Translon: a single term for translated regions.
Explore the source record for details and available documents.
Letter↗
Biomedical subjects
Ola Larsson
Publications and source records attributed to Ola Larsson.
Apoptosis resistance downstream of eIF4E: posttranscriptional activation of an anti-apoptotic transcript carrying a consensus hairpin structure.
Aberrant activation of the translation initiation machinery is a common property of malignant cells, and is essential for breast carcinoma cells to manifest a malignant phenotype. How does sustained activation of the rate limiting step in protein synthesis so fundamentally alter a cell? In this report, we test the post transcriptional operon theory as a possible mechanism, employing a model system in which apoptosis resistance is conferred on NIH 3T3 cells by ectopic expression of eIF4E. We show (i) there is a set of 255 transcripts that manifest an increase in translational efficiency during eIF4E-mediated escape from apoptosis; (ii) there is a novel prototype 55 nt RNA consensus hairpin structure that is overrepresented in the 5'-untranslated region of translationally activated transcripts; (iii) the identified consensus hairpin structure is sufficient to target a reporter mRNA for translational activation under pro-apoptotic stress, but only when eIF4E is deregulated; and (iv) that osteopontin, one of the translationally activated transcripts harboring the identified consensus hairpin structure functions as one mediator of the apoptosis resistance seen in our model. Our findings offer genome-wide insights into the mechanism of eIF4E-mediated apoptosis resistance and provide a paradigm for the systematic study of posttranscriptional control in normal biology and disease.
5' Untranslated Regions↗
Lack of correct data format and comparability limits future integrative microarray research.
Explore the source record for details and available documents.
Data Collection↗
Comparative microarray analysis.
Microarrays enable high-throughput parallel gene expression analysis, and their use has grown exponentially during the past decade. We are now in a position where individual experiments could benefit from using the swelling public data repositories to allow microarrays to progress from being a hypothesis-generating tool to a powerful resource that can be used to test hypothesis about biology. Comparative microarray analysis could better distinguish phenotypes from associated phenotypes; identify valid differentially expressed genes by combining many studies; test new hypothesis; and discover fundamental patterns of gene regulation. This review aims to describe the additional methodology needed for such comparative microarray analysis, and we identify and discuss a number of problems such as loss of published data, lack of annotations, and variable array quality, which need to be solved before comparative microarray analysis can be used in a more systematic and powerful manner.
Animals↗
The expression signature of in vitro senescence resembles mouse but not human aging.
BACKGROUND: The biological mechanisms that underlie aging have not yet been fully identified. Senescence, a phenomenon occurring in vitro, limits the number of cell divisions in mammalian cell cultures and has been suggested to contribute to aging. RESULTS: We investigated whether the changes in gene expression that occur during mammalian aging and induction of cellular senescence are similar. We compared changes of gene expression in seven microarray datasets from aging human, mouse and rat, as well as four microarray datasets from senescent cells of man and mouse. The datasets were publicly available or obtained from other laboratories. Correlation measures were used to establish similarities of the expression profiles and gene ontology analyses to identify functional groups of genes that are co-regulated. Robust similarities were established between aging in different species and tissues, indicating that there is an aging transcriptome. Although some cross-species comparisons displayed high correlation, intra-species similarities were more reliable. Similarly, a senescence transcriptome was demonstrated that is conserved across cell types. A similarity between the expression signatures of cellular senescence and aging could be established in mouse, but not in human. CONCLUSION: Our study is the first to use microarray data from several studies and laboratories for dissection of a complex biological phenotype. We demonstrate the presence of a mammalian aging transcriptome, and discuss why similarity between cellular senescence and aging is apparent in aging mice only.
Aging↗
Considerations when using the significance analysis of microarrays (SAM) algorithm.
BACKGROUND: Users of microarray technology typically strive to use universally acceptable data analysis strategies to determine significant expression changes in their experiments. One of the most frequently utilised methods for gene expression data analysis is SAM (significance analysis of microarrays). The impact of selection thresholds, on the output from SAM, may critically alter the conclusion of a study, yet this consideration has not been systematically evaluated in any publication. RESULTS: We have examined the effect of discrete data selection criteria (qualification criteria for inclusion) and response thresholds (out-put filtering) on the number of significant genes reported by SAM. The use of a reduced data set by applying arbitrary restrictions vis-à-vis abundance calls (e.g. from D-chip) or application of the fold change (FC) option within SAM (named the FC hurdle hereafter), can substantially alter the significant gene list when running SAM in Microsoft Excel. We determined that for a given final FC criteria (e.g. 1.5 fold change) the FC hurdle applied within Microsoft Excel SAM alters the number of reported genes above the final FC criteria. The reason is that the FC hurdle changes the composition of the control data set, such that a different significance level (q-value) is obtained for any given gene. This effect can be so large that it changes subsequent post hoc analysis interpretation, such as ontology overrepresentation analysis. CONCLUSION: Our results argue for caution when using SAM. All data sets analysed with SAM could be reanalysed taking into account the potential impact of the use of arbitrary thresholds to trim data sets before significance testing.
Adult↗
Human muscle gene expression responses to endurance training provide a novel perspective on Duchenne muscular dystrophy.
Global gene expression profiling is used to generate novel insight into a variety of disease states. Such studies yield a bewildering number of data points, making it a challenge to validate which genes specifically contribute to a disease phenotype. Aerobic exercise training represents a plausible model for identification of molecular mechanisms that cause metabolic-related changes in human skeletal muscle. We carried out the first transcriptome-wide characterization of human skeletal muscle responses to 6 wk of supervised aerobic exercise training in 8 sedentary volunteers. Biopsy samples before and after training allowed us to identify approximately 470 differentially regulated genes using the Affymetrix U95 platform (80 individual hybridization steps). Gene ontology analysis indicated that extracellular matrix and calcium binding gene families were most up-regulated after training. An electronic reanalysis of a Duchenne muscular dystrophy (DMD) transcript expression dataset allowed us to identify approximately 90 genes modulated in a nearly identical fashion to that observed in the endurance exercise dataset. Trophoblast noncoding RNA, an interfering RNA species, was the singular exception-being up-regulated by exercise and down-regulated in DMD. The common overlap between gene expression datasets may be explained by enhanced alpha7beta1 integrin signaling, and specific genes in this signaling pathway were up-regulated in both datasets. In contrast to these common features, OXPHOS gene expression is subdued in DMD yet elevated by exercise, indicating that more than one major mechanism must exist in human skeletal muscle to sense activity and therefore regulate gene expression. Exercise training modulated diabetes-related genes, suggesting our dataset may contain additional and novel gene expression changes relevant for the anti-diabetic properties of exercise. In conclusion, gene expression profiling after endurance exercise training identified a range of processes responsible for the physiological remodeling of human skeletal muscle tissue, many of which were similarly regulated in DMD. Furthermore, our analysis demonstrates that numerous genes previously suggested as being important for the DMD disease phenotype may principally reflect compensatory integrin signaling.
Biopsy↗
Functional comparison of single- and double-stranded siRNAs in mammalian cells.
The concept of small interfering RNA (siRNA) has been extended to include not only short double-stranded RNA of 19-25bp, but also single-stranded antisense RNA of the same length, since such single-stranded antisense siRNAs were recently found to be able to inhibit gene expression as well. We made comprehensive comparison of double- and single-stranded siRNA functions in RNA interference (RNAi), targeting multiple sites and different mRNAs, measuring RNAi effects at different time-points and in different cell lines, and examining response curves. Duplex siRNAs were found to be more potent than single-stranded antisense siRNAs. This was verified by the observation that single-stranded antisense siRNAs, which were inefficient in some cases when used alone, could be rescued from inefficiency by sequentially transfecting with the sense siRNAs. This result suggests that the structural character of siRNA molecules might be a more important determinant of siRNA efficiency than the cellular persistence of them.
Animals↗
Kinetics of senescence-associated changes of gene expression in an epithelial, temperature-sensitive SV40 large T antigen model.
Replicative senescence limits the number of times primary cells can divide and is therefore regarded as a potential checkpoint for cancer progression. The majority of studies examining changes of gene expression upon senescence have been made with stationary senescent cells. We wanted to study the transition from normal growth to senescence in detail and identify early regulators of senescence by analyzing early changes in global gene expression, using Affymetrix microarrays. For this purpose, we used a murine epithelial senescence model, where senescence is abrogated by SV40 large T antigen and can be induced by using a temperature-sensitive form of SV40 large T antigen (SV40ts58). Comparisons were made to wild-type SV40 large T antigen-expressing cells and to cells expressing SV40ts58 large T antigen grown to confluence. After removal of genes that are similarly regulated in wild-type and temperature-sensitive SV40 large T antigen-expressing cells, 60% of the remaining genes were shared between cells arrested by inactivation of SV40 T antigen and by confluence. We identified 125 up-regulated and 39 down-regulated candidate genes/expressed sequence tags that are regulated upon SV40 T antigen inactivation and not during heat shock or confluence and classified these based on their kinetic profiles. Our study identified genes that fall into different functional clusters, such as transforming growth factor-beta-related genes and transcription factors, and included genes not identified previously as senescence associated. The genes are candidates as early regulators of the senescence checkpoint and may be potential molecular targets for novel anticancer drugs.
Animals↗
Effective small interfering RNAs and phosphorothioate antisense DNAs have different preferences for target sites in the luciferase mRNAs.
Antisense DNA target sites can be selected by the accessibility of the mRNA target. It remains unknown whether a mRNA site that is accessible to an antisense DNA is also a good candidate target site for a siRNA. Here, we reported a parallel analysis of 12 pairs of antisense DNAs and siRNA duplexes for their potency to inhibit reporter luciferase activity in mammalian cells, both of the antisense DNA and siRNA agents in a pair being directed to same site in the mRNA. Five siRNAs and two antisense DNAs turned out to be effective, but the sites targeted by those effective siRNAs and antisense DNAs did not overlap. Our results indicated that effective antisense DNAs and siRNAs have different preferences for target sites in the mRNA.
Animals↗
Quantitative codon optimisation of DNA libraries encoding sub-random peptides: design and characterisation of a novel library encoding transmembrane domain peptides.
Codons for amino acids sharing similar chemical properties seem to cluster on the genetic codon table. Such a geographical distribution of the codons was exploited to create chemically synthesised DNA that encodes peptide libraries containing only a subset of the 20 natural amino acids. The frequency of each amino acid in the subset was further optimised by quantitatively manipulating the ratio of the four phosphoamidites during chemical synthesis of the libraries. Peptides encoded by such libraries show a reduced complexity and could be enriched in peptides of a desired property, which are thus more suitable when screening for functional peptides. Proof of concept for the codon-biased design of peptide libraries was shown by design, synthesis, and characterisation of a transmembrane peptide library that contains >80% transmembrane peptides, representing a 160-fold enrichment compared with a fully randomised library.
Amino Acid Sequence↗
Antisense PNA effects in Escherichia coli are limited by the outer-membrane LPS layer.
Antisense peptide nucleic acids (PNAs) can inhibit Escherichia coli gene expression and cell growth through sequence-specific RNA binding, and this opens possibilities for novel anti-infective agents and tools for microbial functional genomics. However, the cellular effects of PNAs are limited relative to effects in cell extracts, presumably because of cell barrier components such as the outer-membrane lipopolysaccharide (LPS) layer or drug efflux pumps, both of which function to exclude antibiotics and other foreign molecules. To evaluate the importance of such cellular factors on PNA effects, the authors developed a positive assay for antisense inhibition by targeting the lac operon repressor and compared PNA susceptibilities in mutant and wild-type E. coli by assessing lacZ induction. Strains with defective LPS (AS19 and D22) were more permeable to the antibiotic nitrocefin and more susceptible to PNA than the wild-type. Also, PNA potency was improved in wild-type cells grown in the presence of certain cell-wall-permeabilizing agents. In contrast, the activities of the Acr and Emr drug efflux pumps were not found to affect PNA susceptibility. The results show that the LPS layer is a major barrier against cell entry, but PNAs that can enter E. coli are likely to remain active inside cells.
Bacterial Proteins↗