Project description:To identify differentially expressed genes by anti cancer treatments (miRNA and siRNAs) in human cancer, several cell lines (esophagus cancer and renal cancer) were subjected to Agilent whole genome microarrays. Overall design: Human cell lines (TE1, TE8, 786-O and A498) were treated with miRNA (miR-143-5p and miR-143-3p) and siRNAs (si-DONSON and si-AQP9).
Project description:In meiotic prophase, chromosomes are organized into compacted loop arrays to promote homolog pairing and recombination. Here, we probe the architecture of the mouse spermatocyte genome in early and late meiotic prophase using Hi-C. We show that early-prophase chromosomes are arranged as linear arrays of 0.8-1 Mb loops, which extend to 1.5-2 Mb in late prophase as chromosomes compact and homologs undergo synapsis. Topologically associating domains (TADs) are lost in meiotic prophase, suggesting that assembly of the meiotic chromosome axis dramatically reduces the dynamics of chromosome-associated cohesin complexes. While TADs are lost, physically-separated A and B compartments are maintained in meiotic prophase. Moreover, meiotic DNA breaks and inter-homolog crossovers preferentially form in the gene-dense A compartment, revealing a role for chromatin organization in meiotic recombination. Finally, direct detection of inter-homolog contacts genome-wide reveals the structural basis for homolog alignment and juxtaposition by the synaptonemal complex. Overall design: Hi-C detection of intra- and inter-homolog contacts in B6 x CAST F1 hybrid mouse spermatocytes. Two independent biological replicates of zygonema-stage spermatocytes, and three independent biological replicates of pachynema-stage spermatocytes.
Project description:Human DNA-methylation data have been used to develop highly accurate biomarkers of aging ("epigenetic clocks"). Recent studies demonstrate that similar epigenetic clocks for mice (Mus Musculus) can be slowed by gold standard anti-aging interventions such as calorie restriction and growth hormone receptor knock-outs. Using DNA methylation data from previous publications with data collected in house for a total 1189 samples spanning 193,651 CpG sites, we developed 4 novel epigenetic clocks by choosing different regression models (elastic net- versus ridge regression) and by considering different sets of CpGs (all CpGs vs highly conserved CpGs). We demonstrate that accurate age estimators can be built on the basis of highly conserved CpGs. However, the most accurate clock results from applying elastic net regression to all CpGs. While the anti-aging effect of calorie restriction could be detected with all types of epigenetic clocks, only ridge regression based clocks replicated the finding of slow epigenetic aging effects in dwarf mice. Overall, this study demonstrates that there are trade-offs when it comes to epigenetic clocks in mice. Highly accurate clocks might not be optimal for detecting the beneficial effects of anti-aging interventions. Overall design: Reduced representation bisulfite-sequencing (RRBS) data from public and in-house sources were processed consistently and merged to form a set of over 1000 mouse samples from various tissues and chronological ages. These data were used to investigate the relationship between chronologic age and epigenetic age.
Project description:In the present work we have applied analytical methods to map repair events in rDNA using data generated by the newly developed XR-seq genome-wide single nucleotide repair technology. We find that in human and mouse cell lines, rDNA is not subject to TCR of damage caused by UV or by cisplatin. Overall design: We perform XR-seq in mouse skin fibroblast under UV irradiation and collect cells after incubation 3 hours. For human cell lines NHF1, CSB and XPC, we perform XR-seq under UV irradiation and collect cells after incubation 1 hour. For GM12878, we perform XR-seq under cisplatin and collect cells after incubation 2 hours. Then we mapped all the reads to rDNA or DHFR. This dataset includes re-analysis of five GSE67941 Samples and two GSE82213 Samples.
Project description:Goal: Comparison of protein and phospho-protein levels in primary tumors based on activation of different AKT paralogs Methods: RPPA performed at the MD Anderson RPPA Core Facility. Detailed procedures available in methods section. Results: Focus of study compared protein and phospho-protein levels of BRAFV600E;Cdkn2a-/-;Pten-/- and BRAFV600E;Cdkn2a-/-;Pten-/-;AKT1E17K cohorts. P-FAK and paxillin were upregulated in tumors that express AKT1E17K compared with controls Conclusion: AKT1E17K has a critical role in upregulating P-FAK and paxillin Overall design: Thirty primary tumor samples included in analyses: Six BRAFV600E;Cdkn2a-/-, Six BRAFV600E;Cdkn2a-/-;Pten-/-, Six BRAFV600E;Cdkn2a-/-;Pten-/-;AKT1E17K, Six BRAFV600E;Cdkn2a-/-;Pten-/-;AKT2E17K, Six BRAFV600E;Cdkn2a-/-;Pten-/-;AKT3E17K
Project description:Background: Polycomb repressive complex 2 (PRC2) is responsible for establishing and maintaining histone H3K27 methylation during cell differentiation and proliferation. H3K27 can be mono-, di-, or tri-methylated, resulting in differential gene regulation. However, it remains unknown how PRC2 specifies the degree and biological effects of H3K27 methylation within a given cellular context. One way to determine PRC2 specificity may be through alternative splicing of Ezh2, PRC2’s catalytic subunit, during cell differentiation and tissue maturation. Results: We fully characterized the alternative splicing of Ezh2 in somatic cells and male germ cells and found that Ezh’s exon 14 was differentially regulated during mitosis and meiosis. The Ezh2 isoform containing exon 14 (ex14-Ezh2) is upregulated during cell cycle progression, consistent with a role in maintaining H3K27 methylation during chromatin replication. In contrast, the isoform lacking exon 14 (ex14D-Ezh2) was almost exclusively present in spermatocytes when new H3K27me2 is established during meiotic differentiation. Moreover, Ezh2’s transcript is normally controlled by E2F transcription activators, but in spermatocytes, Ezh2’s transcription is controlled by the meiotic regulator MYBL1. Compared to ex14-EZH2, ex14D-EZH2 has a diminished efficiency for catalyzing H3K27me3 and promotes embryonic stem cell differentiation. Conclusions: Ezh2’s expression is regulated at transcriptional and post-transcriptional levels in a cellular context-dependent manner. EZH2 variants determine functional specificity of PRC2 in histone methylation during cell proliferation and differentiation. Overall design: ChIP-seq in wild-type and Ezh2 knock-out E14 Embryonic Stem Cells for H3k27me3 and Ezh2
Project description:Differential ChIP-Seq data monitoring changes in active enhancer marks (H3K27ac sites) after treatment with siGRHL2 in MCF7 and T47D breast cancer models. Comparing sites altered by treatment with siGRHL2 after 48hours revealed these sites to be enriched for Estrogen Receptor (ER) binding. Overall design: ChIP-seq data monitoring H3K27ac in MCF7 and T47D cells after treatment with siGRHL2 and siCTRL for 48 hours.
Project description:The goal of this study is to obtain a genomic view of the Fur regulatory network under both iron replete and iron deficient conditions in Bacillus subtilis using ChIP-seq. Besides the known Fur target sites, 70 putative DNA binding sites were identified, and the vast majority had higher occupancy under iron sufficient conditions. In addition,we discovered a role for catechol degradation in bacillibactin metabolism, and provided evidence that catechol 2,3-dioxygenase can detoxify endogenously produced catechol substrates in addition to its more widely studied role in biodegradation of environmental aromatic compounds and pollutants. Overall design: To modulate intracellular iron levels, we employed a high-affinity Fe2+ exporter FrvA from L. monocytogenes to impose iron starvation. Bacillus wild-type cells (with C-terminal FLAG-tagged Fur at its native locus and an IPTG-inducible ectopic copy of frvA integrated at amyE locus) were harvested at 0 and 30 min after IPTG induction to study Fur-dependent regulation under iron sufficient and deficient conditions, respectively.
Project description:The health impacts of endocrine disrupting chemicals (EDCs) remain debated and their tissue and molecular targets are poorly understood. Here, we leveraged systems biology approaches to assess the target tissues, molecular pathways, and gene regulatory networks associated with prenatal exposure to the model EDC Bisphenol A (BPA). Prenatal BPA exposure led to scores of transcriptomic and methylomic alterations in the adipose, hypothalamus, and liver tissues in mouse offspring, with cross-tissue perturbations in lipid metabolism as well as tissue-specific alterations in histone subunits, glucose metabolism and extracellular matrix. Network modeling prioritized main molecular targets of BPA, including Pparg, Hnf4a, Esr1, Srebf1, and Fasn. Lastly, integrative analyses identified the association of BPA molecular signatures with cardiometabolic phenotypes in mouse and human. Our multi-tissue, multi-omics investigation provides strong evidence that BPA perturbs diverse molecular networks in central and peripheral tissues, and offers insights into the molecular targets that link BPA to human cardiometabolic disorders. Overall design: We utilized next-generation sequencing technologies to characterize perturbations in both the transcriptome (RNA-seq) and the epigenome (RRBS) across three tissues (white adipose tissue, hypothalamus, liver) in mouse offspring who had experienced in utero exposure to BPA