Tonje Strømmen Steigedal

Semantic annotation of biomedical literature using google

With the increasing amount of biomedical literature, there is a need for automatic extraction of information to support biomedical researchers. Due to incomplete biomedical information databases, the extraction is not straightforward using dictionaries, and several approaches using contextual rules and machine learning have previously been proposed. Our work is inspired by the previous approaches, but is novel in the sense that it is using Google for semantic annotation of the biomedical words. The semantic annotation accuracy obtained – 52% on words not found in the Brown Corpus, Swiss-Prot or LocusLink (accessed using Gsearch.org) – is justifying further work in this direction.

Inducible cAMP early repressor splice variants ICER I and IIγ both repress transcription of c‐fos and chromogranin A

Inducible cAMP early repressor (ICER) splice variants are generated upon activation of an alternative, intronic promoter within the CREM gene. ICER is proposed to downregulate both its own expression, and the expression of other genes, containing cAMP‐responsive promoter elements. To examine the biological function of the two ICER splice variants, I and IIγ, in comparable cellular systems, we generated HEK 293 cell variants with controllable overexpression of either ICER I or IIγ. These two splice variants contain two different variants of DNA binding domains. Overexpression of either ICER I or IIγ strongly represses CRE‐driven reportergene transcription but not AP1‐ or NFκB‐driven transcription. Thus, high specificity is maintained even at ICER overexpression. We here show that both ICER I and IIγ repress Pituitary adenylate cyclase‐activating polypeptide (PACAP)‐mediated c‐fos mRNA induction with similar efficiency, indicating that both splice variants play an important role in modulating PACAP‐mediated transcriptional activation of the c‐fos gene. ICER I and IIγ also repress cAMP‐mediated activation of chromogranin A (CgA), indicating that these splice variants may function as negative feedback regulators in CgA synthesis. The proliferation rate was not altered in cells overexpressing ICER I or IIγ. Thus, in the epithelial cells HEK 293, ICER I and IIγ splice variants seem to exert similar biological function. J. Cell. Biochem. J. Cell. Biochem. 101: 1532–1544, 2007.

Gastrin-induced proliferation involves MEK partner 1 (MP1)

The peptide hormone gastrin is an important factor for the maintenance and homeostasis of the gastric mucosa. We show that gastrin stimulates proliferation in a dose-dependent manner in the human gastric adenocarcinoma cell line AGS-GR. Furthermore, we demonstrate that the MAPK scaffold protein MEK partner 1 (MP1) is important for gastrin-induced phosphorylation of ERK1 and ERK2 and that MP1 promotes gastrin-induced proliferation of AGS-GR cells. Our results suggest a role of MP1 in gastrin-induced cellular responses involved in proliferation and homeostasis of the gastric mucosa.

The duration of gastrin treatment affects global gene expression and molecular responses involved in ER stress and anti-apoptosis

BackgroundHow cells decipher the duration of an external signal into different transcriptional outcomes is poorly understood. The hormone gastrin can promote a variety of cellular responses including proliferation, differentiation, migration and anti-apoptosis. While gastrin in normal concentrations has important physiological functions in the gastrointestine, prolonged high levels of gastrin (hypergastrinemia) is related to pathophysiological processes.ResultsWe have used genome-wide microarray time series analysis and molecular studies to identify genes that are affected by the duration of gastrin treatment in adenocarcinoma cells. Among 403 genes differentially regulated in transiently (gastrin removed after 1 h) versus sustained (gastrin present for 14 h) treated cells, 259 genes upregulated by sustained gastrin treatment compared to untreated controls were expressed at lower levels in the transient mode. The difference was subtle for early genes like Junb and c-Fos, but substantial for delayed and late genes. Inhibition of protein synthesis by cycloheximide was used to distinguish between primary and secondary gastrin regulated genes. The majority of gastrin upregulated genes lower expressed in transiently treated cells were primary genes induced independently of de novo protein synthesis. This indicates that the duration effect of gastrin treatment is mainly mediated via post-translational signalling events, while a smaller fraction of the differentially expressed genes are regulated downstream of primary transcriptional events. Indeed, sustained gastrin treatment specifically induced prolonged ERK1/2 activation and elevated levels of the AP-1 subunit protein JUNB. Enrichment analyses of the differentially expressed genes suggested that endoplasmic reticulum (ER) stress and survival is affected by the duration of gastrin treatment. Sustained treatment exerted an anti-apoptotic effect on serum starvation-induced apoptosis via a PKC-dependent mechanism. In accordance with this, only sustained treatment induced anti-apoptotic genes like Clu, Selm and Mcl1, while the pro-apoptotic gene Casp2 was more highly expressed in transiently treated cells. Knockdown studies showed that JUNB is involved in sustained gastrin induced expression of the UPR/ER stress related genes Atf4, Herpud1 and Chac1.ConclusionThe duration of gastrin treatment affects both intracellular signalling mechanisms and gene expression, and ERK1/2 and AP-1 seem to play a role in converting different durations of gastrin treatment into distinct cellular responses.

Functional studies on transfected cell microarray analysed by linear regression modelling.

Transfected cell microarray is a promising method for accelerating the functional exploration of the genome, giving information about protein function in the living cell. The microarrays consist of clusters of cells (spots) overexpressing or silencing a particular gene product. The subsequent analysis of the phenotypic consequences of such perturbations can then be detected using cell-based assays. The focus in the present study was to establish an experimental design and a robust analysis approach for fluorescence intensity data, and to address the use of replicates for studying regulation of gene expression with varying complexity and effect size. Our analysis pipeline includes measurement of fluorescence intensities, normalization strategies using negative control spots and internal control plasmids, and linear regression (ANOVA) modelling for estimating biological effects and calculating P-values for comparisons of interests. Our results show the potential of transfected cell microarrays in studying complex regulation of gene expression by enabling measurement of biological responses in cells with overexpression and downregulation of specific gene products, combined with the possibility of assaying the effects of external stimuli. Simulation experiments show that transfected cell microarrays can be used to reliably detect even quantitatively minor biological effects by including several technical and experimental replicates.

ProtChew: Automatic Extraction of Protein Names from Biomedical Literature

With the increasing amount of biomedical literature, there is a need for automatic extraction of information to support biomedical researchers. Due to incomplete biomedical information databases, the extraction is not straightforward using dictionaries, and several approaches using contextual rules and machine learning have previously been proposed. Our work is inspired by the previous approaches, but is novel in the sense that it is fully automatic and doesn’t rely on expert tagged corpora. The main ideas are 1) unigram tagging of corpora using known protein names for training examples for the protein name extraction classi- fier and 2) tight positive and negative examples by having protein-related words as negative examples and protein names/synonyms as positive examples. We present preliminary results on Medline abstracts about gastrin, further work will be on testing the approach on BioCreative benchmark data sets.

Salt-Inducible Kinase 1 (SIK1) Is Induced by Gastrin and Inhibits Migration of Gastric Adenocarcinoma Cells

Salt-inducible kinase 1 (SIK1/Snf1lk) belongs to the AMP-activated protein kinase (AMPK) family of kinases, all of which play major roles in regulating metabolism and cell growth. Recent studies have shown that reduced levels of SIK1 are associated with poor outcome in cancers, and that this involves an invasive cellular phenotype with increased metastatic potential. However, the molecular mechanism(s) regulated by SIK1 in cancer cells is not well explored. The peptide hormone gastrin regulates cellular processes involved in oncogenesis, including proliferation, apoptosis, migration and invasion. The aim of this study was to examine the role of SIK1 in gastrin responsive adenocarcinoma cell lines AR42J, AGS-GR and MKN45. We show that gastrin, known to signal through the Gq/G11-coupled CCK2 receptor, induces SIK1 expression in adenocarcinoma cells, and that transcriptional activation of SIK1 is negatively regulated by the Inducible cAMP early repressor (ICER). We demonstrate that gastrin-mediated signalling induces phosphorylation of Liver Kinase 1B (LKB1) Ser-428 and SIK1 Thr-182. Ectopic expression of SIK1 increases gastrin-induced phosphorylation of histone deacetylase 4 (HDAC4) and enhances gastrin-induced transcription of c-fos and CRE-, SRE-, AP1- and NF-κB-driven luciferase reporter plasmids. We also show that gastrin induces phosphorylation and nuclear export of HDACs. Next we find that siRNA mediated knockdown of SIK1 increases migration of the gastric adenocarcinoma cell line AGS-GR. Evidence provided here demonstrates that SIK1 is regulated by gastrin and influences gastrin elicited signalling in gastric adenocarcinoma cells. The results from the present study are relevant for the understanding of molecular mechanisms involved in gastric adenocarcinomas.

gProt: annotating protein interactions using google and gene ontology

With the increasing amount of biomedical literature, there is a need for automatic extraction of information to support biomedical researchers. Due to incomplete biomedical information databases, the extraction cannot be done straightforward using dictionaries, so several approaches using contextual rules and machine learning have previously been proposed. Our work is inspired by the previous approaches, but is novel in the sense that it combines Google and Gene Ontology for annotating protein interactions. We got promising empirical results – 57.5% terms as valid GO annotations, and 16.9% protein names in the answers provided by our system gProt. The total error-rate was 25.6% consisting mainly of overly general answers and syntactic errors, but also including semantic errors, other biological entities (than proteins and GO-terms) and false information sources.

Functional Studies on RNA-Transfected Cell Microarrays

RNA-transfected cell microarray shows great promise in functional genomics. By printing siRNA complexed with transfection reagent on glass slides, arrays of transfected cells are formed in which the phenotypic consequences of gene suppression can be investigated. Using reporter plasmids with fluorescence intensity as output data, we have developed a strategy for statistical analysis of the intensity data from medium-scale functional studies using data from several experimental replicates.

Nephronectin is Correlated with Poor Prognosis in Breast Cancer and Promotes Metastasis via its Integrin-Binding Motifs

Most cancer patients with solid tumors who succumb to their illness die of metastatic disease. While early detection and improved treatment have led to reduced mortality, even for those with metastatic cancer, some patients still respond poorly to treatment. Understanding the mechanisms of metastasis is important to improve prognostication, to stratify patients for treatment, and to identify new targets for therapy. We have shown previously that expression of nephronectin (NPNT) is correlated with metastatic propensity in breast cancer cell lines. In the present study, we provide a comprehensive analysis of the expression pattern and distribution of NPNT in breast cancer tissue from 842 patients by immunohistochemical staining of tissue microarrays from a historic cohort. Several patterns of NPNT staining were observed. An association between granular cytoplasmic staining (in <10% of tumor cells) and poor prognosis was found. We suggest that granular cytoplasmic staining may represent NPNT-positive exosomes. We found that NPNT promotes adhesion and anchorage-independent growth via its integrin-binding and enhancer motifs and that enforced expression in breast tumor cells promotes their colonization of the lungs. We propose that NPNT may be a novel prognostic marker in a subgroup of breast cancer patients.