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W Zhang

Publications and source records attributed to W Zhang.

At least 199 records · Page 11Linked to original sources
PubMed2001

The novel peptide apelin lowers blood pressure via a nitric oxide-dependent mechanism.

Apelin is an endogenous ligand of the human orphan receptor APJ. We detected apelin-like immunoreactivity in the adipocytes, gastric mucosa, and Kupffer cells in the liver. We also detected apelin-like immunoreactivity localized within the endothelia of small arteries in various organs. Further, it was found that mean arterial pressure after the administration of apelin-12, apelin-13, and apelin-36 at a dose of 10 nmol/kg in anaesthetized rats was reduced by 26+/-5, 11+/-4, and 5+/-4 mm Hg, respectively. In the presence of a nitric oxide (NO) synthase inhibitor, the effect of apelin-12 on blood pressure was abolished. Furthermore, the administration of apelin-12 (10 nmol/kg) in rats produced a transitory elevation of the plasma nitrite/nitrate concentration from a basal level of 21.4+/-1.6 to 27.0+/-1.5 microM. Thus, apelin may lower blood pressure via a nitric oxide-dependent mechanism.

Animals↗
PubMed2001

[Molecular mechanism underlying calcium handling in diastolic heart failure].

OBJECTIVE: To elucidate the molecular mechanism underlying calcium handling in diastolic heart failure (DHF) from mRNA level and protein expression, including L-type calcium channel, sarcoplasmic reticulum (SR) Ca(2+)-ATPase, phospholamban, ryanodine receptor, calsequestrin. METHODS: DHF was produced in rabbits by abdominal aortic coarctation. The mRNA amounts of these calcium-handling genes were measured by RT-PCR, while the protein levels of SR Ca(2+)-ATPase and phospholamban were analyzed by Western blot analysis. RESULTS: The content of calcium was significantly increased in myocardium of rabbits with DHF than in the myocardium of sham-operated rabbits. The SR Ca(2+)-ATPase activity of DHF rabbits was significantly reduced compared with that in sham-operated rabbits (21.1 micromol.mg(-1).h(-1) +/- 5.7 micromol.mg(-1).h(-1) vs 10.5 micromol.mg(-1).h(-1) +/- 2.8 micromol.mg(-1).h(-1), P < 0.01). RT-PCR analyses showed that the steady-state level of mRNA encoding the L-type calcium channel and SR Ca2+-ATPase was decreased significantly in rabbits with DHF compared with that in the sham-operated rabbits (micromol.mg(-1).h(-1)): 0.75 +/- 0.11 vs 1.20 +/- 0.33; 0.76 +/- 0.12 vs 1.24 +/- 0.38, P < 0.05). The SR Ca(2+)-ATPase mRNA level correlated negatively well with left ventricular relaxation time constant and left ventricular end-diastolic pressure (r = -0.81, -0.64, respectively, P < 0.05 approximately 0.01); the mRNA level of L-type calcium channel correlated negatively with left ventricular end-diastolic pressure (r = -0.74, P < 0.05). The mRNA level of ryanodine receptor correlated negatively with the left ventricular relaxation time constant too (r = -0.71, P < 0.05). Protein level of SR Ca(2+)-ATPase was significantly lower in rabbits with DHF than in the sham-operated rabbits (0.76 +/- 0.6 vs 1.02 +/- 0.09, P < 0.05), whereas the protein level of phospholamban was unchanged. CONCLUSION: The L-type calcium channel and SR Ca(2+)-ATPase were down regulated in DHF. These changes may be a contributory factor for DHF.

Animals↗
PubMed2001

An eukaryotic-type serine/threonine protein kinase involved in the carbon source-dependent pigment biosynthesis in Amycolatopsis mediterranei U32.

The structural gene, pkmA, was cloned and sequenced from a rifamycin SV-producing Amycolatopsis mediterranei U32 strain. The N-terminal portion of the deduced amino acid sequence of pkmA showed significant similarity to the family of serine/threonine protein kinases. It contains all the structural features which are highly conserved in protein kinases, including the Gly-X-Gly-X-X-Gly motif of ATP binding and the essential amino acids known to be important for the recognition of the correct hydroxyamino acid in serine/threonine protein specific kinases. The protein possesses a region rich in Ala and Pro residues around the middle of pkmA open reading frame, which might be involved in the transmembrane function, as suggested by PhoA fusion protein analysis. The pkmA gene was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein, and the protein was found to have the activity of autophosphorylation. A double crossover gene replacement was achieved by inserting an aparmycin resistance gene into pkmA in A. mediterranei chromosomal DNA. The phenotypic analysis of the mutant suggested that pkmA gene is involved in carbon source-dependent pigment formation in A. mediterranei U32.

Alkaline Phosphatase↗
PubMed2001

Beta(3)-integrin-deficient mice but not P-selectin-deficient mice develop intimal hyperplasia after vascular injury: correlation with leukocyte recruitment to adherent platelets 1 hour after injury.

BACKGROUND: Intimal hyperplasia contributes to restenosis after percutaneous vascular interventions. Both beta(3)-integrins, alpha(V)beta(3) and alpha(IIb)beta(3) (glycoprotein IIb/IIIa), and leukocytes have been implicated in neointimal formation, based in part on the results obtained using antagonists to 1 or both receptors in animal models. METHODS AND RESULTS: The responses in wild-type mice, beta(3)-integrin-deficient mice, and P-selectin-deficient mice were studied in a model of transluminal endothelial injury of the femoral artery. At 4 weeks, beta(3)-integrin-deficient mice were not protected from developing intimal hyperplasia, whereas P-selectin-deficient mice were protected. Within 1 hour of injury, several layers of platelets deposited on the arteries of wild-type mice and a single layer of platelets deposited on the vessels of beta(3)-integrin-deficient mice; in both cases, leukocytes were recruited to the platelet layer. In P-selectin-deficient mice, the platelet layer was less compact and extended further into the lumen but did not recruit leukocytes. CONCLUSIONS: In a model of transluminal arterial injury, absence of early leukocyte recruitment and not deficiency of beta(3)-integrins correlated with a reduction in neointimal formation. Blockade of P-selectins may be an effective therapeutic strategy to decrease restenosis after percutaneous vascular interventions.

Animals↗
PubMed2001

Enhanced reporter gene expression in the rat brain from helper virus-free HSV-1 vectors packaged in the presence of specific mutated HSV-1 proteins that affect the virion.

Herpes simplex virus (HSV-1) gene expression is hypothesized to shut off promoters in HSV-1 vectors, but in a helper virus-free HSV-1 vector system, a number of promoters support only short-term expression. Thus, recombinant gene expression remains short-term in the absence of approximately 99% of the HSV-1 genome. To resolve this paradox, we hypothesized that specific HSV-1 proteins that affect the virion can shut off recombinant gene expression. This study evaluated expression from HSV-1 vectors, containing neuronal-specific promoters, that were packaged in the presence of specific mutated HSV-1 proteins that affect the virion. The mutated HSV-1 proteins that were examined included two protein kinases (U(L)13 and U(S)3), the virion host shut-off factor (vhs), the transactivator of immediate early promoters (VP16), and a virion protein that affects RNA metabolism (U(S)11). Helper virus-free packaging could occur in the presence of each mutated protein alone or specific combinations of two or three mutated proteins. In BHK and PC12 cells, vectors packaged in the presence of each mutated protein increased ( approximately 2-fold) the level of expression per cell, and vectors packaged in the presence of specific combinations of mutated proteins supported larger (4-7-fold) increases. In the rat striatum, vectors packaged in the presence of a mutated U(S)3 displayed enhanced gene transfer (13-18-fold increases in the number of cells at 4 days), and vectors packaged in the presence of mutated U(L)13 or VP16 enhanced long-term expression (2 months). Vectors packaged in the presence of mutated vhs or U(S)11 displayed minimal changes in expression.

Animals↗
PubMed2001

The JIL-1 tandem kinase mediates histone H3 phosphorylation and is required for maintenance of chromatin structure in Drosophila.

To analyze the function of the chromosomal kinase JIL-1, we generated an allelic series of hypomorphic and null mutations. JIL-1 is an essential kinase for viability, and reduced levels of JIL-1 kinase activity lead to a global change in chromatin structure. In JIL-1 hypomorphs, euchromatic regions of polytene chromosomes are severely reduced and the chromosome arms condensed. This is correlated with decreased levels of histone H3 Ser10 phosphorylation. These levels can be restored by a JIL-1 transgene placing JIL-1 directly in the pathway mediating histone H3 phosphorylation. We propose a model where JIL-1 kinase activity is required for maintaining the structure of the more open chromatin regions that facilitate gene transcription.

Alleles↗
PubMed2001

Cloning and characterization of Siglec-10, a novel sialic acid binding member of the Ig superfamily, from human dendritic cells.

The Siglecs (sialic acid-binding Ig-like lectins) are a subfamily of I-type lectins, which specifically recognize sialic acids. Nine members of the family have been identified thus far. We have obtained a novel cDNA clone from a human dendritic cell cDNA library encoding a protein with sequence and structural features of the Siglec family, hence designated as Siglec-10. The full-length Siglec-10 cDNA encodes a type 1 transmembrane protein containing four extracellular immunoglobulin-like domains, a transmembrane region, and a cytoplasmic tail with two classical immunoreceptor tyrosine-based inhibitory motifs. The N-terminal V-set Ig domain has most of the amino acid residues typical of the Siglecs. Siglec-10 shows the closest homology to Siglec-5 and Siglec-3/CD33. Various cells and cell lines including monocytes and dendritic cells express Siglec-10. High levels of mRNA expression were seen in peripheral blood leukocytes, spleen, and liver. When expressed on COS-7 cells, Siglec-10 was able to bind human red blood cells and soluble sialoglycoconjugates in a sialic acid-dependent manner. The identification of Siglec-10 as a new Siglec family member and its expression profile, together with its sialic acid-dependent binding capacity, suggest that it may be involved in cell-cell recognition by interacting with sialylated ligands expressed on specific cell populations.

Amino Acid Motifs↗
PubMed2001

EGR-1, a UV-inducible gene in p53(-/-) mouse cells.

Changes in gene expression were examined in p53(-/-) and p53(+/+) mouse cells after ultraviolet (UV) irradiation. Differential display was used to identify differentially expressed gene(s) in UV-treated p53(-/-) and p53(+/+) cells. One of the differentially expressed genes was EGR-1 (early growth response gene-1), which was shown to be induced only in p53(-/-) cells. The induction of this gene by UV was detected as early as 0.5 h, peaked at 2 h, and returned to normal levels by 4 h. De novo protein synthesis was not required for UV-induced EGR-1 expression in p53(-/-) cells. Pretreatment of p53(-/-) cells with suramin, an inhibitor of growth factor receptors, completely suppressed UV-induced EGR-1 expression, suggesting that the induction may be mediated via the growth factor receptors. The presence of wild-type p53 suppressed the induction of EGR-1 after UV treatment. Overexpression of EGR-1 promoted the UV-induced transformation in p53(+/+) cells, but not in p53(-/-) cells. These data suggested that EGR-1 may be an important player in the UV responses of mammalian cells and may influence UV-induced transformation.

Animals↗
PubMed2001

Resetting baroreceptors to a lower arterial pressure level by enalapril avoids baroreflex mediated activation of sympathetic nervous system by nifedipine.

Baroreceptor-unloading-mediated activation of sympathetic nervous system (SNS) by antihypertensive agents, such as dihydropyridine calcium channel blockers (CCB), has been considered to compromise the beneficial effects of the therapy and lead to unsatisfying clinical outcome. The present study was aimed at finding a novel way of using CCB without activating SNS. In anaesthetized Wistar rats, baroreceptor-unloading-mediated reflex activation of SNS, as indicated by tachycardia and increase of plasma catecholamines, was observed after mean arterial pressure (MAP) was decreased by 15 mmHg during 4-h administration of nifedipine, a CCB. However an angiotensin-converting enzyme inhibitor (ACEI), enalapril did not cause tachycardia or increase plasma catecholamine levels when it decreased MAP by 15 mmHg. After 100 min (supposedly baroreceptor resetting or adaptation to hypotension had occurred), enalapril infusion was gradually replaced by nifedipine infusion in 40 min. Nifedipine was infused for another 100 min, which kept the lowered MAP unchanged and did not activate SNS. In anaesthetized spontaneously hypertensive rats (SHR), baroreceptor-mediated reflex activation of SNS was observed after MAP was decreased by 25 mmHg during 4-h nifedipine administration. However enalapril did not cause tachycardia or increase plasma catecholamine levels when it decreased MAP by 25 mmHg. After 100 min, enalapril infusion was gradually replaced by nifedipine infusion in 40 min. Nifedipine was then infused for another 100 min, which kept the lowered MAP unchanged and did not activate SNS. The present study indicated that reflex activation of SNS caused by antihypertensive effect of CCB could be avoided if, prior to CCB administration, baroreceptors have been reset to a lower MAP by a drug that does not activate baroreceptor reflex.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2001

Impaired modulation of sympathetic vasoconstriction in contracting skeletal muscle of rats with chronic myocardial infarctions: role of oxidative stress.

Skeletal muscle perfusion during exercise is impaired in heart failure, but the underlying mechanisms are poorly understood. One possibility is that sympathetic vasoconstriction is enhanced in exercising muscle in heart failure as a result of impaired counterregulatory mechanisms that normally act to attenuate vasoconstrictor responses. In healthy animals, sympathetic vasoconstriction in contracting skeletal muscle is attenuated by endogenously produced nitric oxide (NO). Because the NO pathway may be dysfunctional in heart failure, we hypothesized that reduced NO in contracting muscle would result in enhanced sympathetic vasoconstriction. In sham rats and rats with chronic myocardial infarctions (MIs) produced by coronary artery ligation, we measured arterial pressure and femoral artery blood flow responses to sympathetic nerve stimulation (1, 2.5, and 5 Hz) in resting and contracting hindlimb. In resting hindlimb, sympathetic stimulation decreased femoral vascular conductance similarly in sham and MI rats. In contracting hindlimb, these vasoconstrictor responses were attenuated to a greater extent in sham than in MI rats. NO synthase inhibition enhanced sympathetic vasoconstriction in contracting hindlimb of sham, but not MI, rats. Conversely, infusion of L-arginine or a superoxide scavenger, tempol or tiron, attenuated sympathetic vasoconstriction in contracting hindlimb of MI rats. NO synthase expression was similar, but malondialdehyde (a marker of free radical damage) was greater in skeletal muscle from MI than from sham rats. These data suggest that impaired metabolic modulation of sympathetic vasoconstriction in contracting skeletal muscle of MI rats is a consequence of superoxide-mediated disruption of the NO pathway.

Animals↗
PubMed2001

The optimal measure of allelic association.

Allelic association between pairs of loci is derived in terms of the association probability rho as a function of recombination theta, effective population size N, linear systematic pressure v, and time t, predicting both rho(rt), the decrease of association from founders and rho(ct), the increase by genetic drift, with rho(t) = rho(rt) + rho(ct). These results conform to the Malecot equation, with time replaced by distance on the genetic map, or on the physical map if recombination in the region is uniform. Earlier evidence suggested that rho is less sensitive to variations in marker allele frequencies than alternative metrics for which there is no probability theory. This robustness is confirmed for six alternatives in eight samples. In none of these 48 tests was the residual variance as small as for rho. Overall, efficiency was less than 80% for all alternatives, and less than 30% for two of them. Efficiency of alternatives did not increase when information was estimated simultaneously. The swept radius within which substantial values of rho are conserved lies between 385 and 893 kb, but deviation of parameters between measures is enormously significant. The large effort now being devoted to allelic association has little value unless the rho metric with the strongest theoretical basis and least sensitivity to marker allele frequencies is used for mapping of marker association and localization of disease loci.

Alleles↗
PubMed2001

Identification and characterization of DPZF, a novel human BTB/POZ zinc finger protein sharing homology to BCL-6.

The C2H2 zinc finger protein family is one of the largest families of transcription factors. We identified a novel BTB/POZ zinc finger gene from human dendritic cells (DC), which encodes a 733-residue protein with a BTB/POZ domain at the N-terminal and 4 C2H2 zinc fingers at C-terminal. It was designated dendritic cell-derived BTB/POZ zinc finger (DPZF). DPZF protein shares closest homology to BCL-6, with the highest homology present in the BTB/POZ and zinc finger domains. Like BCL-6, DPZF gene is localized on chromosome 3. It is widely expressed in hematopoietic tissues, including DC, monocytes, B cells, and T cells. DPZF protein expression is detectable in lymphoid neoplasm with a molecular mass of 100 kD, especially in B lymphoma. These indicate that DPZF may be a transcription factor closely related to BCL-6, and may be involved in hematopoiesis, oncogenesis, and immune responses.

Amino Acid Sequence↗
PubMed2001

Solution structure of the transcriptional activation domain of the bacteriophage T4 protein, MotA.

Bacteriophage T4 encodes a transcription factor, MotA, that binds to the -30 region of middle-mode promoters and activates transcription by host RNA polymerase. The crystal structure of the N-terminal domain of MotA (MotNF) revealed a six-helix domain in which the two C-terminal alpha-helices mediate the formation of a dimer via a coiled-coil motif and hydrophobic interactions. This structure suggested that full-length MotA binds DNA as a dimer, but subsequent biochemical results have shown that a monomeric form of MotA binds DNA. In this study, gel filtration chromatography, dynamic light scattering, and NMR-based diffusion measurements show conclusively that MotNF is a monomer, and not a dimer, in solution. In addition, we have determined the monomeric solution structure of MotNF using NMR spectroscopy, and have compared this with the dimer structure observed in crystals. The core of the protein assumes the same helical conformation in solution and in crystals, but important differences are observed at the extreme C-terminus. In solution, helix alpha5 is followed by five disordered residues that probably link the N-terminal and C-terminal domains of MotA. In crystals, helix alpha5 forms the dimer interface and is followed by a short sixth helix that further stabilizes the dimer configuration. The solution structure of MotNF supports the conclusion that MotA functions as a monomer, and suggests that the existence of the sixth helix in crystals is a consequence of crystal packing. Our work highlights the importance of investigating protein structures in both crystals and solution to fully understand biomolecular structure and to accurately deduce relationships between structure and function.

Amino Acid Sequence↗
PubMed2001

[Expression of apoptosis-related proteins in Hashimoto's thyroiditis and its pathological significance].

OBJECTIVE: To explore the role and significance of apoptosis-related proteins in the pathogenesis and pathological changes in Hashimoto's thyroiditis (HT). METHODS: Apoptotic level in thyroid tissue specimens from 17 patients with HT and 17 patients with nontoxic goiter (NTG) was evaluated by TUNEL method. Expression and distribution of Fas, FasL, Bcl-2 and Bax proteins in the specimens were investigated with immunohistochemical methods. RESULTS: A high percentage of apoptosis (32.5% +/- 12.3%) was observed by TUNEL method in thyroid follicles from HT specimens in comparison with that from NTG specimens (1.3% +/- 0.7%, P < 0.01). Positive rates of Fas, FasL, Bcl-2 and Bax in thyroid cells of NTG (35% approximately 47%) were significantly lower than those in thyroid cells of HT (88% approximately 94%, P < 0.01). The intensity of positive immunostaining for Fas, FasL, Bcl-2 and Bax in thyroid cells of HT was significantly higher than that of the controls (P < 0.01). Apoptosis and strongly positive immunostained thyroid cells with HT for Fas, FasL and Bax were mainly distributed in follicles adjacent to lymphocytic infiltrates, whereas strongly positive stained thyroid cells for Bcl-2 were mainly located in the area remote from the infiltrating lymphocytes. Staining of infiltrating lymphocytes for Fas, FasL, Bcl-2 and Bax was weak. CONCLUSION: Thyroid follicles in HT specimens exhibited high apoptotic levels. The high expression of apoptosis-related proteins Fas, FasL, Bcl-2 and Bax in thyrocytes of HT and weak expression in infiltrating lymphocytes accounted for thyroid follicle destruction and diffuse lymphocytic infiltration in HT.

Adult↗
PubMed2001

From receptor recognition mechanisms to bioinspired mimetic antagonists in HIV-1/cell docking.

Understanding the ways in which two or more proteins interact may give insight into underlying binding and activation mechanisms in biology, methods for protein separation and structure-based antagonism. This review describes ways in which protein recognition has been explored in our laboratory for the HIV-1/cell entry process. Initial contact between an HIV-1 virion particle and a human cell occurs between gp120 (an HIV-1 envelope protein) and CD4 (a human extracellular signaling protein). This interaction leads to a sequence of events which includes a conformational change in gp120, fusion of the HIV-1 and cellular membranes and eventual infection of the cell. Using an optical biosensor and a reporter antibody, we have been able to measure the conformational change in gp120 that occurs upon CD4 binding. We also have used this biosensor system to characterize CD4 mimetics, obtained by peptide synthesis in miniprotein scaffolds. Phage display techniques have been employed to identify novel miniprotein sequences. The combination of biosensor interaction kinetics analysis and phage display provides a useful approach for understanding the recognition mechanisms involved in the HIV/cell docking process. This approach may also be useful in investigating other protein complexes of importance in health and disease.

CD4 Antigens↗
PubMed2001

Inactivation of pyruvate formate-lyase by dioxygen: defining the mechanistic interplay of glycine 734 and cysteine 419 by rapid freeze-quench EPR.

Pyruvate formate-lyase from Escherichia coli (EC 2.3.1.54; PFL) catalyzes the reversible anaerobic conversion of pyruvate and CoA into acetyl-CoA and formate. Active PFL contains a novel alpha-carbon centered glycyl radical at G734 that is required for its catalytic activity. Two adjacent cysteine residues, C418 and C419, are essential for PFL activity according to site-directed mutagenesis studies. Upon exposure to air, active PFL loses its activity with the concomitant loss of the glycyl radical. Previous EPR studies of dioxygen inactivation of PFL revealed protein-based peroxyl and sulfinyl radicals during the manual mixing and quenching process [Reddy et al. (1998) Biochemistry 37, 558-563]. To probe the mechanism of this process, we carried out experiments using rapid freeze-quench EPR spectroscopy. Upon mixing of active wild type or C418A PFL with oxygenated solution, a short-lived radical intermediate appears at the earliest time point (10 ms), followed by the appearance of a long-lived sulfinyl radical. The axial EPR spectrum of this short-lived radical (g = 2.034, 2.007) is characteristic of a peroxyl radical. When C419A PFL or the double mutant [C418A/C419A] PFL was mixed with oxygenated solution, the peroxyl radical was also observed at 10 ms but in this case persisted over 12 s. These observations provide compelling evidence to support a proposed mechanism in which dioxygen quenches the glycyl radical in the active enzyme and the resulting peroxyl radical may react further with the sulfhydryl group of the C419 residue to form the sulfinyl radical.

Acetylation↗
PubMed2001

Involvement of caspases in 4-hydroxy-alkenal-induced apoptosis in human leukemic cells.

4-Hydroxynonenal (HNE), a reactive and cytotoxic end-product of lipid peroxidation, has been suggested to be a key mediator of oxidative stress-induced cell death and in various cell types has been shown to induce apoptosis. We have demonstrated that HNE, at micromolar concentrations, induces dose- and time-dependent apoptosis in a leukemic cell line (CEM-C7). Interestingly, much higher concentrations of HNE (> 15-fold) were required to induce apoptosis in leukocytes obtained from normal individuals. We also demonstrate that HNE causes a decrease in clonogenicity of CEM-C7 cells. Furthermore, our data characterize the caspase cascade involved in HNE-induced apoptosis in CEM-C7 cells. Using specific fluorogenic substrates and irreversible peptide inhibitors, we demonstrate that caspase 2, caspase 3, and caspase 8 are involved in HNE-induced apoptosis, and that caspase 2 is the first initiator caspase that activates the executioner caspase 3, either directly or via activation of caspase 8. Our studies also suggest the involvement of another executioner caspase, which appears to be similar to caspase 8 but not caspases 2 and 3, in its specificity. The demonstration of decreased clonogenicity by HNE in the leukemic cells, and their higher susceptibility to HNE-induced apoptosis as compared to the normal cells, suggests that such compounds may have potential for leukemia chemotherapy.

Aldehydes↗