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Y Zhao

Publications and source records attributed to Y Zhao.

At least 19 recordsLinked to original sources
PubMed1992

CoA-dependent methylmalonate-semialdehyde dehydrogenase, a unique member of the aldehyde dehydrogenase superfamily. cDNA cloning, evolutionary relationships, and tissue distribution.

Three overlapping cDNA clones encoding methylmalonate-semialdehyde dehydrogenase (MMSDH; 2-methyl-3-oxopropanoate:NAD+ oxidoreductase (CoA-propanoylating); EC 1.2.1.27) have been isolated by screening a rat liver lambda gt 11 library with nondegenerate oligonucleotide probes synthesized according to polymerase chain reaction-amplified portions coding for the N-terminal amino acid sequence of rat liver MMSDH. The three clones cover a total of 1942 base pairs of cDNA, with an open reading frame of 1569 base pairs. The authenticity of the composite cDNA was confirmed by a perfect match of 43 amino acids known from protein sequencing. The composite cDNA predicts a 503 amino acid mature protein with M(r) = 55,330, consistent with previous estimates. Polymerase chain reaction was used to obtain the sequence of the 32 amino acids corresponding to the mitochondrial entry peptide. Northern blot analysis of total RNA from several rat tissues showed a single mRNA band of 3.8 kilobases. Relative mRNA levels were: kidney greater than liver greater than heart greater than muscle greater than brain, which differed somewhat from relative MMSDH protein levels determined by Western blot analysis: liver = kidney greater than heart greater than muscle greater than brain. A 1423-base pair cDNA clone encoding human MMSDH was isolated from a human liver lambda gt 11 library. The human MMSDH cDNA contains an open reading frame of 1293 base pairs that encodes the protein from Leu-74 to the C terminus. Human and rat MMSDH share 89.6 and 97.7% identity in nucleotide and protein sequence, respectively. MMSDH clearly belongs to a superfamily of aldehyde dehydrogenases and is closely related to betaine aldehyde dehydrogenase, 2-hydroxymuconic semialdehyde dehydrogenase, and class 1 and 2 aldehyde dehydrogenases.

Aldehyde Oxidoreductases↗
PubMed1992

Increased tyrosine kinase activity of c-Src during calcium-induced keratinocyte differentiation.

In cultured human epidermal keratinocytes, induction of differentiation by Ca2+ and ionophore treatment was found to result in rapid elevation of c-Src tyrosine kinase activity and inactivation of the c-Yes tyrosine kinase. Activation of c-Src kinase was accompanied by tyrosine dephosphorylation, which might be explained by a rapid increase in intracellular protein-tyrosine phosphatase activity. Ca(2+)-induced differentiation was also associated with altered tyrosine phosphorylation of several cellular proteins and correlated with a marked redistribution of intracellular phosphotyrosine from membrane and adhesion sites to the nucleus. Some of the c-Src protein was also found in the nucleus after Ca2+ treatment, and Ca(2+)-activated c-Src bound to three cellular proteins (120 kDa, 65 kDa, and 34 kDa). In agreement with these results, immunohistochemistry on human epidermis revealed an increase in c-Src expression and tyrosine phosphorylation in cells undergoing differentiation, which strongly suggests a possible role of non-receptor tyrosine kinases in epithelial cell maturation.

Calcium↗
PubMed1992

Branched-chain alpha-ketoacid dehydrogenase kinase. Molecular cloning, expression, and sequence similarity with histidine protein kinases.

A cDNA for branched-chain alpha-ketoacid dehydrogenase kinase was cloned from a rat heart cDNA library. The cDNA had an open reading frame encoding a protein of 382 amino acid residues with a calculated molecular weight of 43,280. The clone codes for the branched-chain alpha-ketoacid dehydrogenase kinase based on the following: 1) the deduced amino acid sequence contained the partial sequence of the kinase determined by direct sequencing; 2) expression of the cDNA in Escherichia coli resulted in synthesis of a 43,000-Da protein that was recognized specifically by kinase antibodies; and 3) enzyme activity that phosphorylated and inactivated the branched-chain alpha-ketoacid dehydrogenase complex was found in extracts of E. coli expressing the protein. Northern blot analysis indicated the mRNA for the branched-chain alpha-ketoacid dehydrogenase kinase was more abundant in rat heart than in rat liver, as expected from the relative amounts of kinase activity expressed in these tissues. The deduced sequence of the kinase aligned with a high degree of similarity within subdomains characteristic of procaryotic histidine protein kinases. This first mitochondrial protein kinase to be cloned appears more closely related in sequence to procaryotic histidine protein kinases than to eucaryotic serine/threonine protein kinases.

Amino Acid Sequence↗
PubMed1992

Effects of clofibric acid on the activity and activity state of the hepatic branched-chain 2-oxo acid dehydrogenase complex.

Feeding clofibric acid to rats caused little or no change in total activity of the liver branched-chain 2-oxo acid dehydrogenase complex (BCODC). No change in mass of liver BCODC was detected by immunoblot analysis in response to dietary clofibric acid. No changes in abundance of mRNAs for the BCODC E1 alpha, E1 beta and E2 subunits were detected by Northern-blot analysis. Likewise, dietary clofibric acid had no effect on the activity state of liver BCODC (percentage of enzyme in the dephosphorylated, active, form) of rats fed on a chow diet. However, dietary clofibric acid greatly increased the activity state of liver BCODC of rats fed on a diet deficient in protein. No stable change in liver BCODC kinase activity was found in response to clofibric acid in either chow-fed or low-protein-fed rats. Clofibric acid had a biphasic effect on flux through BCODC in hepatocytes prepared from low-protein-fed rats. Stimulation of BCODC flux at low concentrations was due to clofibric acid inhibition of BCODC kinase, which in turn allowed activation of BCODC by BCODC phosphatase. Inhibition of BCODC flux at high concentrations was due to direct inhibition of BCODC by clofibric acid. The results suggest that the effects of clofibric acid in vivo on branched-chain amino acid metabolism can be explained by the inhibitory effects of this drug on BCODC kinase.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1992

Molecular cloning, stage-specific expression and cellular distribution of a putative protein kinase from Plasmodium falciparum.

A putative protein kinase gene (PfPK2) has been isolated from the human parasite Plasmodium falciparum by using a mixed oligonucleotide pool which corresponds to a highly conserved region of serine/threonine protein kinases. The complete nucleotide sequence of 5 kb suggests the existence of a second transcriptional unit besides that of the PfPK2 gene, separated by a highly (A+T)-rich region and transcribed in a different orientation. No intron sequence exists in PfPK2. The predicted amino acid sequence of PfPK2 contains features characteristic of eukaryotic serine/threonine protein kinases. Within its putative catalytic domain it shares 33%, 30%, and 28% amino acid identities with rat calcium-calmodulin-dependent protein kinase, human protein kinase C, and bovine cAMP-dependent protein kinase, respectively. Outside the catalytic domain, however, PfPK2 has no homology with regulatory domains of other protein kinases, indicating PfPK2 might be modulated by signals different from those of higher eukaryotes or might be associated with other regulatory subunits. Using a specific antiserum raised in rabbits against a recombinant fragment of the protein expressed in Escherichia coli, PfPK2 was found to be expressed in a stage-specific fashion and mainly localized in the parasitic membrane.

Amino Acid Sequence↗
PubMed1992

A conditional yeast mutant deficient in mRNA transport from nucleus to cytoplasm.

Transport of mRNA from nucleus to cytoplasm is critical for eukaryotic gene expression; however, the mechanism of export is unknown. Selection and screening procedures have therefore been used to obtain a family of temperature-sensitive conditional mutants of Saccharomyces cerevisiae that accumulate poly(A)+ RNA in the nucleus when incubated at 37 degrees C, as judged by in situ hybridization. In one such mRNA transport mutant, mtr1-1, RNA synthesis continues, the export of poly(A)+ RNA is inhibited, intranuclear poly(A)+ is remarkably stable, and protein synthesis gradually stops. Thus, there is no tight coupling between RNA synthesis and export. The export lesion is reversible. Although mRNA export is clearly not a default option, neither inhibition of protein synthesis, inhibition of mRNA splicing, nor inhibition of poly(A)-binding protein function blocks export of the average poly(A)+, as judged by in situ hybridization. Further analysis of the family of mtr mutants should help map the path of RNA transport.

Adenosine Triphosphate↗
PubMed1992

Purification, characterization, regulation and molecular cloning of mitochondrial protein kinases.

The mitochondrial kinases responsible for the phosphorylation and inactivation of rat heart pyruvate dehydrogenase complex and the rat liver and heart branched-chain alpha-ketoacid dehydrogenase complexes have been purified to homogeneity. The branched-chain alpha-ketoacid dehydrogenase kinase is composed of one subunit with a molecular weight of 44 kDa; pyruvate dehydrogenase kinase has two subunits with molecular weights of 48 (alpha) and 45 kDa (beta). Proteolysis maps of branched-chain alpha-ketoacid dehydrogenase kinase and the two subunits of pyruvate dehydrogenase kinase are different, suggesting that all subunits are different entities. The alpha subunit of the rat heart pyruvate dehydrogenase kinase was selectively cleaved by chymotrypsin with concomitant loss of kinase activity, as previously shown for the bovine kidney enzyme, suggesting that the catalytic activity of pyruvate dehydrogenase kinase resides in this subunit. Polyclonal antibodies against branched-chain alpha-ketoacid dehydrogenase kinase, purified by an epitope selection method, bound only to the 44 kDa polypeptide of the branched-chain alpha-ketoacid dehydrogenase complex, substantiating that the 44 kDa protein corresponds to the kinase for this complex. Both kinases exhibited strong substrate specificity toward their respective complexes and would not inactivate heterologous complexes. The kinases possessed slightly different substrate specificities toward histones. Phosphorylation and inactivation of the branched-chain alpha-ketoacid dehydrogenase complex by its purified kinase was inhibited by alpha-chloroisocaproate and dichloroacetate, established inhibitors of the phosphorylation of the complex. cDNAs encoding the branched-chain alpha-ketoacid dehydrogenase kinase have been isolated from rat heart and rat liver lambda gt11 libraries. This represents the first successful cloning of a mitochondrial protein kinase. Preliminary data suggest that two different isoforms of the kinase may exist in different ratios in various tissues. No evidence was found for induction of the branched-chain alpha-ketoacid dehydrogenase complex nor its kinase by clofibric acid. Rather, clofibric acid is a potent inhibitor of the activity of the branched-chain alpha-ketoacid dehydrogenase kinase and this may be the molecular mechanism responsible for the myotonic effects of clofibric acid in man.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1992

Assignment of a human DNA double-strand break repair gene (XRCC5) to chromosome 2.

The Chinese hamster ovary (CHO-K1) cell mutant XRS-6 is defective in rejoining of DNA double-strand breaks and is hypersensitive to X-rays, gamma-rays, and bleomycin. Radiation resistance or sensitivity of somatic cell hybrids constructed from the fusion of XRS-6 cells with primary human fibroblasts strongly correlated with the retention of human chromosome 2 isozyme and molecular markers. Discordancies between some chromosome 2 markers and the radiation resistance phenotype in some of the hybrid cells suggested the location of the X-ray repair cross complementing 5 (XRCC5) gene on the p arm of chromosome 2. Introduction of human chromosome 2 by microcell-mediated chromosome transfer into the radiation-sensitive XRS-6 cells resulted in hybrid cells in which the radiation sensitivity was complemented. The chromosome 2p origin of the complementing human DNA in the microcell hybrids was supported by fluorescent in situ hybridization analysis of human metaphases using human DNA amplified from the hybrids by inter-Alu-PCR as chromosome-painting probes. XRCC5 is therefore provisionally assigned to human chromosome 2p.

Animals↗
PubMed1992

Identification and transcriptional analyses of the UL3 and UL4 genes of equine herpesvirus 1, homologs of the ICP27 and glycoprotein K genes of herpes simplex virus.

The DNA sequence of 3,240 nucleotides of the XbaI G fragment located in the unique long (UL) region of the equine herpesvirus 1 genome revealed two major open reading frames (ORFs) designated UL3 and UL4. The UL3 ORF of 470 amino acids (aa) maps at nucleotides (nt) 4450 to 3038 from the long terminus, and its predicted 51.4-kDa protein product exhibits significant homology to the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1; 32% identity) and to the ORF4 protein of varicella-zoster virus (13% identity). Interestingly, a zinc finger motif is conserved in the C-terminal domains of both ICP27 of HSV-1 (aa 483 to 508) and UL3 of equine herpesvirus 1 (aa 441 to 466). The UL4 ORF of 343 aa maps at nt 5618 to 4587 and could encode a protein of 38.1 kDa which exhibits significant homology to the UL53 protein (cell fusion protein or glycoprotein K) of HSV-1 (26% identity) and to the ORF5 protein of varicella-zoster virus (33% identity). Analyses of the UL4 amino acid sequence revealed domains characteristic of a membrane-bound glycoprotein and included potential signature sequences for (i) a signal sequence, (ii) two N-linked glycosylation sites, and (iii) four transmembrane domains. Nucleotide sequence analyses also revealed potential TATA boxes located upstream of the UL3 and UL4 ORFs. However, only a single polyadenylation signal (nt 2988 to 2983) was detected downstream of the UL3 ORF. Northern (RNA) blot hybridization and S1 nuclease analyses were used to map and characterize the UL3 and UL4 mRNAs. Metabolic inhibitors were used to identify the kinetic class of these two genes. The data revealed that UL3 is an early gene that encodes a 1.6-kb mRNA, while UL4 is a late gene encoding a 3.8-kb mRNA that overlaps the UL3 transcript. Both transcripts were shown by S1 nuclease analyses to initiate 24 to 26 nt downstream of their respective TATA boxes and to have a common transcription termination signal as a pair of 3'-coterminal mRNAs.

Amino Acid Sequence↗
PubMed1992

Effects of anoxia on the biphasic response of isolated strips of rabbit bladder to field stimulation, bethanechol, methoxamine and KCl.

The contractile response of the bladder can be divided into two phases: an initial rapid increase in tension and a prolonged period of sustained tension (plateau phase). The bladder empties primarily during the plateau phase of the contractile response. These two phases can be differentiated using both pharmacologic and metabolic agents, indicating that the two phases have independent energy requirements. The present study compares the phasic (peak) and tonic (plateau) components of the responses of isolated strips of bladder body and base to field stimulation, bethanechol, methoxamine and KCl administration. New Zealand White rabbits were anesthetized with pentobarbital, and the bladder was removed. The bladder was divided between body and base at the level of the ureteral orifices. Three strips of bladder body and three strips of bladder base were mounted in separate baths containing Tyrode's solution at 37 degrees C and equilibrated with 95% O2, 5% CO2. Anoxia was produced by changing the gas mixture to 95% nitrogen, 5% CO2. The effects of anoxia on the responses to field stimulation, bethanechol, methoxamine and KCl were determined at different times after the initiation of anoxia. The results of these studies can be summarized as follows: (1) Anoxia induced time-dependent plateau phases of the response to field stimulation (2 and 32 Hz). (2) The rate of inhibition of the plateau phase was significantly and substantially greater than that of the peak phase in both the bladder body and base. (3) Similarly, anoxia inhibited the plateau phase of the bladder body's response to bethanechol to a significantly and substantially greater degree than anoxia inhibited the peak contraction.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Effect of pinacidil on spontaneous and evoked contractile activity.

Uninhibited bladder contractions have been associated with a variety of bladder dysfunctions including outlet obstruction, neurogenic bladder, incontinence, and other neurologic and nonneurogenic bladder disorders. One class of compounds that is gaining popularity and support for the treatment of hyperreflexia is potassium channel openers, such as pinacidil and cromakalim. In general, these agents act by hyperpolarizing the smooth muscle membrane, resulting in an increase in membrane stability which in turn would be expected to inhibit spontaneous and evoked contraction. It is the purpose of this study to compare the potency and selectivity of pinacidil at inhibiting both hyperreflexia in vivo, and several forms of in vitro contractile stimulation in the rabbit. The following is a summary of the results. (1) Pinacidil is an effective inhibitor of hyperreflexia in the in vivo rabbit model. (2) Pinacidil is a substantially more potent inhibitor of the amplitude of the hyperreflexia than the frequency. (3) Pinacidil was substantially more potent at inhibiting the contractile response to 2-Hz stimulation than to 32-Hz stimulation, but was equally effective at inhibiting field stimulation of the bladder base and body. (4) Pinacidil was significantly more potent at inhibiting the peak response to field stimulation than the rate of tension generation. (5) Pinacidil was equally potent and effective at inhibiting the phasic and tonic components of the response to field stimulation. (6) Pinacidil was a more potent inhibitor of methoxamine stimulation of the bladder base than bethanechol stimulation of the bladder body. (7) Pinacidil was a noncompetitive or mixed inhibitor of both methoxamine and bethanechol stimulation, whereas it was a competitive inhibitor of KCl stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Role of tissue renin in the pathophysiology of hypertension in TGR(mREN2)27 rats.

A transgenic rat line, TGR(mREN2)27, was established by introducing the murine Ren-2 gene into the genome of rats by microinjection techniques. These rats exhibit severe hypertension, making them an interesting model in which to study the role of renin in the pathophysiology of hypertension. However, although the additional renin gene is the only genetic difference compared with control rats, the exact mechanism of hypertension in TGR(mREN2)27 rats is still unclear. It cannot be attributed to a stimulation of the endocrine renin-angiotensin system or to an overexpression of renin in the kidney, since plasma and kidney renin and renin gene expression in the kidney are low in these animals. Here we describe recent progress made toward elucidating mechanisms of hypertension in TGR(mREN2)27 rats. 1) TGR(mREN2)27 rats were bred to homozygosity. The development of high blood pressure in homozygous rats is accelerated compared with that of heterozygous rats. This is paralleled by a higher mortality rate in homozygous TGR(mREN2)27 rats. Blood pressure and mortality rate of homozygous transgenic rats were effectively reduced by 10 mg captopril per kilogram body weight. 2) Treatment of 8-week-old heterozygous TGR(mREN2)27 rats with 10 mg/kg body wt per day of the angiotensin II receptor antagonist DuP 753 for 4.5 weeks normalized blood pressure. After withdrawal of the drug, blood pressure increased rapidly, reaching control levels after 3 weeks. In another group of TGR(mREN2)27 rats treated with 0.5 mg/kg per day, there was no change in blood pressure. Plasma renin and plasma angiotensin II were significantly higher in the high-dose group compared with the low-dose group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Cortex Hormones↗
PubMed1992

[Study on casting precision of plat castable ceramic crowns].

The plat castable ceramic crown was made with investment material prepared in our college with our own casting technique by a Chinese-made casting machine. The accuracy and fitness of the crown were studied and appraised. Results show that the margin of fitness is 46 microns and that the cement film thickness of the dies in the spacer material group is 65.13 microns. Hence the results are satisfactory. The Plat castable ceramic crown might be used in clinical dentistry.

Crowns↗
PubMed1992

Thrombomodulin as a marker of radiation-induced endothelial cell injury.

Cultured confluent human umbilical vein endothelial cells were irradiated in vitro with 60Co gamma rays at doses from 0 to 50 Gy. After irradiation thrombomodulin was measured at different times over 6 days in the supernatants of endothelial cell culture medium, on the surface of the cells, and within the cells. At 24 h after irradiation, an increase in the release of thrombomodulin from irradiated endothelial cells and an increase in the number of molecules and the activity of thrombomodulin on the surface of the cells were observed; these reactions were dependent on radiation dose. The capacity of the cells to produce and release thrombomodulin was decreased from 2 to 6 days after exposure to 60Co gamma rays. Our data indicate that radiation can injure endothelial cells, and that thrombomodulin may be used as a marker of radiation-induced injury in endothelial cells. The interrelationship between the dysfunction of irradiated endothelial cells and the pathological mechanisms of acute radiation disease is also discussed.

Biomarkers↗