Biomedical subjects

Ann-Ping Tsou

Publications and source records attributed to Ann-Ping Tsou.

15 recordsLinked to original sources

PubMed2006

DDX3, a DEAD box RNA helicase with tumor growth-suppressive property and transcriptional regulation activity of the p21waf1/cip1 promoter, is a candidate tumor suppressor.

DDX3 is a DEAD box RNA helicase with diverse biological functions. Using colony formation assay, our results revealed that DDX3 inhibited the colony formation ability of various tumor cells, and this inhibition might be due to a reduced growth rate caused by DDX3. Additionally, we identified p21(waf1/cip1), a cyclin-dependent kinase inhibitor, as a target gene of DDX3, and the up-regulation of p21(waf1/cip1) expression accounted for the colony-suppressing activity of DDX3. Moreover, DDX3 exerted its transactivation function on p21(waf1/cip1) promoter through an ATPase-dependent but helicase-independent mechanism, and the four Sp1 sites located within the -123 to -63 region, relative to the transcription start site of p21(waf1/cip1) promoter, were essential for the response to DDX3. Furthermore, DDX3 interacted and cooperated with Sp1 to up-regulate the promoter activity of p21(waf1/cip1). To determine the relevance of DDX3 in clinical cancers, the expression profile of DDX3 in various tumors was also examined. A declined expression of DDX3 mRNA and protein was found in approximately 58% to 73% of hepatoma specimens, which led to the reduction of p21(waf1/cip1) expression in a manner independent of p53 status. Additionally, an alteration of subcellular localization from nuclei to cytoplasm was also observed in >70% of cutaneous squamous cell carcinoma samples. Because DDX3 exhibits tumor suppressor functions, such as a growth-suppressive property and transcriptional activation of the p21(waf1/cip1) promoter, and is inactivated through down-regulation of gene expression or alteration of subcellular localization in tumor cells, all these features together suggest that DDX3 might be a candidate tumor suppressor.

Adenosine Triphosphatases↗

PubMed2006

miRNAMap: genomic maps of microRNA genes and their target genes in mammalian genomes.

Recent work has demonstrated that microRNAs (miRNAs) are involved in critical biological processes by suppressing the translation of coding genes. This work develops an integrated database, miRNAMap, to store the known miRNA genes, the putative miRNA genes, the known miRNA targets and the putative miRNA targets. The known miRNA genes in four mammalian genomes such as human, mouse, rat and dog are obtained from miRBase, and experimentally validated miRNA targets are identified in a survey of the literature. Putative miRNA precursors were identified by RNAz, which is a non-coding RNA prediction tool based on comparative sequence analysis. The mature miRNA of the putative miRNA genes is accurately determined using a machine learning approach, mmiRNA. Then, miRanda was applied to predict the miRNA targets within the conserved regions in 3'-UTR of the genes in the four mammalian genomes. The miRNAMap also provides the expression profiles of the known miRNAs, cross-species comparisons, gene annotations and cross-links to other biological databases. Both textual and graphical web interface are provided to facilitate the retrieval of data from the miRNAMap. The database is freely available at http://mirnamap.mbc.nctu.edu.tw/.

Animals↗

PubMed2006

Biological data warehousing system for identifying transcriptional regulatory sites from gene expressions of microarray data.

Identification of transcriptional regulatory sites plays an important role in the investigation of gene regulation. For this propose, we designed and implemented a data warehouse to integrate multiple heterogeneous biological data sources with data types such as text-file, XML, image, MySQL database model, and Oracle database model. The utility of the biological data warehouse in predicting transcriptional regulatory sites of coregulated genes was explored using a synexpression group derived from a microarray study. Both of the binding sites of known transcription factors and predicted over-represented (OR) oligonucleotides were demonstrated for the gene group. The potential biological roles of both known nucleotides and one OR nucleotide were demonstrated using bioassays. Therefore, the results from the wet-lab experiments reinforce the power and utility of the data warehouse as an approach to the genome-wide search for important transcription regulatory elements that are the key to many complex biological systems.

Algorithms↗

PubMed2006

Database to dynamically aid probe design for virus identification.

Viral infection poses a major problem for public health, horticulture, and animal husbandry, possibly causing severe health crises and economic losses. Viral infections can be identified by the specific detection of viral sequences in many ways. The microarray approach not only tolerates sequence variations of newly evolved virus strains, but can also simultaneously diagnose many viral sequences. Many chips have so far been designed for clinical use. Most are designed for special purposes, such as typing enterovirus infection, and compare fewer than 30 different viral sequences. None considers primer design, increasing the likelihood of cross hybridization to similar sequences from other viruses. To prevent this possibility, this work establishes a platform and database that provides users with specific probes of all known viral genome sequences to facilitate the design of diagnostic chips. This work develops a system for designing probes online. A user can select any number of different viruses and set the experimental conditions such as melting temperature and length of probe. The system then returns the optimal sequences from the database. We have also developed a heuristic algorithm to calculate the probe correctness and show the correctness of the algorithm. (The system that supports probe design for identifying viruses has been published on our web page http://bioinfo.csie.ncu.edu.tw/.)

Algorithms↗

PubMed2006

Effects of Scutellaria baicalensis Georgi on macrophage-hepatocyte interaction through cytokines related to growth control of murine hepatocytes.

The aim of this study is to elucidate the effects of Scutellaria baicalensis Georgi (SbG) extract and its constituents on macrophage-hepatocyte interaction in primary cultures. By using trans-well primary Kupffer cell culture or conditioned medium (CM) from murine macrophage RAW264.7 cell line (RAW cells), effects of SbG on hepatocyte growth were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide and trypan blue exclusion assay. Cytokine production, antibody-neutralization studies, and molecular mechanisms of transforming growth factor (TGF)-beta1 gene expression were elucidated on SbG-treated RAW264.7 cells. In addition, recombinant human TGF-beta1 (r-human TGF-beta1) was added to elucidate the mechanisms of SbG effects on cultured hepatocytes. Immunohistochemistry using anti-NF-kappaB antibody was used to determine the possible signal transduction pathways in primary hepatocyte culture. The results showed that SbG stimulated the proliferation of cultured hepatocytes, possibly through NF-kappaB, but not of Toll-like receptor 4 activation; whereas SbG-RAW-CM and SbG in trans-well significantly suppressed the proliferation of hepatocytes. Antibody-neutralization studies revealed that TGF-beta1 was the main antimitotic cytokine in SbG-treated RAW cells CM. The growth stimulation effect of SbG on cultured hepatocytes was inhibited by exogenous administration of r-human TGF-beta1. Furthermore, SbG induced NF-kB translocation into the nuclei of cultured cells. In the RAW264.7 line, SbG and baicalin stimulated TGF-beta1 gene expression via NF-kappaB and protein kinase C activation. We conclude that SbG stimulates hepatocyte growth via activation of the NF-kappaB pathway and induces TGF-beta1 gene expression through the Kupffer cell-hepatocyte interaction, which subsequently results in the inhibition of SbG-stimulated hepatocyte growth.

Animals↗

PubMed2005

Incorporating hidden Markov models for identifying protein kinase-specific phosphorylation sites.

Protein phosphorylation, which is an important mechanism in posttranslational modification, affects essential cellular processes such as metabolism, cell signaling, differentiation, and membrane transportation. Proteins are phosphorylated by a variety of protein kinases. In this investigation, we develop a novel tool to computationally predict catalytic kinase-specific phosphorylation sites. The known phosphorylation sites from public domain data sources are categorized by their annotated protein kinases. Based on the concepts of profile Hidden Markov Models (HMM), computational models are trained from the kinase-specific groups of phosphorylation sites. After evaluating the trained models, we select the model with highest accuracy in each kinase-specific group and provide a Web-based prediction tool for identifying protein phosphorylation sites. The main contribution here is that we have developed a kinase-specific phosphorylation site prediction tool with both high sensitivity and specificity.

Computational Biology↗

PubMed2005

Integrative genomics based identification of potential human hepatocarcinogenesis-associated cell cycle regulators: RHAMM as an example.

DNA microarray has been widely used to examine gene expression profile of different human tumors. The information generated from microarray analysis usually represents the overall range of cancer-associated abnormality associated with gene regulation. In order to identify key regulatory genes involved in carcinogenesis of human cancer, hypothesis driven data mining of the microarray data plus experimental validation becomes a critical approach in the post-genome era. Here, we present an integrative genomic analysis of published microarray data and homolog gene database. Over 20,000 genes were examined to reveal 16 genes specific to vertebrates, cell cycle G2/M regulated, and overexpressed in human HCC. Using Affymetrix microarray analysis, we found that all 16 genes were up-regulated in human HCC. Among these 16 genes, we experimentally validated the up-regulation of receptor for hyaluronan-mediated motility (RHAMM) in different cell model systems. We first confirmed elevation of RHAMM in the G2/M phase of synchronized HeLa cells. We also found that RHAMM had an elevated level of expression in all the HCC samples we examined and it was induced during the G2/M phase of regenerating mouse hepatocytes after partial hepatectomy. Thus, the expression of RHAMM appears to be tightly regulated during mammalian cell cycle G2/M progression. The ectopic overexpression of RHAMM in 293T cells resulted in the accumulation of cells at G2/M phase. RHAMM-induced mitotic arrest of cells was predominantly in the prophase. Taken together, using an integrated functional genomic approach, we have uncovered a set of genes that may play specific roles in cell cycle progression and in HCC development. To elucidate the function of these genes in cell cycle regulation may shed light on the control mechanism of human HCC in the future.

Amino Acid Sequence↗

PubMed2005

Phosphorylation and stabilization of HURP by Aurora-A: implication of HURP as a transforming target of Aurora-A.

Aurora-A, a mitotic serine/threonine kinase with oncogene characteristics, has recently drawn intense attention because of its association with the development of human cancers and its relationship with mitotic progression. Using the gene expression profiles of Aurora-A as a template to search for and compare transcriptome expression profiles in publicly accessible microarray data sets, we identified HURP (encodes hepatoma upregulated protein) as one of the best Aurora-A-correlated genes. Empirical validation indicates that HURP has several characteristics in common with Aurora-A. These two genes have similar expression patterns in hepatocellular carcinoma, liver regeneration after partial hepatectomy, and cell cycle progression and across a variety of tissues and cell lines. Moreover, Aurora-A phosphorylated HURP in vitro and in vivo. Ectopic expression of either the catalytically inactive form of Aurora-A or the HURP-4P mutant, in which the Aurora-A phosphorylation sites were replaced with Ala, resulted in HURP instability and complex disassembly. In addition, HURP-wild-type stable transfectants were capable of growing in low-serum environments whereas HURP-4P grew poorly under low-serum conditions and failed to proliferate. These studies together support the view that the ability to integrate evidence derived from microarray studies into biochemical analyses may ultimately augment our predictive power when analyzing the potential role of poorly characterized proteins. While this combined approach was simply an initial attempt to answer a range of complex biological questions, our findings do suggest that HURP is a potential oncogenic target of Aurora-A.

Amino Acid Sequence↗

PubMed2005

Gene expression profiling predicts liver responses to a herbal remedy after partial hepatectomy in mice.

The aim of this study was to demonstrate that regenerating liver responses to a herbal remedy could be presented by gene expression profiling. Compositions of the ingredients in the remedy containing Scutellaria baicalensis Georgi and Bupleurum scorzonerifolfium Wild (S/B remedy) were analyzed and quantified by high performance liquid chromatography. By using a 70% partial hepatectomy in BALB/c mice as an in vivo model, the effects of high dose (50 mg/kg) and low dose (1 mg/kg) S/B remedy were evaluated by cDNA microarray, followed by RT-PCR and real-time PCR confirmation. Factors affecting proliferative activities of mouse hepatocytes were measured by DNA flow cytometry, BrdU incorporation assay and serum interleukin-6 (IL-6) level. Based on global gene expression profiles, the results showed that the low dose S/B remedy down-regulated expression of immediate early genes and cell cycle-related genes, whereas the high dose had opposite effects. The gene expression was further verified by real-time RT-PCR. Proliferative activities, in terms of synthetic phase fractions and G2/M phase fractions, in vehicle, low dose, and high dose groups were 18.45+/-2.56%, 14.65+/-1.06%; 9.27+/-0.85%, 7.80+/-0.11%; and 18.90+/-2.17%, 22.95+/-0.25%, respectively. The serum IL-6 level was also dose-dependent in both low and high dose S/B remedy-treated mice. We conclude that in vivo gene expression profiling correlates with liver responses to a herbal remedy, which provides a new direction for pharmaceutical studies on human diseases.

Animals↗

PubMed2004

Identifying transcriptional regulatory sites in the human genome using an integrated system.

This work develops an integrated system which, after a set of genes are inputted, is able to predict transcriptional regulatory sites and to detect the co-occurrence of these regulatory sites. The system integrates several site detection methods such as known site matching, over-presented oligonucleotide detection and DNA motif discovery programs. User profiles and history pages enable users to trace the sequence analyses of these transcriptional regulatory sites. Two groups of co-regulated genes were used to test the proposed system. The results predicted by the proposed system consist of known site homologs and putative regulatory sites. By comparing these sites with previously published results, the proposed system is able to help biologists identify possible candidates for the regulatory sites from groups of co-regulated genes. The integrated system is now available at http://rgsminer. csie.ncu.edu.tw/.

Databases, Nucleic Acid↗

PubMed2003

Selenite-induced survival of HuH7 hepatoma cells involves activation of focal adhesion kinase-phosphatidylinositol 3-kinase-Akt pathway and Rac1.

Selenium has been shown to sustain the growth of selected human hepatocellular carcinoma cell lines under serum-free conditions, but the detailed mechanism remained undetermined. In the present study, the molecular mechanism(s) involving sodium selenite (Na2SO3, Se) as a survival agent were determined. Selenite not only protects HuH7 cells from serum deprivation-induced apoptosis, it also supports its long-term growth in sodium selenite (10(-7)m) supplemented serum-free medium. The anti-apoptotic effect correlates with activation of focal adhesion kinase and the phosphatidylinositol 3-kinase (PI3K)-Akt kinase pathway. Using HuH7 cells stably transfected with a constitutively active Akt kinase and PI3K inhibitor LY294002, selenite-induced cell survival was shown to be PI3K-Akt-dependent. Parallel changes included a significant reduction in the intracellular reactive oxygen species content, the reversal of DNA fragmentation, and the suppression of caspase and apoptosis signal-regulating kinase 1 activities. HuH7 cells stably expressing a Rac1 mutant N17 (Rac1N17-HuH7) are refractory to selenite treatment. In these cells selenite supplement neither triggers Akt activation nor supports cell proliferation. Participation of Rac1 activation in this event is supported by the fact that selenite treatment drastically enhanced activation of Rac1. The exact link between selenite treatment, Rac1 activation, and activation of the focal adhesion kinase-PI 3-kinase, however, remains to be characterized. The mitogenic signaling mediated by selenite may involve unconventional growth stimuli including higher glutathione peroxidase 1 activity and higher transcription levels of selenoprotein P. The selenium-HuH7 system we have established thus provides a unique tool that will allow the biological role of selenite in growth regulation of hepatocytes to be studied in detail.

Apoptosis↗

PubMed2003

Identification of a novel cell cycle regulated gene, HURP, overexpressed in human hepatocellular carcinoma.

An analytic strategy was followed to identify putative regulatory genes during the development of human hepatocellular carcinoma (HCC). This strategy employed a bioinformatics analysis that used a database search to identify genes, which are differentially expressed in human HCC and are also under cell cycle regulation. A novel cell cycle regulated gene (HURP) that is overexpressed in HCC was identified. Full-length cDNAs encoding the human and mouse HURP genes were isolated. They share 72 and 61% identity at the nucleotide level and amino-acid level, respectively. Endogenous levels of HURP mRNA were found to be tightly regulated during cell cycle progression as illustrated by its elevated expression in the G(2)/M phase of synchronized HeLa cells and in regenerating mouse liver after partial hepatectomy. Immunofluorescence studies revealed that hepatoma up-regulated protein (HURP) localizes to the spindle poles during mitosis. Overexpression of HURP in 293T cells resulted in an enhanced cell growth at low serum levels and at polyhema-based, anchorage-independent growth assay. Taken together, these results strongly suggest that HURP is a potential novel cell cycle regulator that may play a role in the carcinogenesis of human cancer cells.

Amino Acid Sequence↗

PubMed2003

Overexpression of a novel imprinted gene, PEG10, in human hepatocellular carcinoma and in regenerating mouse livers.

Many of the promising applications of the microarray technology are pertinent to identifying abnormalities in gene expression that contribute to malignant progression. We developed a bioinformatics tool to identify differentially expressed genes in human hepatocellular carcinoma (HCC). This involved the construction of a liver EST database (http://lestdb.nhri.org.tw) and in silico verification of differentially expressed genes with a human hepatoma microarray database. The stringency of the search was reinforced with a statistical analysis. A novel imprinted gene, paternally expressed 10(PEG10) was identified as having an elevated level of expression in the majority of the HCC samples and was also induced to express during G2/M phase of regenerating mouse liver. Ectopic expression of PEG10 in 293T cells affects cell cycle progression. PEG10 is distributed in the cytosol and associates with the nuclear membrane. This is the first time that an imprinted gene has been found to reexpress in both human HCC and in the regenerating mouse liver. This result indicates that the induction of the paternally imprinted gene may play an important role during liver regeneration or carcinogenesis of the human hepatocyte. Understanding the molecular basis of the abnormal imprinting of PEG10 will shed new light on the process that leads to liver disease.

Animals↗

PubMed2002

Identification of mutations in the arginine vasopressin receptor 2 gene causing nephrogenic diabetes insipidus in Chinese patients.

Congenital nephrogenic diabetes insipidus (NDI) is, in most instances, a rare X-linked recessive renal disorder (MIM 304800) characterized by the clinical symptoms of polyuria, polydipsia, and dehydration. The X-linked NDI is associated with mutations of the arginine vasopressin receptor type 2 (AVPR2) gene, which results in resistance to the antidiuretic action of arginine vasopressin (AVP) in the renal tubules and collecting ducts. Identification of mutations in the AVPR2 gene can facilitate early diagnosis of NDI, which can prevent serious complications such as growth retardation and mental retardation. We analyzed three unrelated Chinese NDI families and identified three mutations: R106C, F287L, and R337X. In addition, an A/G polymorphism at cDNA nucleotide position 927 (codon 309L) was identified. A functional expression assay of the R106C and F287L mutants in COS-7 cells revealed that both mutants show significant dysfunction and accumulate intracellular cyclic adenosine monophosphate in response to AVP hormone stimulation. These results facilitate the diagnosis of NDI at the molecular level in the Chinese population, and provide insight into the molecular pathology of NDI.

Amino Acid Sequence↗

PubMedDate not supplied

MultiProtIdent: identifying proteins using database search and protein-protein interactions.

Protein identification is important in proteomics. Proteomic analyses based on mass spectra (MS) constitute innovative ways to identify the components of protein complexes. Instruments can obtain the mass spectrum to an accuracy of 0.01 Da or better, but identification errors are inevitable. This study shows a novel tool, MultiProtIdent, which can identify proteins using additional information about protein-protein interactions and protein functional associations. Both single and multiple Peptide Mass Fingerprints (PMFs) are input to MultiProtIdent, which matches the PMFs to a theoretical peptide mass database. The relationships or interactions among proteins are considered to reduce false positives in PMF matching. Experiments to identify protein complexes reveal that MultiProtIdent is highly promising. The website associated with this study is http://dbms104.csie.ncu.edu.tw/.

Algorithms↗