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Biomedical subjects

G Bai

Publications and source records attributed to G Bai.

At least 73 records · Page 4Linked to original sources
PubMed1993

Cloning and analysis of the 5' flanking sequence of the rat N-methyl-D-aspartate receptor 1 (NMDAR1) gene.

We cloned and analyzed a 3.8 kb EcoRI fragment of the rat NMDAR1 gene. It contains 3 kb of promoter/enhancer region, exon 1 and a portion of intron 1. Two major transcription start sites were identified at -276 and -238 from the first nucleotide in codon 1. One GSG and two SP1 motifs, but no TATA/CAAT boxes, exist in the region proximal to the transcription start sites. Our results suggest that NMDAR1 has the characteristics of a housekeeping gene and may be regulated by immediate-early genes.

Amino Acid Sequence↗
PubMed1993

Expression and characterization of rat protein phosphatases-1 alpha, -1 gamma 1, -1 gamma 2, and -1 delta.

Four distinct cDNAs for rat protein phosphatase-1 have been isolated from rat tissues (Sasaki et al., Jpn. J. Cancer Res. 81, 1272-1280, 1990). These cDNAs encode proteins of highly similar sequence, the major differences being located at their N and C termini. In order to demonstrate that these cDNAs encode functional proteins and to investigate their enzymatic properties, it would be desirable to obtain purified preparations of these proteins. Using a system that was developed for the expression of rabbit muscle protein phosphatase-1 (Zhang et al., J. Biol. Chem. 267, 1484-1490, 1992) we have expressed these isoforms in Escherichia coli. The four recombinant isoforms were purified to near homogeneity and their properties were examined in terms of substrate specificity and sensitivity to okadaic acid and inhibitor-2.

Animals↗
PubMed1993

Mutagenesis of the catalytic subunit of rabbit muscle protein phosphatase-1.

We have generated site-directed mutants of the catalytic subunit of rabbit muscle ppase-1. Since it is known that ppase-1 and ppase-2A are highly susceptible to inactivation by sulfhydryl reagents, we have mutagenized the six cysteine residues conserved between these two enzymes to serines. The six mutants were purified to near homogeneity by affinity chromatography on inhibitor-2-Sepharose and characterized. All six exhibited enzymatic activity. These results indicate that the catalytic mechanism of ppase-1 is different from that of the protein tyrosine phosphatases which involve a cysteinyl phosphate intermediate.

Amino Acid Sequence↗
PubMed1993

Localization of the multiple calmodulin messenger RNAs in differentiated PC12 cells.

Calmodulin, a ubiquitous calcium-binding protein which is involved in many biological processes, including cell proliferation and differentiation, has been shown to be encoded by three genes from which five calmodulin messenger RNAs are transcribed. In our previous studies, using the PC12 pheochromocytoma cell line as a model system for neuronal differentiation, all five calmodulin messenger RNAs were found to be present, and treatment with both nerve growth factor and dibutyryl cyclic AMP, which induce neurite outgrowth in these cells, increased the level of calmodulin and differentially increased the levels of the various calmodulin messenger RNAs. In an attempt to uncover the nature of the differential increase in the calmodulin messenger RNAs during neuronal differentiation, we examined here the subcellular distribution of the individual calmodulin messenger RNAs in PC12 cells treated with nerve growth factor and dibutyryl cyclic AMP by in situ hybridization cytochemistry, using radiolabeled oligodeoxynucleotide probes. Using an oligodeoxynucleotide probe which detects all of the calmodulin transcripts, the calmodulin messenger RNAs were found to be distributed throughout the cell bodies of differentiated PC12 cells; significant amounts of calmodulin messenger RNAs were also found in most neurites (approximately 70% of the total number). Using specific probes for the calmodulin messenger RNAs derived from each calmodulin gene, distinct patterns of localization of the different calmodulin messenger RNAs were revealed. The messenger RNAs from calmodulin genes I and II were readily detected in all cell bodies and in about one-half of the neurites. In contrast, a weak signal for the messenger RNAs from calmodulin gene III was associated with cell bodies, while no significant signal was found in neurites. A population distribution analysis of the labeling of individual PC12 cell bodies, as determined by counting autoradiographic grains, revealed differences in the relative abundance of each group of messenger RNAs derived from each of the three calmodulin genes. The order of relative abundance of the messenger RNAs in cell bodies was found to be: calmodulin gene II messenger RNA > calmodulin gene I messenger RNAs >> calmodulin gene III messenger RNAs. An analysis of the labeling density along neurites indicated a similar density of neuritic messenger RNAs from calmodulin gene I and calmodulin gene II, whereas there was no significant signal for the messenger RNAs from calmodulin gene III.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Combinatorial regulation by promoter and intron 1 regions of the metallothionein SpMTA gene in the sea urchin embryo.

The SpMTA metallothionein gene of the sea urchin Strongylocentrotus purpuratus is regulated developmentally, histospecifically, and by heavy-metal induction. The sequenced 5' flank of the gene can be divided into proximal, middle, and distal regions, each containing a pair of metal response elements (MREs). Canonical 7-bp core sequences are present in all except the middle-region MREs c and d, which contain 1-bp mismatches. Metal-induced expression in transgenic blastulae was increased with each consecutive addition of the middle and distal regions to a chimeric reporter gene construct containing the proximal SpMTA promoter region. Reduced metal induction through point mutation of the distal MREs e and f indicated that the MREs themselves were largely responsible for the transcriptional increase. These activities were further enhanced by SpMTA intron 1, but not when a specific interior region of the intron had been deleted. The atypical MREs c and d did not support induction by themselves, i.e., when present alone with mutated proximal MREs a and b. However, in the presence of intron 1, they were able to substitute for the nullified MREs a and b in the promotion of metal-induced expression. This capability suggests, furthermore, that these atypical MREs, in addition to responding to an intron 1 region, participate cooperatively with the canonical proximal MREs.

Amino Acid Sequence↗
PubMed1993

Determination of thiocyanate metabolite of sodium nitroprusside in serum by spectrophotometry.

A sensitive, reproducible and rapid method for the spectrophotometric determination of thiocyanate, the metabolite of sodium nitroprusside, was developed. Acetonitrile was used to remove protein in the serum. Uniform design technique was used in arranging the experiments and choosing the reaction conditions between thiocyanate and ferric nitrate reagent. The wavelength of maximum absorption was 456 nm, the calibration curve of SCN- was linear in the range of 1.68-13.4 micrograms/ml (r = 0.9999), the relative average recovery of thiocyanate was 94.0%. The day-to-day and within-day relative standard deviations were less than 4.0% (n = 8) and 3.2% (n = 10) respectively. The limit of determination was 0.3 microgram/ml. Drugs such as lidocaine, etc. presented no interference on the determination.

Aged↗
PubMed1993

Sporotrichosis.

Explore the source record for details and available documents.

Administration, Oral↗
PubMed1993

Apoxesis of primary tracheal tumor. A clinical analysis of 23 cases.

During a 17-year period, 23 patients with primary tracheal tumors underwent surgical treatment in the Department of Thoracic Surgery, PUMC Hospital (11 cases of benign tumor of the trachea, 12 cases of low malignancies). Fourteen times of apoxesis were performed in 11 patients with benign tumors who were followed up for an average of 6.3 years. Fifteen operations were performed in 12 cases including local resection of the tracheal wall and tumor in 4, and curettage of tumor plus electric cauterization on the basis in 10. Eight of 9 patients with adenoid cystic carcinoma received postoperative adjuvant irradiation, with a 5-year postoperative survival rate of 75% (6/8) and 3 cases survived over ten years. The desirability of apoxesis and local resection of tracheal tumor is discussed. The authors suggest that these two surgical patterns can be regarded as a simple and effective treatment for patients with primary tracheal tumors.

Adolescent↗
PubMed1992

Cyclic AMP selectively up-regulates calmodulin genes I and II in PC12 cells.

Calmodulin is encoded by three genes in rat tissues, which collectively transcribe five different mRNAs. We showed previously that the pheochromocytoma cell line, PC12, expresses all five of these mRNAs and that treating PC12 cells with nerve growth factor differentially induces the calmodulin mRNAs; the greatest increase was seen by 24 h in the 1.4 kb transcript from calmodulin gene II. In the present study we found that treating PC12 cells with dibutyryl cyclic AMP also differentially increased the levels of the calmodulin mRNAs. However, dibutyryl cyclic AMP produced increases as early as 3 to 6 h, with the greatest increase (about 3-fold) being seen in the level of the 1.7 kb mRNA transcribed from calmodulin gene I. The transcripts of 4.1 kb (calmodulin gene I) and 1.4 kb (calmodulin gene II) were also increased, but the 2.3 kb transcript from calmodulin gene III remained stable. Another cyclic AMP analogue, chlorophenylthio cyclic AMP, produced effects similar to those of dibutyryl cyclic AMP, but dibutyryl cyclic GMP did not. Pretreatment with cycloheximide blocked the increase in the 4.1 kb calmodulin mRNA induced by dibutyryl cyclic AMP, but only partially blocked the increase in the 1.4 kb and 1.7 kb transcripts. Phorbol 12-myristate 13-acetate, which can induce some cyclic AMP responsive genes, failed to significantly change the levels of any of the calmodulin mRNAs. These studies show that, like nerve growth factor, cyclic AMP can selectively up-regulate the transcripts from calmodulin genes in PC12 cells, but the time-course and type of transcripts induced by cyclic AMP are distinct. These results suggest that the mechanisms by which these two agents alter the calmodulin transcripts are different.

Animals↗
PubMed1992

Expression of the catalytic subunit of phosphorylase phosphatase (protein phosphatase-1) in Escherichia coli.

The catalytic subunit of rabbit skeletal muscle protein phosphatase-1 was expressed in Escherichia coli. Expression of phosphatase-1 in the pET3a vector, which is based on the use of the T7 promoter, resulted in the expression of the enzyme as an insoluble aggregate. The insoluble enzyme could be renatured by high dilutions of the urea-solubilized protein in buffers containing dithiothreitol, Mn2+, and high NaCl concentrations. However, under all conditions tested, only partial (less than 5%) renaturation was achieved. A second attempt was made using a vector with the trp-lac hybrid promoter. In this case it was possible to express the enzyme as a soluble protein at levels of 3-4% of the soluble E. coli protein. The recombinant enzyme was purified by DEAE-Sepharose and heparin-Sepharose chromatography. Approximately 20 mg of purified enzyme was reproducibly obtained from the cells derived from 2 liters of culture. The purified enzyme had a specific activity toward phosphorylase alpha comparable to that reported for the authentic protein and had an Mr of 37,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The recombinant enzyme displayed similar sensitivities to inhibition by inhibitor-2, okadaic acid, and microcystin-LR as for the protein isolated from rabbit muscle. At all stages of purification the recombinant phosphatase behaved as an essentially inactive enzyme that required the presence of microM Mn2+ for full expression of its activity.

Animals↗
PubMed1991

A cDNA encoding rabbit muscle protein phosphatase 1 alpha complements the Aspergillus cell cycle mutation, bimG11.

The bimG11 allele causes a conditional growth defect in the fungus Aspergillus nidulans preventing both progression through mitosis and normal polar growth. Previously, we have shown that the bimG11 mutation increases the phosphorylation of nuclear proteins and that the gene encodes a protein similar to mammalian type 1 protein phosphatase. Assay of protein phosphatase activity in protein extracts of Aspergillus demonstrates directly that type 1 phosphatase activity is greatly reduced in the mutant at restrictive temperature. Expression of a muscle type 1 protein phosphatase fully complements all aspects of the bimG11 phenotype, and restores the level of PP1 activity to nearly normal. Expression of the related phosphatase, PP2A, does not complement the bimG11 mutation, showing that complementation can only be achieved by the type 1 gene. This clearly demonstrates that the phenotype of bimG11 is due to reduced PP1 activity and that the PP1 catalytic subunit is functionally conserved over a wide span of evolution.

Alleles↗
PubMed1991

The increase of calmodulin in PC12 cells induced by NGF is caused by differential expression of multiple mRNAs for calmodulin.

A rat pheochromocytoma cell line (PC12 cells) was used as a model to investigate the role of calmodulin and its multiple mRNAs in NGF-induced neuronal differentiation. The effect of NGF on the degree of differentiation was assayed using a simple differentiation scoring system. Significant increases in the differentiation score were seen by one day, and the scores increased about 10-fold by 8 days of treatment. NGF also increased calmodulin in the PC12 cells; significant increases were seen by 2 days of treatment, and a maximum increase of 3-fold was seen by 4 days. Northern blot analysis using a calmodulin riboprobe revealed that all five calmodulin mRNAs found in rat tissue were present in PC12 cells. The relative abundance of the calmodulin mRNAs was 1.7 greater than 1.4 greater than 2.3 greater than 4.1 greater than 0.9 kb. NGF treatment caused a differential increase in these mRNAs. The 1.4 kb transcript (from Gene II) was increased earlier (at 1 day) and to a greater extent (3-fold) than any of the other mRNAs. Studies of the half-lives (t1/2) of these mRNAs suggested that the t1/2 varied with the mRNA; the smaller the mRNA, the shorter the t1/2. However, there were no significant effects of NGF on the t1/2 of any of the mRNAs. These studies indicate that NGF elevates calmodulin in PC12 cells by causing a differential increase in the multiple mRNAs for calmodulin and that the increase in calmodulin may play some part in NGF-induced neuronal differentiation in PC12 cells.

Animals↗
PubMed1991

Long-term effects of insulin on the enzyme activity and messenger RNA of glycogen synthase in rat hepatoma H4 cells: an effect of insulin on glycogen synthase mRNA stability.

Insulin induced glycogen synthase activity and decreased glycogen synthase mRNA concentrations in rat hepatoma H4 cells. Total enzyme activity measured with glucose 6-phosphate gradually increased during a 24-h insulin incubation. The time course of glycogen synthase activation measured by the activity ratio (low G-6-P/high G-6-P) in response to insulin was biphasic with the first peak at 15 min and the second peak at 4 to 6 h. When cells were incubated with insulin and cycloheximide, the first peak persisted while the second peak was abolished. These data suggest that the first activation peak derives from the classic effect of insulin via dephosphorylation and the second peak from an insulin-induced protein synthesis of a glycogen synthase activator. Ribonuclease protection assays with a cloned rat liver glycogen synthase cDNA were used to quantitate glycogen synthase mRNA. Insulin unexpectedly decreased glycogen synthase mRNA in a time- and a dose-dependent manner. After incubation with the RNA synthesis inhibitor, 5, 6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB) without and with insulin, the half time of glycogen synthase mRNA decreased from 6.0 +/- 0.80 to 3.9 +/- 0.75 h, respectively. Nuclear run-off experiments with isolated nuclei showed no change of transcription of glycogen synthase mRNA. These data suggest that insulin in this system affects glycogen synthase mRNA stability rather than transcription.

Animals↗
PubMed1991

Neuronal localization and modulation of the D2 dopamine receptor mRNA in brain of normal mice and mice lesioned with 6-hydroxydopamine.

A novel oligonucleotide probe was designed, characterized and utilized to study the distribution and modulation of the mRNA encoding the D2 dopamine receptor in the brain of the mouse. Using in situ hybridization histochemistry, the highest levels of the D2 receptor mRNA were found in regions of the brain containing the cell bodies and the terminal projection fields of the nigrostriatal, mesolimbic and mesocortical dopaminergic systems. Particularly high levels of the D2 receptor mRNA were found in substantia nigra pars compacta, ventral tegmental area, caudate-putamen and olfactory tubercle. This distribution generally paralleled that of the D2 dopamine receptor. Some areas, not usually associated with dopaminergic systems, also contained significant levels of the D2 receptor mRNA signal. These areas included the hippocampus, certain thalamic nuclei, the inferior colliculus and the spinal trigeminal nucleus of the medulla and spinal cord. Lesioning the corpus striatum with 6-hydroxydopamine had little effect on the level of the D2 receptor mRNA in the striatum but greatly reduced the hybridization signal in the substantia nigra pars compacta and ventral tegmental area. Similarly, lesioning the substantia nigra, nearly abolished the signal in the pars compacta but failed to substantially alter the D2 receptor mRNA signal in the striatum. These results suggest that the D2 receptor mRNA in the substantia nigra pars compacta was localized largely to dopaminergic cell bodies, the terminal projections of which lie in the striatum and codes for D2 autoreceptors and that the D2 receptor mRNA of the striatum is in non-dopaminergic cell bodies that are intrinsic to the striatum and probably codes for post-synaptic D2 receptors. Further, the evidence that lesions of striatum and substantia nigra induced with 6-hydroxydopamine greatly reduced the D2 receptor mRNA signal in the substantia nigra, without concomitantly increasing the D2 receptor mRNA in the striatum, suggests that the increase in dopamine receptor binding in the striatum that is ipsilateral to the lesion with 6-hydroxydopamine and the enhanced behavioral sensitivity to dopaminergic agonists, cannot be accounted for solely by an increase in D2 receptor mRNA.

Animals↗
PubMed1991

[Preliminary investigation of effect of pyocyanin on common bacteria in burn wound].

Pyocyanin was extracted from culture medium of Ps. aeruginosa by Frank's method. Minimum inhibitory concentration (MIC) & minimum bactericidal concentration (MBC) of pyocyanin for 6 species of bacteria isolated from burn wounds were determined. The results of MIC & MBC: S. aureus 12.5 micrograms/ml, 50 micrograms/ml; E. coli 25 micrograms/ml, 100 micrograms/ml; P. vulgaris 50 micrograms/ml, 200 micrograms/ml; C. freundii 100 micrograms/ml, 400 micrograms/ml; S. epidermidis 12.5 micrograms/ml, 50 micrograms/ml; no inhibitory effect on Ps. aeruginosa. According to the laboratory results and clinical findings, the authors believe that when there is a dominant growth of Ps. aeruginosa in burn wounds there may be pyocyanin production to inhibit or kill other species of bacteria.

Burns↗
PubMed1990

The primary structure of rat liver glycogen synthase deduced by cDNA cloning. Absence of phosphorylation sites 1a and 1b.

The cDNA for rat liver glycogen synthase was isolated by screening a rat liver cDNA library constructed in lambda gt11. The cDNA was 2.4 kilobases in length and encoded a protein of 703 amino acid residues with a molecular mass of 80.5 kDa. Comparison of the rat liver and the human muscle sequences show that the amino- and carboxyl-terminal regions are quite divergent as compared to the internal sequences which show an 80% identity. The rat liver carboxyl-terminal region is truncated by 33 residues and has only 46% identity with the muscle sequence but retains the common feature of a low content of hydrophobic amino acids (13%). Phosphorylation sites 1a and 1b, which are the primary targets for phosphorylation by cAMP-dependent protein kinase, are absent in the liver sequence. The presence of these divergent, structurally anomalous carboxyl-terminal regions in liver and muscle glycogen synthase suggests the absence of the requirement that they possess a tertiary structure that is integral to that of the protein core. A model is proposed in which this region interacts with a catalytic core to maintain the I state, and in which phosphorylation serves to uncouple this interaction.

Amino Acid Sequence↗