Search PubMed⌕ Search

Biomedical subjects

H Li

Publications and source records attributed to H Li.

At least 1,387 records · Page 77Linked to original sources
PubMed1996

Induction of phosphoglycerate kinase 1 gene expression by hypoxia. Roles of Arnt and HIF1alpha.

To identify new dimerization partners for the aromatic hydrocarbon receptor nuclear translocator (Arnt), we used its N-terminal region (amino acids 1-470) as a target in a two-hybrid screening procedure, and we cloned the murine form of hypoxia-inducible factor 1alpha (HIF1alpha). Sequence comparisons reveal substantial identity between mouse and human HIF1alpha. Hypoxia induces a 10-fold accumulation of phosphoglycerate kinase 1 mRNA in wild type mouse hepatoma (Hepa 1c1c7) cells; the induction mechanism is Arnt dependent because induction does not occur in Arnt-defective cells. Furthermore, induction of phosphoglycerate kinase 1 mRNA requires Arnt's N-terminal region, which mediates DNA binding and heterodimerization; in contrast, induction does not require Arnt's C-terminal region, which mediates transactivation. We also show that a GAL4-HIF1alpha fusion protein transactivates a GAL4-dependent gene in the absence of Arnt, that HIF1alpha's transactivation capability is inducible by hypoxia, and that both hypoxia responsiveness and transactivation capability reside within the C-terminal 83 amino acids of HIF1alpha. Our findings generate new insights into the mechanism by which Arnt and HIF1alpha induce transcription in response to hypoxia.

Amino Acid Sequence↗
PubMed1996

Identification of novel regions of deletion in familial Wilms' tumor by comparative genomic hybridization.

Wilms' tumor, an embryonic renal neoplasm diagnosed primarily in young children, can occur in either a noninheritable (sporadic) or a familial form, with the latter presenting earlier and more often at bilateral sites. Although familial Wilms' tumor is thought to develop through inherited and acquired mutational inactivation of the two alleles of predisposing tumor suppressor genes, only a small percentage of cases can be accounted for by mutations affecting the WT1 gene or linkage to the Beckwith-Weidemann syndrome of the BWS region on the short arm of chromosome 11. To find chromosomal regions that might contain genes important in the development of this disease, we used comparative genomic hybridization to analyze tumor specimens from familial cases for chromosomal regions that were consistently lost. Although inherited lesions of tumor suppressors are most often inactivating point mutations, accompanying somatic lesions in the malignant clones are often chromosomal deletions; therefore, consensus regions of loss in familial tumors are likely to harbor genes linked to familial predisposition. There were extensive genomic aberrations among the eight familial cases studied, with an average of 6.5 changes/tumor (range, 0-22). The most consistent findings with likely biological relevance were deletions of chromosomes 4 (consensus, 4q21-qter), 9 (consensus, 9p21-pter), 20p, and 3 (consensus, 3q12-q21). These regions have not been previously implicated in Wilms' tumor and may harbor novel genes that could aid attempts to understand the familial predisposition as well as the development and progression of these tumors.

Chromosome Deletion↗
PubMed1996

Alternatively spliced cyclin C mRNA is widely expressed, cell cycle regulated, and encodes a truncated cyclin box.

The cyclin C protein has recently been shown to associate with a unique cyclin dependent protein kinase (cdk8) and it has been proposed that this complex may regulate RNA transcription during the cell cycle. In addition, the human cyclin C gene has been localized to human chromosome 6q21 and it was found to be frequently deleted in a subset of acute lymphoblastic leukemias (ALL's). Screening of an avian T-cell cDNA library resulted in the isolation of a cyclin C homologue as well as an abundant, yet distinct, cyclin C-related cDNA. The predicted open reading frame (ORF) of the cyclin C cDNA predicted a 283 amino acid protein that was > 99% identical to the human protein and 72% identical to the Drosophila melanogaster protein. However, the predicted ORF of the cyclin C-related cDNA predicted a much smaller 105 amino acid protein that was identical to cyclin C well into the cyclin-box region (amino acid residue 98), where it abruptly diverges and then terminates. Using PCR analysis of cDNA derived from a range of cell lines and tissues, alternative splicing of the avian cyclin C gene has been demonstrated. Furthermore, a smaller approximately 19 kDa protein that co-migrates with the in vitro transcribed and translated truncated cyclin C protein was detected in normal and virally-transformed avian cells with a cyclin C-specific antibody. Expression of alternatively spliced cyclin C mRNA and protein is regulated in a cell cycle-dependent manner reminiscent of cyclin B2. The function of this truncated cyclin C protein is not known, but its expression in avian cells suggest that this truncated cyclin C protein may participate as an early endogenously encoded cyclin C inhibitor.

Alternative Splicing↗
PubMed1996

Emergence of preferred structures in a simple model of protein folding.

Protein structures in nature often exhibit a high degree of regularity (for example, secondary structure and tertiary symmetries) that is absent from random compact conformations. With the use of a simple lattice model of protein folding, it was demonstrated that structural regularities are related to high "designability" and evolutionary stability. The designability of each compact structure is measured by the number of sequences that can design the structure-that is, sequences that possess the structure as their nondegenerate ground state. Compact structures differ markedly in terms of their designability; highly designable structures emerge with a number of associated sequences much larger than the average. These highly designable structures possess "proteinlike" secondary structure and even tertiary symmetries. In addition, they are thermodynamically more stable than other structures. These results suggest that protein structures are selected in nature because they are readily designed and stable against mutations, and that such a selection simultaneously leads to thermodynamic stability.

Amino Acid Sequence↗
PubMed1996

The production of macrophage inflammatory protein-1 alpha by human basophils.

Macrophage inflammatory protein-1alpha (MIP-1alpha) has previously been shown to be produced by mononuclear cells, eosinophils, and neutrophils. Its production by basophils has not been investigated. The objective of this study was to investigate the production of MIP-1alpha by basophils. Peripheral blood basophils were separated by Percoll gradient centrifugation, cultured overnight, and processed for double immunocytochemistry using Abs against MIP-1alpha and FcepsilonRIalpha (alpha subunit of IgE receptor type 1). We demonstrated that basophils expressed immunoreactive MIP-1alpha upon stimulation with anti-IgE. Less than 5% of the basophils stained for MIP-1alpha without stimulation. The secretion of MIP-1alpha by basophils was studied by ELISA. In these experiments, basophils were further enriched to 65 to 99% (median, 86%) by a negative selection method. Basophils released MIP-1alpha when stimulated by Abs against IgE and FCepsilonRIalpha as well as IL-3 and the calcium ionophore, A23187. In parallel experiments, PBMC, eosinophils, and neutrophils did not produce MIP-1alpha in response to anti-IgE, but they did so in response to A23187. No MIP-1alpha release was detected in platelet preparations. Preincubation with IL-3 (15 min or 18 h) augmented anti-IgE-included basophil MIP-1alpha production. The secretion of MIP-1alpha by basophils was detectable shortly after stimulation and gradually increased over 24 h. Since MIP-1alpha has potent inflammatory and histamine-releasing activities, its production by basophils may indicate a positive feedback mechanism for allergic inflammation.

Antibodies, Anti-Idiotypic↗
PubMed1996

Malignant catarrhal fever virus. Characterization of a United States isolate and development of diagnostic assays.

Malignant catarrhal fever (MCF) is a severe lymphoproliferative disease of certain domestic and wild ruminants. Two distinct but closely related viruses cause clinically indistinguishable syndromes in susceptible ruminant species: wildebeest-associated MCF virus (WA-MCFV) and sheep-associated MCF virus (SA-MCFV). Neither the pathogenesis nor the epidemiology of SA-MCF is understood, primarily because of a lack of adequate detection methods for the etiologic agent or antibody against that agent. Work designed to develop these tests has been under way in our laboratory. To obtain basic information about the virus, the in vitro growth properties of a US isolate of MCF virus were studied and its major viral proteins identified and characterized by a panel of monoclonal antibodies generated against the isolate. A monoclonal antibody to a broadly conserved epitope of MCF virus was identified, and a competitive-inhibition ELISA (CI-ELISA) was developed for detection of anti-MCF antibody in sheep and other ruminants. The monoclonal antibody (15-A) reacted with an epitope located on a glycoprotein complex, which was present in all isolates of MCF virus examined. Antibody from a wide variety of ruminants infected with MCF virus of both sheep and wildebeest origin competed with the monoclonal antibody 15-A for the epitope, which was not present on 14 other common ruminant viruses. The assay detected antibody in inapparently infected sheep, and in cattle, deer, and bison with clinical MCF. A PCR assay for DNA of the sheep-associated virus was developed, based on previously reported primers. Comparative studies demonstrated that the CI-ELISA was specific for MCFV antibody and that the PCR was more reliable for diagnosis of clinical MCF.

Animals↗
PubMed1996

The relation between codon usage, base correlation and gene expression level in Escherichia coli and yeast.

Based on the investigation of the relation between gene expression and the usage of synonymous codons, a method of classifying and predicting the gene expression level is proposed which is called the Self-consistent Information Clustering (SCIC). Using the modified Codon Adaption Index (CAI) values, we have accomplished the linear regression analysis on the relation between base composition, base correlation and gene expression level in Escherichia coli and yeast. The assumption of Expression-Enhancing-Network Site (EENS) is proposed, the existence of which can be demonstrated by the linear equations between gene expression and base correlations in a codon, in adjacent codons and in non-adjacent codons. The modes of base correlation of E. coli and yeast which are important to gene expression have been found and listed in this paper.

Base Sequence↗
PubMed1996

Cloning of PL33: a novel probable serpentine membrane receptor associated with human cytotrophoblast and lineage-specific HL-60 cell differentiation.

From a subtracted library of differentiating human cytotrophoblast cells, we have cloned a gene termed PL33, a novel probable member of the G-protein-linked seven transmembrane domain receptor family. The 496 amino acid open reading frame of PL33 codes for a protein of predicted molecular weight of 54.7 kDa. Northern blot analysis shows that the two mRNA transcripts of PL33 are expressed in testes, keratinocytes, differentiating cytotrophoblast and in monocyte/macrophage lineage-specific HL-60 cell differentiation.

Amino Acid Sequence↗
PubMed1996

Sex- and tissue-specific expression of a cytochrome P450 2C2-luciferase transgene.

To study the transcriptional regulation of the rabbit cytochrome P450 2C2 gene (CYP2C2), transgenic mice were generated using a DNA fragment containing the CYP2C2 5'-flanking region fused to a luciferase reporter gene. In one of three transgenic lines, this transgene was expressed in the liver and at a lower level in the kidney, similar to the tissue-specific pattern of this gene in rabbits. Unlike expression in rabbits, the transgene was expressed in a male-specific pattern in liver and kidney, and in the brain of both sexes. Expression of the transgene in male mice was repressed after castration, while expression in females was stimulated after testosterone treatment and periodic injection of rat growth hormone (GH). The expression of this transgene was also regulated developmentally in the liver. These results suggest that this 3500 bp of CYP2C2 5'-flanking region contains the basal transcriptional regulation elements for tissue-specific and developmental expression. Although a position effect can not be ruled out, this transgene may also include the cis-regulatory element for sexual dimorphic expression.

Animals↗
PubMed1996

IL-13 released by and localized in human basophils.

We and others have shown that human basophils can synthesize and release IL-4. However, IL-13, a cytokine that closely resembles IL-4, has not hitherto been described as a basophil product. The production of IL-13 by basophils was demonstrated by immunocytochemistry. Approximately 70% of basophils stimulated with anti-FcepsilonRIalpha (antibody to the alpha subunit of IgE receptor type I) stained for IL-13. Under similar experimental conditions, mononuclear cells failed to stain for IL-13. The cytokine was localized to basophilic granules by electron microscopic examination of immunogold staining. The secretion of IL-13 into the culture supernatant was assayed by ELISA. Kinetic studies showed detectable IL-13 release at 3 h, which steadily increased up to 24 h. This is significantly different from the kinetics of basophil histamine and IL-4 release. IL-13 production was also observed upon stimulation with anti-IgE, anti-FcepsilonRIalpha, IL-3, and A23187 in a dose-dependent manner. PBMC, neutrophils, and eosinophils isolated from the same donors did not release IL-13 after anti-IgE stimulation. The anti-IgE-induced basophil IL-13 synthesis could be enhanced by IL-3 preincubation (with and without IL-3 preincubation, anti-IgE-induced IL-13 production was 227 +/- 99 and 42 +/- 13 pg/10(6) basophils, respectively). PBMC produced a significant amount of IL-1 3 upon stimulation with PHA, but a low level of IL-13 in response to A23187 and/or PMA. Eosinophils and neutrophils did not produce IL-13 when cultured with A23187, IL-5, and anti-FcepsilonRIalpha. This is the first demonstration of IL-1 3 production by basophils. Our data suggest that basophils, in addition to secreting mediators, can represent an important source of proallergic cytokines.

Basophils↗
PubMed1996

Two-peak kinetic curve of the chemiluminescence in phorbol-induced macrophage.

Phorbol-1,2-myristate-1,3-acetate can induce rat macrophage to emit chemiluminescence. The kinetic curve of the chemiluminescence clearly showed two peaks when a proper ratio of phorbol-1,2-myristate-1,3-acetate and macrophage were mixed. It was proved that the first one mainly came from the oxygen free radicals and the second one from NO generated in the phorbol-1,2-myristate-1,3-acetate stimulated cells by using the substitute and inhibitor of NO synthase.

Animals↗
PubMed1996

Characterization of the denaturation and renaturation of human plasma vitronectin. II. Investigation into the mechanism of formation of multimers.

Unfolding and refolding of plasma vitronectin appear irreversible under near physiological conditions, with rearrangements of disulfides and self-association to a multimeric form observed as prominent structural alterations which accompany denaturation. A mechanism for the folding reactions of vitronectin has been proposed (Zhuang, P., Blackburn, M. N., and Peterson, C. B.(1996) J. Biol. Chem. 270, 14323-14332) in which vitronectin acquires a partially folded intermediate structure which is highly prone to oligomerize into a multimeric form. Strongly oxidizing conditions adopted for refolding from urea were effective at preventing disulfide rearrangement which disrupts distal disulfides near the C terminus of the protein. Prohibiting disulfide rearrangement under these conditions, however, was not sufficient to achieve reversibility in folding. In contrast, variations in the ionic strength of the refolding medium affect the partitioning of species so that refolded monomers are obtained at high ionic strength, and self-association is precluded. The effects of ionic strength on the partially folded intermediate in the vitronectin folding pathway appear to favor intramolecular hydrophobic collapse to form a stable hydrophobic core for the monomer versus intermolecular hydrophobic interactions which stabilize multimeric vitronectin. Although both ionic and hydrophobic interactions presumably contribute to subunit interfaces within the multimer, the basic heparin-binding region near the C terminus of the protein does not provide binding interactions which are important for self-association of vitronectin.

Amino Acid Sequence↗
PubMed1996

Magnetic resonance imaging (MRI) detection of the murine brain response to light: temporal differentiation and negative functional MRI changes.

Using a 9.4 T MRI instrument, we have obtained images of the mouse brain response to photic stimulation during a period between deep anesthesia and the early stages of arousal. The large image enhancements we observe (often >30%) are consistent with literature results extrapolated to 9.4 T. However, there are also two unusual aspects to our findings. (i) The visual area of the brain responds only to changes in stimulus intensity, suggesting that we directly detect operations of the M visual system pathway. Such a channel has been observed in mice by invasive electrophysiology, and described in detail for primates. (ii) Along with the typical positive response in the area of the occipital portion of the brain containing the visual cortex, another area displays decreased signal intensity upon stimulation.

Animals↗
PubMed1996

Base-boronated dinucleotides: synthesis and effect of N7-cyanoborane substitution on the base protons.

Boron-modified nucleic acids comprise a new set of DNA mimics that have potential biological and therapeutic applications. A series of nine dinucleotides containing N7-cyanoborane-2'-deoxyguanosine ((7b)dG) at the 3', 5' or both positions of the phosphodiester linkage have been synthesized using solution phase phosphoramidite chemistry. Fmoc was used as the 5'-protecting group because of incompatibility of the cyanoborane moiety with 5'-DMT cations generated during the deprotection step. The presence of the cyanoborane group was confirmed on the basis of Fab-MS and 1H NMR spectroscopy. The H-8 proton of (7b)dG in the dinucleotides shifted 0.35-0.80 p.p.m. downfield relative to that of unmodified dG. A comparison of the D20 exchange kinetics of the H-8 proton at 60 degrees C showed that H-8 of (7b)dG is very labile relative to unmodified dG, indicating that the N7-cyanoborane modification increases the acidity of the H-8 proton of (7b)dG. These studies illustrate the feasibility of synthesizing boron-containing oligonucleotides which are modified at the N7-guanine to block Hoogsteen pairing in the DNA major groove.

Amides↗
PubMed1996

Monocyte chemotactic protein 4 (MCP-4), a novel structural and functional analogue of MCP-3 and eotaxin.

A novel human CC chemokine complementary DNA was identified in a library constructed from human fetal RNA, cloned into a baculovirus vector, and expressed in Sf9 insect cells. The mature recombinant protein that was released had the NH2-terminal sequence pyro-QPDALNVPSTC...and consisted of 75 amino acids. Minor amounts of two variants of 77 and 82 residues (NH2 termini: LAQPDA...and FNPQGLAQPDA...) were released as well. The novel chemokine was designated monocyte chemotactic protein 4 (MCP-4) and the variants were designated (LA)MCP-4 and (FNPQGLA)MCP-4. MCP-4 shares the pyroglutamic acidproline NH2-terminal motif and 56-61% sequence identity with the three known monocyte chemotactic proteins and is 60% identical to eotaxin. It has marked functional similarities to MCP-3 and eotaxin. Like MCP-3, MCP-4 is a chemoattractant of high efficacy for monocytes and T lymphocytes. On these cells, it binds to receptors that recognize MCP-1, MCP-3, and RANTES. On eosinophils, MCP-4 has similar efficacy and potency as MCP-3, RANTES, and cotaxin. It shares receptors with eotaxin and shows full cross-desensitization with this cosinophil-selective chemokine. Of the two variants, only (LA)MCP-4 could be purified in sufficient quantities for testing and was found to be at least 30-fold less potent than MCP-4 itself. This suggests that the 75-residue form with the characteristic NH2 terminus of an MCP is the biologically relevant species.

Acetylglucosaminidase↗
PubMed1996

Unexpectedly strong binding of a large metal ion (Bi3+) to human serum transferrin.

Large metal ions (>0.9 A ionic radius) have previously been found to bind only weakly to human serum transferrin (hTF, 80 kDa), presumably because the interdomain cleft cannot close around the metal and synergistic anion. Surprisingly, therefore, we report that Bi3+ (ionic radius 1.03 A), a metal ion widely used in anti-ulcer drugs, binds strongly to both the N- and C-lobes with log K1* = 19.42 and log K2* = 18.58 (10 mM Hepes, 5 mM bicarbonate, 310 K). The uptake of Bi3+ by apo-hTF from bismuth citrate complexes is very slow (hours), whereas that from bismuth nitrilotriacetate is rapid (minutes). Evidence from absorption and NMR spectroscopy is presented to show that Bi3+ binds to the specific Fe3+ binding sites along with carbonate as the synergistic anion. Under the conditions used, preferential binding of Bi3+ to the C-lobe of hTF is observed. Linear free energy relationships show that there is a strong correlation between the strength of binding of Bi3+ and Fe3+ to a wide variety of ligands which include transferrin. Therefore we conclude that the strength of metal ion binding to transferrin is determined more by the ligand donor set than by the size of the ion.

Bismuth↗
PubMed1996

Multiplex genotype determination at a large number of gene loci.

To facilitate large-scale genotype analysis, an efficient PCR-based multiplex approach has been developed. For simultaneously amplifying the target sequences at a large number of genetic loci, locus-specific primers containing 5' universal tails are used. Attaching the universal tails to the target sequences in the initial PCR steps allows replacement of all specific primers with a pair of primers identical to the universal tails and converts the multiplex amplification into "uniplex." Simultaneous amplification of 26 genetic loci with this approach is described. The multiplex amplification can be coupled with genotype determination. By incorporating a single-base mismatch between a primer and the template into the target sequences, a polymorphic site can be converted into a desirable restriction fragment length polymorphism when it is necessary. In this way, the allelic PCR products for the polymorphic loci can be discriminated by gel electrophoresis after restriction enzyme digestion. In this study, 32 loci were typed in such a multiplex way.

Alleles↗