromosome 1 where the ACAT gene is located. Northern blot analysis and RNase protection assays of mouse tissues revealed that ACAT mRNA is expressed most highly in the adrenal gland, ovary, and preputial gland and is least abundant in skeletal muscle, adipose tissue, heart, and brain. To study the dietary regulation of ACAT mRNA expression in mouse tissues, we fed C57BL/6J mice a high-fat, high-cholesterol (HF/HC) atherogenic diet for 3 weeks and measured ACAT mRNA levels in various tissues by RNase protection. The HF/HC diet had little effect on ACAT mRNA levels in the small intestine, aorta, adrenal, or peritoneal macrophages, whereas hepatic ACAT mRNA levels were doubled in mice fed the atherogenic diet. ACAT activity in liver microsomes was similarly increased in cholesterol-fed mice, suggesting that mouse ACAT is regulated at least in part at the level of mRNA abundance. Additionally, a significant positive correlation was observed between ACAT activity and microsomal free cholesterol levels in chow- and cholesterol-fed mice, supporting the concept of cholesterol availability as a regulator of ACAT. To further investigate the regulation of ACAT activity under controlled conditions, ACAT-deficient Chinese hamster ovary cells were stably transfected with the mouse ACAT cDNA clone driven by a cytomegalovirus promoter. Two transfected Chinese hamster ovary cell lines that expressed the mouse ACAT transgene regained the ability to esterify cholesterol. Cholesterol esterification activity in both of these cell lines was further increased by exposure of these cells to low density lipoprotein. Thus we have demonstrated that mouse ACAT expression in vivo and in vitro is regulated by at least two mechanisms: control of mRNA abundance and post-transcriptional regulation of the enzyme activity, probably by cholesterol availability.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_________::ae68b8f13cce88f38713bb04eb091b32Test
https://doi.org/10.1074/jbc.270.44.26192Test

المؤلفون: Nancy S. Shipley, Mitchell D. Miller, Merry Sullivan, Kurt L. Krause, Michael J. Benedik

المصدر: Journal of Molecular Biology. 222:27-30

مصطلحات موضوعية: chemistry.chemical_classification, Nuclease, Endodeoxyribonucleases, biology, RNA, Endonucleases, biology.organism_classification, Enterobacteriaceae, Serratia, chemistry.chemical_compound, Endonuclease, Crystallography, Enzyme, X-Ray Diffraction, chemistry, Structural Biology, Endoribonucleases, Serratia marcescens, biology.protein, Molecular Biology, DNA

الوصف: Crystals have been obtained of the extracellular endonuclease from the bacterial pathogen Serratia marcescens. This magnesium-dependent enzyme is equally active against single and double-stranded DNA, as well as RNA, without any apparent base preference. The Serratia nuclease is not homologous with staphylococcal nuclease, the only other broad specificity endonuclease for which a structure exists, nor is it homologous with other nucleases that have been solved by X-ray diffraction. The structure of this enzyme should, therefore, provide new information about this class of enzyme. At present we have succeeded in obtaining large, high quality crystals using ammonium sulfate. They crystallize in the orthorhombic space group P2(1)2(1)2(1), with cell dimensions a = 106.7 A, b = 74.5 A, c = 68.9 A, and diffract to beyond 2 A. Low-resolution native data sets have been recorded and a search is under way for heavy-atom derivatives.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::bb99b9ae6605ef95899bd787d2211b82Test
https://doi.org/10.1016/0022-2836Test(91)90734-n

المؤلفون: Lawrence Chan, Keh-Weei Tzung, Kazuhiro Oka, Merry Sullivan, Antonio Baldini, Elizabeth A. Lindsay

المصدر: Genomics. 20:298-300

مصطلحات موضوعية: chemistry.chemical_classification, DNA, Complementary, cDNA library, Hybridization probe, Molecular Sequence Data, Nucleic acid sequence, Chromosome Mapping, Very Low-Density Lipoprotein Receptor, Biology, Amino acid, Receptors, LDL, chemistry, Biochemistry, Complementary DNA, Genetics, Animals, Humans, Amino Acid Sequence, Rabbits, Chromosomes, Human, Pair 9, Peptide sequence, In Situ Hybridization, Fluorescence, VLDLR Gene

الوصف: A complementary DNA for the very-low-density lipoprotein receptor (VLDLR) that codes for a protein of 873 amino acids was cloned from a human heart cDNA library. The mature protein of 846 amino acids, preceded by a 27-residue signal peptide, shares 97% amino acid sequence identity with the rabbit VLDLR. Like the low-density lipoprotein receptor, the VLDLR contains five different domains, all of which are highly conserved between human and rabbit. A tetrapeptide NPVY that potentially serves as a signal for clustering of the VLDLR on coated pits is present in the cytoplasmic domain, which is 100% conserved between human and rabbit. We localized the VLDLR gene to chromosome 9p24 by fluorescence in situ hybridization using the cloned cDNA as hybridization probe. The high amino acid sequence homology of the VLDLR between two mammalian species suggests that the receptor plays a fundamental role in lipoprotein metabolism and that energy metabolism mediated by triglyceride utilization may be an evolutionarily highly conserved mechanism.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::f16c304cd0e0dd1c58376e43e8ccdae4Test
https://doi.org/10.1006/geno.1994.1171Test

المؤلفون: M. Nakamuta, Kathy Robinson, Julie Martinez, Lawrence Chan, Merry Sullivan, Kazuhiro Oka, Kazumi Ishimura-Oka, Oliver Tiebel

المصدر: Atherosclerosis. 145(2)

مصطلحات موضوعية: Apolipoprotein E, medicine.medical_specialty, Low-density lipoprotein receptor gene family, Apolipoprotein B, Molecular Sequence Data, Very Low-Density Lipoprotein Receptor, Adipose tissue, Down-Regulation, Biology, Lipoproteins, VLDL, Muscle Development, Polymerase Chain Reaction, Mice, Pregnancy, Internal medicine, medicine, Animals, RNA, Messenger, Muscle, Skeletal, VLDLR Gene, Regulation of gene expression, Mice, Knockout, Base Sequence, Myocardium, Gene Expression Regulation, Developmental, Heart, RNA Probes, Blotting, Northern, Mice, Inbred C57BL, Endocrinology, Adipose Tissue, Receptors, LDL, LDL receptor, biology.protein, Diet, Atherogenic, lipids (amino acids, peptides, and proteins), Female, Cardiology and Cardiovascular Medicine

الوصف: The very low density lipoprotein receptor (VLDLR) is a multifunctional apolipoprotein (apo) E receptor that shares a common structural feature as well as some ligand specificity to apo E with members of the low density lipoprotein receptor gene family. We have isolated and characterized the mouse VLDLR gene. The mouse VLDLR gene contains 19 exons spanning approximately 50 kb. The exon-intron organization of the gene is completely conserved between mouse and human. Since the 5'-flanking region of the mouse VLDLR gene contains two copies of a sterol regulatory element-1 like sequence (SRE-1), we next studied regulation of the VLDLR mRNA expression in heart, skeletal muscle and adipose tissue in C57BL/6, LDLR-/-, apo E-/- and LDLR-/-apo E-/- mice fed normal chow or atherogenic diet. The VLDLR mRNA expression was down-regulated 3-fold by feeding atherogenic diet in heart and skeletal muscle only in LDLR-/- mice. In contrast, VLDLR mRNA expression was up-regulated by atherogenic diet in adipose tissue in all animal models except double knockout mice. These results suggest that SRE-1 may be functional and VLDLR plays a role in cholesterol homeostasis in heart and skeletal muscle when LDLR is absent and that apo E is required for this modulation. Developmental regulation of the VLDLR mRNA expression was also tissue-specific. VLDLR mRNA expression in heart displayed significant up and down regulation during development. Maximal level was detected on post-natal day 3. However, the VLDLR mRNA levels in skeletal muscle remained relatively constant except a slight dip on post-natal day 7. In kidney and brain, VLDLR mRNA also peaked on post-natal day 3 but remained relatively constant thereafter. In liver, VLDLR mRNA expression was very low; it was barely detectable at day 19 of gestation and was decreased further thereafter. In adipose tissue, the VLDLR mRNA level showed an increase on post-natal day 13, went down again during weaning and then continued to increase afterwards. This developmental pattern as well as dietary regulation in adipose tissue supports the notion that VLDLR plays a role in lipid accumulation in this tissue. Although the primary role of VLDLR in heart, muscle and adipose tissue is likely in lipid metabolism, developmental pattern of this receptor in other tissues suggests that VLDLR has functions that are unrelated to lipid metabolism.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::4984513458765cc71a0988443bd1f1e2Test
https://pubmed.ncbi.nlm.nih.gov/10488949Test

المؤلفون: Julie Martinez, Merry Sullivan, Kunihisa Kobayashi, Ba-Bie Teng, Kazuhiro Oka, Kazumi Ishimura-Oka, Lawrence Chan

المصدر: The Journal of biological chemistry. 272(3)

مصطلحات موضوعية: Enzyme complex, Apolipoprotein B, APOBEC-1 Deaminase, Mutant, Biology, Biochemistry, Adenoviridae, chemistry.chemical_compound, Mice, In vivo, Cytidine Deaminase, Animals, Humans, Luciferase, RNA, Messenger, Molecular Biology, Apolipoproteins B, Gene Transfer Techniques, Fast protein liquid chromatography, Cell Biology, Molecular biology, Lipoproteins, LDL, chemistry, Liver, RNA editing, Low-density lipoprotein, Mutation, biology.protein, Mutagenesis, Site-Directed, RNA Editing

الوصف: APOBEC-1 is a catalytic subunit of an apolipoprotein B (apoB) mRNA editing enzyme complex. In humans it is expressed only in the intestine, whereas in mice it is expressed in both the liver and intestine. APOBEC-1 exists as a spontaneous homodimer (Lau, P. P., Zhu, H.-J., Baldini, A., Charnsangavej, C., and Chan, L. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 8522-8526). We tested the editing activity and dimerization potential of three different mouse APOBEC-1 mutants using in vitro editing activity assay and immunoprecipitation in the presence of epitope-tagged APOBEC-1. One catalytically inactive mutant, mu1 (H61K/C93S/C96S), that retains its capacity to dimerize with wild-type APOBEC-1 was found to inhibit the editing activity of the latter and was thus a dominant negative mutant. Two other inactive mutants that dimerized poorly with APOBEC-1 failed to inhibit its activity. Intravenous injection of a mu1 adenovirus, Admu1, in C57BL/6J mice in vivo resulted in liver-specific expression of mu1 mRNA. On days 4 and 9 after virus injection, endogenous hepatic apoB mRNA editing was 23.3 +/- 5.0 and 36.8 +/- 5.7%, respectively, compared with 65.3 +/- 11 and 71.3 +/- 5.2%, respectively, for luciferase adenovirus-treated animals. Plasma apoB-100 accounted for 95 and 93% of total plasma apoB in Admu1 animals on days 4 and 9, respectively, compared with 78 and 72% in luciferase adenovirus animals. Plasma cholesterol on day 9 was 98 +/- 17 mg/dl in the mu1-treated animals, substantially higher than phosphate-buffered saline-treated (57 +/- 9 mg/dl) or luciferase-treated (71 +/- 12 mg/dl) controls. Fast protein liquid chromatography analysis of mouse plasma showed that the intermediate density/low density lipoprotein fractions in the animals treated with the dominant negative mutant adenovirus were much higher than those in controls. We conclude that active APOBEC-1 functions as a dimer and its activity is inhibited by a dominant negative mutant. Furthermore, apoB mRNA editing determines the availability of apoB-100, which in turn limits the amount of intermediate density/low density lipoprotein that can be formed in mice. Liver-specific inhibition of apoB mRNA editing is an important component of any strategy to enhance the value of mice as a model for human lipoprotein metabolism.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::3dc6f958e29e25e6d07e42e2a7e67bbbTest
https://pubmed.ncbi.nlm.nih.gov/8999814Test

المؤلفون: Mei-jin Chu, Kazuhiro Oka, Lawrence Chan, Merry Sullivan, M. Nakamuta, Kazumi Ishimura-Oka

المصدر: Genomics. 29(3)

مصطلحات موضوعية: Molecular Sequence Data, Biology, Polymerase Chain Reaction, Exon, Islets of Langerhans, Mice, Complementary DNA, Glucokinase, Genetics, Consensus sequence, Animals, Humans, Genomic library, Amino Acid Sequence, Gene, Peptide sequence, DNA Primers, Gene Library, Base Sequence, Sequence Homology, Amino Acid, cDNA library, Nucleic acid sequence, Molecular biology, Rats, Isoenzymes, Liver, Organ Specificity

الوصف: A complementary DNA for glucokinase (GK) was cloned from mouse liver total RNA by a combination of the polymerase chain reaction (PCR) and mouse liver cDNA library screening. Liver- and {beta}-cell-specific exons 1 were isolated by PCR using mouse and rat genomic DNAs. These clones were then used to screen a mouse genomic library; three genomic clones were isolated and characterized. The mouse GK gene spans over 20 kb, containing 11 exons including a liver- or {beta}-cell-specific exon 1, which encodes a tissue-specific 15-aa peptide at the N-terminus of the protein. Both types of GK contain 465 amino acid residues. The predicted amino acid sequence of mouse {beta}-cell-specific GK showed 98 and 96% identity to the rat and human enzymes, respectively; the corresponding values are 98 and 95% respectively, for the liver-specific GK. Several transcription factor-binding consensus sequences are identified in the 5{prime} flanking region of the mouse GK gene. 21 refs., 1 fig.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::d5c892c653f2079b5fccc9f7e722bb15Test
https://pubmed.ncbi.nlm.nih.gov/8575769Test

المؤلفون: Kazuhiro Oka, Lawrence Chan, Wen-Hsiung Li, Merry Sullivan, Kazumi Ishimura-Oka, Julia Krushkal, Mei-jin Chu

المصدر: European journal of biochemistry. 224(3)

مصطلحات موضوعية: Very low-density lipoprotein, Low-density lipoprotein receptor gene family, DNA, Complementary, Molecular Sequence Data, Very Low-Density Lipoprotein Receptor, VLDL receptor, Biology, Lipoproteins, VLDL, Biochemistry, Mice, Animals, Humans, Amino Acid Sequence, Cloning, Molecular, Peptide sequence, Phylogeny, Genetics, Base Sequence, cDNA library, Biological Evolution, Cell biology, Receptors, LDL, LDL receptor, lipids (amino acids, peptides, and proteins), Rabbits, Energy source

الوصف: The very-low-density-lipoprotein receptor (VLDLR) is a recently described lipoprotein receptor that shows considerable similarity to the low-density-lipoprotein receptor (LDLR). This receptor has been suggested to be important for the metabolism of apoprotein-E-containing triacylglycerol-rich lipoproteins, such as very-low-density-lipoprotein (VLDL), beta-migrating VLDL and intermediate-density lipoprotein. cDNA clones that code for the VLDLR were isolated from a mouse heart cDNA library. The deduced amino acid sequence predicts a mature protein of 846 amino acids preceded by a 27-residue signal peptide. Three mRNA species for the VLDLR with sizes of 3.9, 4.5 and 7.9 kilobases were present in high concentration in heart and muscle, which utilize triacylglycerols as an energy source. VLDLR mRNA is also detected in decreasing amounts in kidney, brain, ovary, testis, lung and adipose tissue. It is essentially absent in liver and small intestine. The amino acid sequence of the VLDLR is highly conserved among rabbit, human and mouse. VLDLR contains five structural domains very similar to those in LDLR, except that the ligand-binding domain in VLDLR has an eightfold repeat instead of a sevenfold repeat in LDLR. Sequence conservation among animal species is much higher for the VLDLR than the LDLR. Sequences of the VLDLR from three vertebrate species and the LDLR from five vertebrate species were aligned and a phylogenetic tree was reconstructed. Although both receptors contain five domains and share amino acid sequence similarity, our computations showed that they diverged before the divergence between mammals and amphibians. In addition, sequence comparison of both receptor sequences suggests that the rabbit is evolutionarily closer to man than to the mouse. These results are consistent with the hypothesis that the VLDLR and the LDLR have evolved from a common ancestral gene to play distinct roles in lipoprotein metabolism and that the metabolic handling of triacylglycerol by the body via the VLDLR is a highly conserved mechanism.

الوصول الحر: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::4f68d897e9858f9675fde0962049b992Test
https://pubmed.ncbi.nlm.nih.gov/7925422Test