Project description:To examine whether the mutations in NLS1/2 affect the interaction with mRNAs, we determined the in vitro and in vivo interaction of WT Xrn1 and Xrn1 Delta NLS1/2 mutant with cellular mRNAs.
Project description:We performed ATAC-seq in the GR18 cell line (derived from U2OS ATTC:HTB-96, stably transfected with an expression construct for GR), upon glucocorticoid (dexamethasone) or vehicle (ethanol) treatment for 90 minutes.
Project description:The aim of the study was to evaluate excretory-secretory protein set produced by nematode H. polygyrus L4 stage male and female developed in colitic milienu. Mass spectrometry was used to identify proteins. OmicsBox was used to investigate the functions of the discovered proteins.
Project description:Trisomy 21 (T21) is the most frequent genetic cause of cognitive impairment. To assess the perturbations of gene expression in T21, and to eliminate the noise of the genomic variability, we studied the transcriptome of fetal fibroblasts from a pair of monozygotic twins discordant for T21. Here we show that the differential expression between the twins is organized in domains along all chromosomes that are either up- or downregulated. These gene expression dysregulation domains (GEDDs) can be defined by the expression level of their gene content, and are well conserved in induced pluripotent stem cells derived from the twins’ fibroblasts. Comparison of the transcriptome of the Ts65Dn mouse model of DS and wild-type, also showed GEDDs along the mouse chromosomes that were syntenic in human. The GEDDs correlate with the lamina-associated (LADs) and replication domains of mammalian cells. The overall LADs position was not altered in trisomic cells. However, the H3K4me3 profile of the trisomic fibroblasts was modified and accurately followed the GEDD pattern. These results suggest that the nuclear compartments of trisomic cells undergo modifications of the chromatin environment influencing the overall transcriptome and that GEDDs may therefore contribute to some T21 phenotypes. DNaseI HS mapping in monozygotic twins discordant for trisomy 21 (2 replicates of each).
Project description:The Crabtree effect, in which fermentative metabolism is preferred at the expense of respiration, is a hallmark of budding yeast’s glucose response and a model for the Warburg effect in human tumors. While the glucose-responsive transcriptional repressors Mig1p and Mig2p play well-characterized roles in the Crabtree effect, little function for the related Mig3p transcription factor has been uncovered despite numerous investigations of laboratory yeast strains. Here we studied a wild isolate of Saccharomyces cerevisiae to uncover a critical role for Mig3p that has been lost in S288c-derived laboratory strains. We found that Mig3p affects the expression of hundreds of glucose-responsive genes in the oak strain YPS163, both during growth under standard conditions and upon ethanol treatment. Our results suggest that Mig3p may act as a multifunctional activator/repressor that plays separate roles under standard versus stress conditions, but this function has been largely lost in the lab strains. Population analysis suggests that the lab strain, and several wild strains, harbor mutations that diminish Mig3p function. Thus, by expanding our attention to multiple genetic backgrounds, we have uncovered an important missing link in a key metabolic response. We performed a series of microarray experiments comparing the gene expression response of unstressed or EtOH stressed wild-type or mig3∆ strains in either the BY4741 or YPS163 background (biological triplicates). We also compared gene expression for in reciprocal hemizygous strains (YPS163/BY4741mig3∆ and YPS163mig3∆/BY4742; biological duplicates). Lastly, we measured the gene expression of BY4741 over-expressing BY_MIG3 via galactose induction (biological triplicates).
Project description:ppGpp accumulation caused by ectopic expression of RelA in Saccharomyces cerevisiae gave rise to marked changes in gene expression with both upregulation and downregulation, including changes in mitochondrial gene expression. The most prominent upregulation (38-fold) was detected in the function-unknown hypothetical gene YBR072C-A, followed by many other known stress-responsive genes. ppGpp acuumulation resulted in enhancement of tolerance against various stress stimuli, such as osmotic stress, ethanol, hydrogen peroxide, and high temperature. A two chip study using total RNA recoverd from the Saccharomyces cerevisiae TN2080 (accumulating ppGpp) and TN2077 (vector control) grown to mid-growth phase (8h) in SC-uracil medium. The plasmid pYC2/CT (V5-epitope tag vector) was used as a vector to express Sj-RSH.
Project description:The differentiation of cells into distinct cell types, each of which is heritable for many generations, underlies many biological phenomena. White and opaque cells of the fungal pathogen Candida albicans are two such heritable cell types, each thought to be adapted to unique niches within their human host. To systematically investigate the differences between the two cell types, we performed strand-specific massively-parallel sequencing of RNA from C. albicans white and opaque cells. Combining the resulting data from both cell types, we first substantially re-annotated the C. albicans transcriptome, finding 1443 novel coding and non-coding transcriptionally active regions. Using the new annotation, we compared differences in transcript abundance between the two cell types with the genomic regions bound by the master regulator of the white-opaque switch (Wor1). We found that the revised transcriptional landscape considerably alters our understanding of the circuit governing differentiation. In particular, we can now resolve the poor concordance between binding of the master regulator and the differential expression of adjacent genes, a discrepancy observed in many other studies of cell differentiation. More than one third of the Wor1-bound differentially-expressed transcripts were previously unannotated, which explains the formerly puzzling presence of Wor1 at these positions along the genome. Indeed, many of these newly identified Wor1-regulated genes are non-coding and transcribed antisense to coding transcripts. We also found that 5' and 3' untranslated regions (UTRs) of mRNAs in the circuit are unusually long and that 5' UTRs often differ in length between white and opaque cells. These observations suggest that the use of alternative promoters is widespread in the circuit and that important regulatory information is carried in the long UTRs. Further analysis revealed that the revised Wor1 circuit bears several striking similarities to the Oct4 circuit that specifies the pluripotency of mammalian embryonic stem cells. Additional characteristics shared with the Oct4 circuit suggest a set of general hallmarks characteristic of heritable differentiation states in eukaryotes. RNA-Seq was applied to Candida albicans white and opaque cells to identify novel transcripts and UTRs that are differentially regulated between the two cell types. Two biological replicates each of white and opaque cell cultures. One of the white cell RNA samples was split just after isolation to allow a comparison of the poly(A)-selection and ribo-depletion sample preparation strategies.
Project description:Mouse WT129 ESCs and differentiated from them within 12 days glutamatergic neurons were used for the ChiP-seq experiment with an antibody against Sox2 protein. ES stands for mESCs, NP for neurons, inputs are provided for both cell types.
Project description:ChIP-seq was performed to assess changes in the activity of the MYC SE upon deletion of its modules or CTCF binding sites. Immunoprecipitation of crosslinked chromatin was performed with antibodies directed against H3K27Ac (Diagenode C15410196), H3K9Ac (Diagenode C15410004), H3K4me3 (Diagenode C15410003), RUNX1 (Abcam ab23980) or CTCF (Cell Signalling, 2899S). Crosslinks were reversed overnight at 65°C in the presence of proteinase K (New England Biolabs). De-crosslinked material was purified using a QIAGEN PCR Purification Kit. The purified DNA was processed according to the Nextflex ChIP Sample Preparation Protocol (Perkin Elmer) or the Microplex library preparation kit V2 (Diagnode C05010013) and sequenced on the Illumina NovaSeq6000 platform.
Project description:We used bs-ATLAS-seq to comprehensively map the genomic location and assess the DNA methylation status of human full-length LINE-1 elements (L1) in the genome of 2102Ep cells (E-MTAB-10895). We also achieved targeted nanopore sequencing to assay DNA methylation over a subset of loci (E-MTAB-12247). To further study the link between L1 DNA methylation and expression, we performed, in the same cell line, RNA-seq (E-MTAB-12246), as well as YY1 and H3K4me3 ChIP-seq (this dataset).