Misregulated alternative splicing appears to be a major factor in the pathogenesis of myotonic dystrophy. The present study was done to further explore alternative splicing in this condition by doing exon-level analysis of mRNA from skeletal muscle of 8 subjects with type 1 myotonic dystrophy, 7 subjects with type 2 myotonic dystrophy, 8 disease controls (subjects with facioscapulohumeral muscular dystrophy), and 8 healthy controls . The ratios of signals from the various exons of a gene provided an index of altered exon inclusion/exclusion that was independent of the overall expression of that gene. There were numerous transcripts for which there was evidence of abnormal alternative splicing in subjects with myotonic dystrophy. For many of these transcripts, the abnormal splicing was confirmed by an independent RT-PCR approach.
Splicing biomarkers of disease severity in myotonic dystrophy.
Specimen part
View SamplesGlobal transcription machinery engineering (gTME) is an approach for reprogramming gene transcription to elicit cellular phenotypes important for technological applications. Here we show the application of gTME to Saccharomyces cerevisiae for improved glucose/ethanol tolerance, a key trait for many biofuels programs. Mutagenesis of the transcription factor Spt15p and selection led to dominant mutations that conferred increased tolerance and more efficient glucose conversion to ethanol. The desired phenotype results from the combined effect of three separate mutations in the SPT15 gene [serine substituted for phenylalanine (Phe177Ser) and, similarly, Tyr195His, and Lys218Arg]. Thus, gTME can provide a route to complex phenotypes that are not readily accessible by traditional methods.
Engineering yeast transcription machinery for improved ethanol tolerance and production.
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View SamplesThe transcriptional data from an integrative analysis of transcriptional and metabolic stress responses that provides a more complete understanding of the mechanisms by which genetic regulatory circuits mediate metabolic phenotype.
Linking high-resolution metabolic flux phenotypes and transcriptional regulation in yeast modulated by the global regulator Gcn4p.
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View SamplesIn this study we used microarrays to examine relative genes expression within the aorta of ApoE-/- infused with angiotensin II in relation to aneurysm formation. Infusion of angiotensin II induces aortic dilatation particularly of the suprarenal aorta in ApoE-/- mice. Based on studies carried out in our and other laboratories the response to angiotensin II is variable, with some mice developing large aneurysms but other animals appearing resistant to aneurysm formation with aortic diameters similar to that of saline controls. We compared RNA expression from whole aortas of 17 week old male ApoE-/- mice exposed to angiotensin II (1.44 g/kg/min) for 4 weeks where there was clear evidence of aortic aneurysm formation (n=5) with that of mice failing to develop aneurysms (n=7) and those exposed to saline infusion (n=6). AAA was defined as diameter of suprarenal aorta greated than 1.5mm measured on photographs of aortas at necroscopy.
Whole genome expression analysis within the angiotensin II-apolipoprotein E deficient mouse model of abdominal aortic aneurysm.
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View SamplesVitis vinifera endogenous small RNAs Overall design: Size fractionated small RNA from total RNA extracts of Vitis vinifera leaves, inflorescences, tendrils and small berries were ligated to adapters, purified again and reverse transcribed. After PCR amplification the sample was subjected to Solexa/Illumina high throughput pyrosequencing. Please see www.illumina.com for details of the sequencing technology.
Identification of grapevine microRNAs and their targets using high-throughput sequencing and degradome analysis.
Specimen part, Subject
View SamplesVitis vinifera RNA degradome Overall design: Isolated polyadenylated RNA from total RNA extracts of Vitis vinifera leaves, were ligated to 5'-adapter that include san MmeI recognition site. The ligated products were purified again, reverse transcribed and cleaved with MmeI. The 5' fragments were purified from gel and to a 3'- dsDNA adapter and PCR amplified. After PCR amplification the sample was subjected to Solexa/Illumina high throughput pyrosequencing. Please see www.illumina.com for details of the sequencing technology.
Identification of grapevine microRNAs and their targets using high-throughput sequencing and degradome analysis.
Specimen part, Subject
View SamplesPatients with palliative SCCHN were treated with figitumumab, an IGF-1R inhibitor. This receptor plays an important role in cell growth, proliferation and differentiation and is often overexpressed in SCCHN. No significant clinical activity was observed in our study
Phase II study of figitumumab in patients with recurrent and/or metastatic squamous cell carcinoma of the head and neck: clinical activity and molecular response (GORTEC 2008-02).
Specimen part
View SamplesIt is fundamentally unknown how normal cellular processes or responses to extracellular stimuli may invoke polyadenylation and degradation of ncRNA substrates or if human disease processes exhibit defects in polyadenylation of ncRNA substrates as part of their pathogenesis. Our results demonstrate that mononuclear cells from subjects with relapsing-remitting multiple sclerosis (RRMS) exhibit pervasive increases in levels of polyadenylated ncRNAs including Y1 RNA, 18S and 28S rRNA, and U1, U2, and U4 snRNAs and these defects are unique to RRMS. Defects in expression of both Ro60 and La proteins in RRMS appear to contribute to increased polyadenylation of ncRNAs. Further, IFN-ß1b, a common RRMS therapy, restores both Ro60 and La levels to normal as well as levels of polyadenylated Y1 RNA and U1 snRNA suggesting that aberrant polyadenylation of ncRNA substrates may have pathogenic consequences. Overall design: We extracted RNA from peripheral whole blood in healthy control subjects and patients with established relapsing-remitting multiple sclerosis using PaxGene tubes.
Defective structural RNA processing in relapsing-remitting multiple sclerosis.
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View SamplesTo improve our understanding of lncRNA expression in T cells, we used whole genome sequencing (RNA-seq) to identify lncRNAs expressed in human T cells and those selectively expressed in T cells differentiated under TH1, TH2, or TH17 polarizing conditions. The majority of these lineage-specific lncRNAs are co-expressed with lineage-specific protein-coding genes. These lncRNAs are predominantly intragenic with co-expressed protein-coding genes and are transcribed in sense and antisense orientations with approximately equal frequencies. Further, genes encoding TH lineage specific mRNAs are not randomly distributed across the genome but are highly enriched in the genome in genomic regions also containing genes encoding TH lineage-specific lncRNAs. Our analyses also identify a cluster of antisense lncRNAs transcribed from the RAD50 locus that are selectively expressed under TH2 polarizing conditions and co-expressed with IL4, IL5 and IL13 genes. Depletion of these lncRNAs via selective siRNA treatment demonstrates the critical requirement of these lncRNAs for expression of the TH2 cytokines, IL-4, IL-5 and IL-13. Collectively, our analyses identify new lncRNAs expressed in a TH lineage specific manner and identify a critical role for a cluster of lncRNAs for expression of genes encoding TH2 cytokines. Overall design: Human peripheral blood mononuclear cells (PBMC) were cultured under TH1, TH2, and TH17 polarizing conditions. TH1, TH2, and TH17 primary and effector cultures were isolated and poly(A)+ and total RNA sequencing performed.
Expression and functions of long noncoding RNAs during human T helper cell differentiation.
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View SamplesWe used Affymetrix DNA arrays to investigate the extent to which nuclear HDAC4 accumulation affects neuronal gene expression.
HDAC4 governs a transcriptional program essential for synaptic plasticity and memory.
Specimen part
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