Project description:To examine the effects of microRNAs including miR-223 on murine neutrophil function, small RNAs cloning and sequencing from neutrophils were performed. Neutrophils were isolated from a wild-type mouse and total RNAs were prepared. Small RNAs were cloned and sequenced by Solexa/Illumina genome analyzer.
Project description:Array-based analysis of genome-wide DNA methylation changes induced by the demethylating drugs azacytidine and decitabine on HCT116 and HL60 cells for 24 hours
Project description:To identify potential PTTG1-targeted genes, total RNA from PTTG1+/+ and PTTG1-/- HCT 116 cells was isolated using the RNeasy Kit (Qiagen) and analyzed using Illumina microarrays (HumanHT-12_V4). Raw data were processed according to the manufacturerâ??s standard procedure and further analyzed using Partek 6.5. PTTG1-/- and parent HCT116 cells. Experiment is performed in triplicate
Project description:The goal of the study was to investigate the effect of inducible ZXDA expression in HCT116 cell line. HCT116 clones expressing inducible versions of human ZXDA gene (2 clones wild-type; 2 clones ERP386-388AAA) were incubated with or without doxycycline.
Project description:Hypoxia is associated with poor prognosis in most solid tumors due to its multiple effects on therapy resistance, angiogenesis, apoptotic resistance, and tumor invasion/metastasis. Here we used a comprehensive omics profiling to investigate hypoxia-regulated gene expression in HCT116 colon cancer cells. Quantitative analyses of proteome and secretome were performed in HCT116 cells cultured under hypoxic or normoxic conditions. A total of 5,700 proteins were quantified in proteome analysis and 722 proteins were quantified in secretome analysis. Both datasets were combined with the transcriptome and translatome datasets for further analysis. Verification of candidate proteins/genes in this integrated omics analysis was performed using immunoblotting and quantitative real-time RT-PCR analyses. We also performed polysome profiling to assess changes in translational efficiency of hypoxia-induced genes. Notably, several genes were differently regulated at the transcriptional and translational levels in HCT116 cells during hypoxia. Bioinformatics analysis suggested that hypoxia regulates translation of genes involved in extracellular matrix organization, extracellular exosomes, and protein processing in endoplasmic reticulum. Aberrations in these metabolic pathways appear to be correlated with an increased risk of tumor invasion/metastasis.
Project description:array-based analysis of genome-wide DNA methylation changes induced by the demethylating drugs azacytidine and decitabine on HCT116 cells for 24 hours
Project description:Wdr13 has been implicated in memory and mental disorders, particularly in X-linked intellectual disability (XLID) in animal studies and in humans. The exact molecular function of Wdr13 is still largely unknown. In order to find out the role of Wdr13 in brain, we chose to characterize Wdr13 gene knockout mice and dissect the molecular mechanisms behind its action. We found that Wdr13 expresses majorly in hippocampal formation, cerebral cortex, cerebellum and striatum. Wdr13-/0 mice show mild anxiety but fared better than the wild type mice in memory tests. This phenotype was found to be correlated with increase in level of synaptic genes like Syn1, Camk2a, Sv2b and voltage gated ion channels. Interestingly, exposure to three/four weeks of social isolation caused symptoms of major depressive disorder in the Wdr13-/0 mice which was associated with loss in dendritic branches in hippocampal CA1 neurons and significant decrease in synaptic genes like Syn1, Rab3a, Nrxn2. We found GATA1, a common negative Transcription Factor for the above-mentioned and also a marker of MDD (Major Depressive Disorder) to be upregulated in the Wdr13-/0 mice after social isolation. WDR13 overexpression caused decrease in GATA1 expression in IMR32- human neuroblastoma cell line indicating a possible regulation of GATA1 by WDR13. Wdr13 thus becomes an important candidate gene to be involved in Major Depression.
Project description:Mutations in the DJ-1 (Park7) gene cause autosomal recessive Parkinson's disease in humans, but the function of the DJ-1 protein is poorly characterized. In an effort to understand more about the biology of DJ-1, we performed iTRAQ analyses on subcellular fractions enriched from DJ-1 knockout rat and mouse brains. We generated iTRAQ datasets for mitochondria and cytosol enriched fractions from 6-month-old rat brains, and the cytosol enriched fraction from 14-15-month old mouse brains. Our subsequent analyses of these datasets led to our discovery that the Hexokinase 1 protein was increased in the cytosol components of both DJ-1 knockout species.
Project description:One of the strongest associated type 2 diabetes (T2D) loci reported to date resides within the TCF7L2 gene. Previous studies point to the T allele of rs7903146 in intron 3 as the causal variant at this locus. To aid in the identification of the actual gene(s) under the influence of this variant, we first generated a CRISPR/Cas9 mediated 1.4kb deletion of the genomic region harboring rs7903146 in the HCT116 cell line followed by global gene expression analysis. HCT116 cells with or without a CRISPR/Cas9 mediated1.4kb deletion of the genomic region harboring the SNP rs7903146 were analyzed for expression, with 3 replicates per condition (DEL vs WT). We observed 99 genes with significant differential expression (FDR cut-off=10%) and an effect size of at least two-fold. We then carried out high-throughput chromosome conformation capture assays in the HCT116 and NCM460 cell lines and in colon tissue (see experiment E-MTAB-4845) in order to ascertain which of these perturbed genesâ promoters made consistent physical contact with the genomic region harboring the variant. This revealed just one gene, ACSL5, which resides in the same topologically associating domain as TCF7L2.