Biomedical subjects

P C Liu

Publications and source records attributed to P C Liu.

At least 19 recordsLinked to original sources

PubMed2002

Pathogenesis of gastroenteritis caused by Vibrio carchariae in cultured marine fish.

Serious mortality among the cultured grouper Epinephelus coioides, characterized by a swollen intestine containing yellow fluid (gastroenteritis), occurred in 1993 in Taiwan. A bacterium isolated from the intestinal fluid and head kidney of moribund groupers was identified as Vibrio carchariae. Since then, the same Vibrio species has also been isolated from moribund black sea bream Acanthopagrus schlegeli, yellowfin sea bream A. latus, Japanese sea bass Lateolabrax japonicus, and red drum Sciaenops ocellatus suffering from the same syndrome. Each isolate was virulent to the respective fish. Recently, a similar syndrome, flounder infectious necrotizing enteritis, also caused by V. carchariae in summer flounder Paralichthys dentatus, was reported in Rhode Island. The extracellular products (ECPs) of V. carchariae strains EmI82KL (from grouper), Rd (from red drum), and SfUSA (from summer flounder, U.S.A.) were virulent to the grouper or red drum. A 33-kDa serine protease partially purified from the ECP of strain EmI82KL was lethal to the fish. All the moribund or killed fish exhibited gastroenteritis except those killed within 12 hours. This report is the first to show that intraperitoneal injection of the ECP or protease in the fish is virulent and can reproduce gastroenteritis. The serine protease was suggested as a major toxin in the grouper or red drum secreted by V. carchariae.

Journal Article↗

PubMed2001

The loop domain of heat shock transcription factor 1 dictates DNA-binding specificity and responses to heat stress.

Eukaryotic heat shock transcription factors (HSF) regulate an evolutionarily conserved stress-response pathway essential for survival against a variety of environmental and developmental stresses. Although the highly similar HSF family members have distinct roles in responding to stress and activating target gene expression, the mechanisms that govern these roles are unknown. Here we identify a loop within the HSF1 DNA-binding domain that dictates HSF isoform specific DNA binding in vitro and preferential target gene activation by HSF family members in both a yeast transcription assay and in mammalian cells. These characteristics of the HSF1 loop region are transposable to HSF2 and sufficient to confer DNA-binding specificity, heat shock inducible HSP gene expression and protection from heat-induced apoptosis in vivo. In addition, the loop suppresses formation of the HSF1 trimer under basal conditions and is required for heat-inducible trimerization in a purified system in vitro, suggesting that this domain is a critical part of the HSF1 heat-stress-sensing mechanism. We propose that this domain defines a signature for HSF1 that constitutes an important determinant for how cells utilize a family of transcription factors to respond to distinct stresses.

Amino Acid Sequence↗

PubMed2001

Novel stress-responsive genes EMG1 and NOP14 encode conserved, interacting proteins required for 40S ribosome biogenesis.

Under stressful conditions organisms adjust the synthesis, processing, and trafficking of molecules to allow survival from and recovery after stress. In baker's yeast Saccharomyces cerevisiae, the cellular production of ribosomes is tightly matched with environmental conditions and nutrient availability through coordinate transcriptional regulation of genes involved in ribosome biogenesis. On the basis of stress-responsive gene expression and functional studies, we have identified a novel, evolutionarily conserved gene, EMG1, that has similar stress-responsive gene expression patterns as ribosomal protein genes and is required for the biogenesis of the 40S ribosomal subunit. The Emg1 protein is distributed throughout the cell; however, its nuclear localization depends on physical interaction with a newly characterized nucleolar protein, Nop14. Yeast depleted of Nop14 or harboring a temperature-sensitive allele of emg1 have selectively reduced levels of the 20S pre-rRNA and mature18S rRNA and diminished cellular levels of the 40S ribosomal subunit. Neither Emg1 nor Nop14 contain any characterized functional motifs; however, isolation and functional analyses of mammalian orthologues of Emg1 and Nop14 suggest that these proteins are functionally conserved among eukaryotes. We conclude that Emg1 and Nop14 are novel proteins whose interaction is required for the maturation of the 18S rRNA and for 40S ribosome production.

Amino Acid Sequence↗

PubMed2001

Pathogenicity of Vibrio alginolyticus isolated from diseased small abalone Haliotis diversicolor supertexta.

Outbreaks of mass mortality among cultured small abalone Haliotis diversicolor supertexta with abscess/ulcers in the mantle occurred in 1998 at Kao-Hsiung, Taiwan. A swarming bacterium, strain H-11 was isolated from the haemolymph of the moribund small abalone using tryptic soy agar supplemented with 3% NaCl and/or thiosulphate citrate bile salt sucrose agar. This strain was characterized and identified as Vibrio alginolyticus on the basis of various biochemical tests. The H-11 strain and its extracellular products were virulent to small abalones with LD50 values of 3.6 x 10(5) colony forming units and 2.96 microg protein/g body weight, respectively.

Animals↗

PubMed2000

Virulence of Vibrio parahaemolyticus isolated from cultured small abalone, Haliotis diversicolor supertexta, with withering syndrome.

Outbreaks of mass mortality among cultured small abalone Haliotis diversicolor supertexta with withering syndrome occurred in May and September 1998 in Kao-Hsiung, Taiwan. Bacterial strains CH-1 and B4 were isolated from the haemolymph of the moribund small abalone using tryptic soy agar supplemented with 3% NaCl and/or thiosulphate citrate bile salt sucrose agar. These two strains were characterized and identified as Vibrio parahaemolyticus on the basis of various biochemical tests. The B4 strain and its extracellular products were virulent to small abalone with LD(50) values of 1.6 x 10(5) colony-forming units and 7.58 microg protein g-1 body weight, respectively.

Animals↗

PubMed1999

The mammalian HSF4 gene generates both an activator and a repressor of heat shock genes by alternative splicing.

The expression of heat shock genes is controlled at the level of transcription by members of the heat shock transcription factor family in vertebrates. HSF4 is a mammalian factor characterized by its lack of a suppression domain that modulates formation of DNA-binding homotrimer. Here, we have determined the exon structure of the human HSF4 gene and identified a major new isoform, HSF4b, derived by alternative RNA splicing events, in addition to a previously reported HSF4a isoform. In mouse tissues HSF4b mRNA was more abundant than HSF4a as examined by reverse transcription-polymerase chain reaction, and its protein was detected in the brain and lung. Although both mouse HSF4a and HSF4b form trimers in the absence of stress, these two isoforms exhibit different transcriptional activity; HSF4a acts as an inhibitor of the constitutive expression of heat shock genes, and hHSF4b acts as a transcriptional activator. Furthermore HSF4b but not HSF4a complements the viability defect of yeast cells lacking HSF. Moreover, heat shock and other stresses stimulate transcription of target genes by HSF4b in both yeast and mammalian cells. These results suggest that differential splicing of HSF4 mRNA gives rise to both an inhibitor and activator of tissue-specific heat shock gene expression.

Alternative Splicing↗

PubMed1999

A mouse gene knockout model for juvenile ceroid-lipofuscinosis (Batten disease).

The human hereditary ceroid-lipofuscinoses are a group of autosomal recessively inherited diseases characterized by massive accumulations of autofluorescent lysosomal storage bodies in the cells of many tissues and by neuronal degeneration throughout the central nervous system. There are a number of clinically and genetically distinct forms of ceroid-lipofuscinosis, the most common of which is the juvenile type, also known as Batten disease and CLN3. To study the mechanisms that lead to pathology in CLN3 and to evaluate potential therapies, a mouse model has been generated by targeted disruption of the mouse ortholog of the CLN3 gene (Cln3). As in affected humans, mice homozygous for the disrupted Cln3 allele show accumulation of autofluorescent storage material in neurons and other cell types. The storage material consists of membrane-bounded intracellular inclusions with ultrastructural features typical of the ceroid-lipofuscinoses. The accumulation of this storage material validates the Cln3 knockout mice as a model for the human disorder.

Animals↗

PubMed1999

Modulation of human heat shock factor trimerization by the linker domain.

Heat shock transcription factors (HSFs) are stress-responsive proteins that activate the expression of heat shock genes and are highly conserved from bakers' yeast to humans. Under basal conditions, the human HSF1 protein is maintained as an inactive monomer through intramolecular interactions between two coiled-coil domains and interactions with heat shock proteins; upon environmental, pharmacological, or physiological stress, HSF1 is converted to a homotrimer that binds to its cognate DNA binding site with high affinity. To dissect regions of HSF1 that make important contributions to the stability of the monomer under unstressed conditions, we have used functional complementation in bakers' yeast as a facile assay system. Whereas wild-type human HSF1 is restrained as an inactive monomer in yeast that is unable to substitute for the essential yeast HSF protein, mutations in the linker region between the DNA binding domain and the first coiled-coil allow HSF1 to homotrimerize and rescue the viability defect of a hsfDelta strain. Fine mapping by functional analysis of HSF1-HSF2 chimeras and point mutagenesis revealed that a small region in the amino-terminal portion of the HSF1 linker is required for maintenance of HSF1 in the monomeric state in both yeast and in transfected human 293 cells. Although linker regions in transcription factors are known to modulate DNA binding specificity, our studies suggest that the human HSF1 linker plays no role in determining HSF1 binding preferences in vivo but is a critical determinant in regulating the HSF1 monomer-trimer equilibrium.

Amino Acid Sequence↗

PubMed1999

Electrophoretic characterization of a novel cysteine protease produced by Vibrio harveyi.

Electrophoretic characterization of a novel cysteine protease produced by pathogenic luminous Vibrio harveyi, originally isolated from diseased tiger prawn Penaeus monodon in Taiwan, is demonstrated in the present study using native polyacrylamide gel electrophoresis (native PAGE), sodium dodecyl sulfate-PAGE (SDS-PAGE), crossed immunoelectrophoresis (CIE) and isoelectric focusing (IEF) gels. The protease has a pI of 6.4 and exhibits a fast-migrating feature in native-PAGE and CIE gels indicating that it is a negatively charged protease. The protease electrophoresed as a 22 kDa protein band in native- and SDS-PAGE (in SDS - buffer with or without the presence of 2-mercaptoethanol) while it electrophoresed as a 38 kDa protein band in SDS-PAGE when the samples were boiled for 10 min prior to electrophoresis. The results reveal that the enzyme is an SDS-resistant monomeric protease and its high negative charge is not influenced by SDS (detergent) without boiling the sample. The present results are useful in determining proteins of similar nature to this unique cysteine protease.

Cysteine Endopeptidases↗

PubMed1999

Characterization and chromosomal mapping of a mouse ortholog of the late-infantile ceroid-lipofuscinosis gene CLN2.

Late-infantile ceroid-lipofuscinosis (CLN2) is an autosomal recessively inherited, neurodegenerative disease in humans. The CLN2 locus has been mapped to Chromosome (Chr) 11p15, and its sequence and genomic organization have recently been reported. In the present study, the cDNA sequence, exon/intron organization, and chromosomal localization of a mouse ortholog of the CLN2 gene are described. The mouse cDNA contains an open reading frame that predicts a protein product of 562 amino acids. The mouse and human coding regions are 86% and 88% identical at the nucleic acid and amino acid levels, respectively. One less codon appears in the mouse cDNA when compared with the human ortholog. The mouse gene (Cln2) spans more than 6 kb and consists of 13 exons separated by introns ranging in size from 111 to 1259 bp. Length polymorphism in an (AC)(n) microsatellite in intron 3 of the mouse Cln2 gene was used to perform segregation analysis with The Jackson Laboratory DNA Panel Mapping Resource. On the basis of this analysis, the Cln2 gene was localized to a region of mouse Chr 7 that corresponds to human Chr 11p15. Characterization of the mouse Cln2 gene will facilitate generation of a mouse model for late-infantile ceroid-lipofuscinosis by gene targeting and identification of functionally important regions of the Cln2 protein.

Amino Acid Sequence↗

PubMed1999

Dual challenges of infectious pancreatic necrosis virus and Vibrio carchariae in the grouper, Epinephelus sp.

The grouper industry in Taiwan faces serious threats from various disease problems. The present study investigated dual challenges with infectious pancreatic necrosis virus (IPNV) and Vibrio carchariae in the grouper (Epinephelus sp.). The fish were infected with IPNV for 2 weeks prior to a secondary infection with the bacteria, or vice versa, by either immersion (10(3)-10(4) TCID50 IPNV per ml, 10(6)-10(7) colony forming units (CFU) Vibrio per ml) or by intraperitoneal injection (10(3)-10(4) TCID50 IPNV per g fish or 10(7) CFU Vibrio/g fish) challenges. Mass mortalities occurred in fish infected with IPNV for 2 weeks prior to the infection with the bacteria, or vice versa, in either immersion or intraperitoneal injection challenges. The bacterium could only survive in seawater or brackish water similar to that of cultured groupers.

Animals↗

PubMed1999

Cysteine protease is a major exotoxin of pathogenic luminous Vibrio harveyi in the tiger prawn, Penaeus monodon.

The role of an extracellular cysteine protease, produced by pathogenic luminous Vibrio harveyi strain 820514 originally isolated from diseased tiger prawn (Penaeus monodon), in the disease process in the prawns was studied. The protease was lethal to P. monodon with an LD50 value of 0.3 microgram protein g-1 prawn. The lethal toxicity of the extracellular products (ECP) of the bacterium was neutralized by pre-incubation of the ECP with rabbit antiserum to the cysteine protease. Pre-incubation of ECP with CuCl2 (an inhibitor of cysteine protease) also inhibited toxicity. This suggests that cysteine protease is the major toxin produced by the bacterium. The present protease is the first toxic cysteine protease to be found in Vibrio species.

Animals↗

PubMed1999

An evaluation of chromogenic substrates for characterization of serine protease produced by pathogenic Vibrio alginolyticus.

Four chromogenic substrates for characterizing serine protease of Vibrio alginolyticus were evaluated. The protease activity of bacterial extracellular products, or the fractions of 33 kD protease purified by the AKTA purifier system with various columns, was completely inhibited by ethylenediamine tetra-acetic acid, ethylene glycol-bis(beta-amino-ethyl ether) N,N,N',N'-tetraacetic acid (EGTA), antipain and phenylmethylsulphonyl fluoride (PMSF) using water-soluble substrates (azoalbumin and azocasein). It was only completely inhibited by antipain and PMSF using water-insoluble substrates (azocoll and hide powder azure). The protease activity was not, or only partially, inhibited by 1,10-phenanthroline and sodium dodecyl sulphate (SDS) using all four substrates. Since chelating agents and 1,10-phenanthroline are commonly employed as inhibitors to identify metalloprotease, the two water-soluble substrates may not be appropriate for this purpose, except for using 1,10-phenanthroline as an inhibitor. Chelating agents may be still applicable as inhibitors using water-insoluble substrates and 1,10-phenanthroline is highly recommended in the characterization for metalloprotease to avoid confusion. In the present study, the 33 kD protease was further confirmed as an SDS-resistant serine protease and not a metalloprotease.

Animals↗

PubMed1999

Purification and partial characterization of a toxic serine protease produced by pathogenic Vibrio alginolyticus.

An extracellular lethal toxin produced by Vibrio alginolyticus strain Swy originally isolated from diseased kuruma prawn (Penaeus japonicus) was purified using the AKTA purifier system with hydrophobic interaction chromatography, anion exchange and gel filtration columns. The toxin is an alkaline serine protease, inhibited by phenyl methylsulphonyl fluoride (PMSF), antipain and shows maximal activity at pH 8 to 11, having a pI of 4.3 and a molecular weight of approximately 33 kD. The toxin was completely inhibited by FeCl2 but partially inhibited by 3,4-dichloroisocoumarin (3,4-DCI), ethylenediamine tetraacetic acid (EDTA), ethylene glycol-bis(beta-amino-ethyl ether) N,N,N',N'-tetraacetic acid (EGTA), CuCl2 and ZnCl2. The purified protease was lethal for kuruma prawn at an LD50 of 0.29 microgram protein/g body weight. The haemolymph withdrawn from the moribund prawns injected with the toxic protease was unable to clot. The coagulogen in the kuruma prawn plasma showed an increased migration rate after incubation with this serine protease, and a plasma colour change from blue to pink was recorded. The addition of PMSF completely inhibited the lethal toxicity of the purified protease, indicating that this serine protease was a lethal toxin produced by the bacterium. The 33 kD protease was therefore a toxic protease produced by V. alginolyticus strain Swy.

Animals↗

PubMed1998

Suppression of C/EBPalpha and induction of C/EBPbeta by 2,3,7,8-tetrachlorodibenzo-p-dioxin in mouse adipose tissue and liver.

We examined the effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on two transcription factors, CAAT/enhancer binding protein-alpha (C/EBPalpha) and beta (C/EBPbeta), involved in the coordination of gene expression in adipose and liver. A single dose of TCDD (100 microg/kg) to male C57BL mice resulted in a time- and dose-dependent decrease in the level of C/EBPalpha mRNA in adipose tissue and liver, and a reciprocal increase in C/EBPbeta mRNA. Gel shift analysis using hepatic nuclear extracts from control and TCDD-treated mice and an oligonucleotide containing a C/EBP recognition element revealed a time-dependent change in DNA-protein complexes formed. Bands corresponding to C/EBPalpha, as determined by supershift analysis, diminished in TCDD-treated animals over a 7-day time period, whereas two new bands corresponding to C/EBPbeta, not present in control extracts, were increased significantly in treated samples. TCDD induced C/EBPbeta mRNA in wild-type mouse hepatoma cells, but not in aryl hydrocarbon receptor (AhR) nuclear translocator-deficient hepatoma cells. Induction in wild-type hepatoma cells was antagonized effectively by a molar excess of alpha-naphthoflavone. These results showed that TCDD caused rapid, reciprocal changes in C/EBPalpha and C/EBPbeta mRNAs and DNA binding in the adipose and liver of male C57BL mice and induced C/EBPbeta in hepatoma cells in an AhR-dependent manner. C/EBPs play vital roles in the coordination of energy homeostasis, and their alteration by TCDD may provide insight into the mechanism by which TCDD perturbs energy storage and utilization in vivo.

Adipose Tissue↗

PubMed1998

Coding sequence and exon/intron organization of the canine CLN3 (Batten disease) gene and its exclusion as the locus for ceroid-lipofuscinosis in English setter dogs.

Hereditary ceroid-lipofuscinosis in English setters has been proposed to be the canine equivalent of human juvenile ceroid-lipofuscinosis, which results from defects in the CLN3 gene. Analyses were performed to determine whether the disease in English setters is also the consequence of a CLN3 gene mutation. Canine CLN3 cDNA was found to contain a 1,314-bp open reading frame predicting a derived amino acid sequence which is 89%, 85%, and 84% identical to the predicted amino acid sequences for the human, mouse, and rabbit CLN3 proteins, respectively. The canine gene has sixteen exons. No differences were detected when cDNA nucleotide sequences from an English setter with ceroid-lipofuscinosis and from a normal dog were compared. Moreover, alleles of the canine CLN3 gene distinguished by an intragenic marker segregated independently from the disease in an English setter family, eliminating CLN3 as the locus for the canine disease. A ceroid-lipofuscinosis-affected Tibetan terrier was homozygous for a Gly70Glu CLN3 variant; however, this allele is common in dog breeds considered free of ceroid-lipofuscinosis.

Amino Acid Sequence↗

PubMed1998

Inhibition of cyclin D1 expression and phosphorylation of retinoblastoma protein by phosmidosine, a nucleotide antibiotic.

In this report, we studied the effect of phosmidosine, a proline-containing nucleotide on the serum-induced cell cycle progression in human lung fibroblast WI-38 cells. Phosmidosine suppressed S-phase entry and arrested cell cycle progression at the G1 phase. In serum-stimulated cells, phosmidosine did not affect the activation of the mitogen-activated protein kinase cascade. However, phosmidosine inhibited hyperphosphorylation of retinoblastoma (RB) protein by RB-kinases such as cyclin-dependent kinase 4 and cyclin-dependent kinase 2, probably as a result of the inhibition of cyclin D1 expression. Furthermore, in tsFT210 cells, a temperature-sensitive cdc2 mutant isolated from the mouse mammary carcinoma cell line FM3A, phosmidosine, irreversibly inhibited the cell cycle progression at G1 without affecting the G2 to M transition. Phosmidosine acts at an earlier point in G1 compared with mimosine or aphidicolin, well-known cell cycle blockers at the G1-S boundary. Taken together, phosmidosine arrested cells at a specific point between the start point and restriction point in G1 and is a useful drug that may contribute to the understanding of the regulatory mechanisms of G1 progression.

Animals↗

PubMed1998

Allelotype analysis of uterine leiomyoma: localization of a potential tumor suppressor gene to a 4-cM region of chromosome 7q.

Uterine leiomyoma is a benign smooth muscle tumor of the myometrium and is the most commonly encountered neoplasm in women of reproductive age. As for most benign tumors, the pathogenesis of leiomyoma remains obscure, especially at the molecular genetic level. The purpose of this study was to perform a genome-wide allelotype analysis to identify potential sites of tumor suppressor gene inactivation. Fifty-two cases of uterine leiomyoma were subjected to allelotype analysis by using matched pairs of tumor and blood DNA. Loss of heterozygosity (LOH) was assessed at 61 microsatellite markers distributed throughout the genome and representing all 41 chromosome arms. In general, LOH was very rare except on chromosome 7q, where LOH was observed in 34% of all informative tumors. Fine-deletion mapping with 25 microsatellite markers from the 7q22 region revealed a minimal deletion unit of approximately 4 cM, bounded by the markers D7S2453 proximally and D7S496 distally, that probably harbors a novel tumor suppressor gene involved in the etiology of this tumor.

Alleles↗