HeLa Cells infected with Shigella in the presence/absence of Staurosporin
ABSTRACT: Question Addressed: What is the host response (specifically apoptosis pathways) to Shigella infection and what is the contribution of MxiE to the host response? The arrays are apoptosis Exon hit arrays, thus splice variation can be determined. Array data from HeLa cells that remained Uninfected or were infected with wildtype S. flexneri serotype 2a strain 2457T or an isogenic mixE mutant. These groupings are further divided such that Uninfected or Infected cells remained untreated or were treated with the apopotosis inducer staurosporine (STS). The design of the experiment was a time course and samples were harvested at the indicated time points (hr). All hybridizations were conducted against a common reference sample that was made from pooled samples of normal uninfected healthy HeLa cells. Infection: HeLa cells were infected with wt Shigella (wt), mxiE mutant (mixE) or were left uninfected (uninfected) Compound Based Treatment: Cultures were treated with Staurosporin (STS) or were left untreated (untreated) Infection time: Cells were harvested at the indicated time after treatment 20 Samples
Project description:Study of host response in mice to uropathogenic E. coli (UPEC) infection and the contribution of deletion of the Cnf and HlyA1 toxins to the host response. Mice were infected with UPEC in the bladder for 24 hrs and RNA was isolated from whole bladder to analyze on Agilent whole mouse 2-channel arrays against normal untreated mouse bladder RNA. Biological replicates were analyzed for each condition. Infection: genotype of E. coli CP9 strain strain_or_line_design
Project description:Gene expression was studied in samples from the stomach of Rhesus monkeys that have been infected with Helicobacter pylori and treated with a known carcinogen (Ethyl-nitro-nitrosoguanidine (ENNG)) that is similar to known nitrosamine dietary carcinogens. The samples represent the following: Biopsy 4689 is from animal 81G taken 55 months after inoculation with H. pylori and the start of the carcinogen ENNG Biopsy 4691 is from animal 63G taken 55 months after inoculation with H. pylori and the start of the carcinogen ENNG Biopsy 4709 is from animal 92F taken after 55 months of observation and no treatment (control). Gastric biopsies were obtained at the indicated times and RNA was harvested from the sample. All hybridizations were conducted against a common reference that was prepared using commercially obtained RNA isolated from the stomach of uninfected monkeys. Infection: Inoculated with H. pylori and treated with carcinogen ENNG (Treated) or control
Project description:INTRODUCTION AND OBJECTIVE: Obstruction and other processes that lead to renal damage can have different effects in adult versus developing kidneys. These differences are important for understanding pathophysiology, but also could influence selection of biomarkers used for detection of renal damage that could be used to guide intervention. In mice, significant renal development occurs postnatally, providing a model for understanding kidney development. To date, gene expression changes that occur mouse kidney growth and development postnatally have been poorly characterized. We describe here a comprehensive gene expression of the developing mouse kidney based on microarray profiling. METHODS: C57BL/6 mice were sacrificed and kidneys were harvested at embryonic day E19.5, and postnatal days P1, P3, P5, P7, P10, P14, P21, P28 and P35. RNA was extracted from kidneys and transcript profiling was performed using Agilent microarrays. Transcripts that undergo abrupt transitions in expression level over the time course were identified using StepMiner analysis (Sahoo, 2007). Ingenuity pathway analysis (IPA) was used to analyze the biological function and gene networks of gene expression data. RESULTS: Transcript levels for 13645 probes (representing 12769 genes) were modulated significantly over the time course, with 6949 up-regulated and 6696 down-regulated. IPA was used to identify genetic pathways, networks and functions significantly altered over the time course. Interestingly, in days P10 to P14 gene functions significantly altered were up-regulated exclusively with the functions represented including lipid metabolism, small molecule biochemistry and molecular transport. Between days P5 to P7 down-regulated genes predominated with enriched functions including, gene expression, cell cycle, protein synthesis and embryonic development. For P14 to P21 enriched functions included DNA replication, recombination and repair, cell cycle, tissue development, cellular assembly and organization, protein trafficking and cell morphology. In the late stages (P21 to P28) functional enrichment was seen for cell-to-cell signaling, tissue development, cellular movement. CONCLUSIONS: This study provides the most comprehensive temporal survey of postnatal kidney development to date. This data set provides a framework for interpreting nephropathies, such as those induced by congenital obstruction. time series design
Project description:Study of host response in mice to uropathogenic E. coli (UPEC) infection and the contribution of deletion of the Cnf and HlyA1 toxins to the host response. Mice were infected with UPEC in the bladder for 24 hrs and RNA was isolated from whole bladder to analyze on Agilent whole mouse 2-channel arrays against normal untreated mouse bladder RNA. Biological replicates were analyzed for each condition. Infection: genotype of E. coli CP9 (UPEC) strain strain_or_line_design
Project description:Genome rearrangements are associated with eukaryotic evolutionary processes ranging from tumorigenesis to speciation. Such rearrangements are especially common following the shock of interspecific hybridization, and some of these could be expected to have strong selective value. To test this expectation we created de novo interspecific yeast hybrids between two diverged but largely syntenic species, Saccharomyces cerevisiae and S. uvarum, then experimentally evolved them under continuous ammonium limitation. We discovered that a characteristic interspecific genome rearrangement arose multiple times in independently evolved populations. We uncovered nine different breakpoints, all occurring in a ~1 kb region of chromosome 14, and all producing an “interspecific fusion junction” within the MEP2 gene coding sequence, such that the 5’ portion derives from S. cerevisiae and the 3’ portion derives from S. uvarum. In most cases the rearrangements altered both chromosomes, resulting in what can be considered to be an introgression of a several-kb region of S. uvarum into an otherwise intact S. cerevisiae chromosome 14, while the S. uvarum chromosome 14 experienced an interspecific reciprocal translocation at the same breakpoint within MEP2, yielding a chimaeric chromosome. The net result is the presence in the cell of two MEP2 fusion genes having identical breakpoints. Given that MEP2 encodes for a high-affinity ammonium permease, that MEP2 fusion genes arise repeatedly under ammonium-limitation, and that three independent evolved isolates carrying MEP2 fusion genes are each more fit than their common ancestor, the novel MEP2 fusion genes are very likely adaptive under ammonium limitation. Our results suggest that when homoploid hybrids form, the admixture of two genomes enables swift and otherwise unlikely evolutionary innovations. Furthermore, the architecture of the MEP2 rearrangement suggests a model for rapid introgression, a phenomenon seen in numerous eukaryotic phyla, that does not invoke repeated backcrossing to one of the parental species. Nomenclature: GSY86 TimeZeroInoculum = ancestral interspecific hybrid used to inoculate ammonium-limited chemostats into 3 replicate vessels A, B, C. 150gen = various single-clone isolates from 150 generations of evolutions from vessels A, B or C. 200gen = various isolates from 200 generations of evolutions from vessels A, B or C. Logical Set: Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc.
Project description:Darinaparsin (DPS) is an arsenic cytotoxin with a more favorable toxicity profile than the established anti-leukemic drug, arsenic trioxide (ATO). Here we report effects of DPS on a variety of solid tumor cell lines under normoxia as well as hypoxia, the latter an important characteristic of the tumor microenvironment. Using MTT and clonogenic assays, we demonstrated that DPS had potent cytotoxic activities under both normoxia and hypoxia, with IC50??s ~ 2- to 3-fold lower than ATO. Xenograft studies using tumor cell lines as well as patient-derived tumor tissues implanted under the renal capsule in mice confirmed the anti-tumor activity of DPS in vivo. DPS was also a more potent radiosensitizer under hypoxia than ATO in vitro, and sensitized solid tumors to radiation in vivo at doses that had no systemic toxicities. Interestingly, in contrast to previous reports of DPS effects on leukemic cells under normoxia, DPS-induced killing of hypoxic solid tumor cells was not dependent on ROS generation and oxidative damage. Instead, cDNA microarray analysis suggested that DPS inhibited oncogene- (such as RAS and MYC) associated gene expression. In fact, compared to normal mouse embryonic fibroblasts (MEFs), oncogene (RAS/E1A) transformed MEFs were markedly more sensitive to DPS-induced apoptosis under both normoxia and hypoxia. Altogether, these results demonstrate that DPS has significant and preferential cytotoxicity against solid tumor as compared with normal cells, and is an effective radiosensitizer. Since DPS is in early clinical development as a single agent, these findings have near term translational potential. time_series_design
Project description:INTRODUCTION AND OBJECTIVE: Loss of renal function is often the impetus for operative intervention in renal obstruction. Obstructive nephropathy is characterized discrete morphological and physiologic changes including tubular dilatation, apoptosis, and atrophy, as well as interstitial cellular infiltration and progressive interstitial fibrosis. We hypothesize that there are gene expression alterations correlated with obstructive nephropathy that could serve as biomarkers for early intervention. METHODS: C57BL/6 mice were subjected to Unilateral Urethral Obstruction (UUO) or sham surgery at postnatal day 21 of life, and kidneys were harvested after 1, 2, 5 and 9 days postoperatively. RNA was extracted from kidneys and comprehensive gene expression profiling was performed with Agilent microarrays. We used biological replicates: 13 UUO replicates, 9 sham replicates. 2-Channel microarrays were hybridized using universal reference in the Cy3 channel. Ingenuity pathway analysis was used to analyze the biological function and gene networks of gene expression data. RESULTS: Microarray analysis revealed more than 1800 transcripts that were up- or down-regulated over days 1 through 9 following obstruction, and included many transcripts reported previously (FOS, CD44, CLU, SPP1 and EGF). Pathway analysis revealed significant enrichment of transcripts in cell activation/differentiation, immune/inflammatory responses, cell cycle, metabolic process and transport. Interestingly, IPA network analysis showed that transcriptional regulatory pathway involving CEBPB/HNF4A is involved in obstructive nephropathy. CONCLUSIONS: Transcript profiling identified numerous gene expression changes following UUO and suggests a role for CEBPB/HNF4A pathway in obstructive nephropathy. This dataset provides a foundation for development of biomarkers for obstructive nephropathy. Time: day samples were collected post operation Somatic Modification: mice were operated on to generate urethral obstruction (Obstructed/Control) time_series_design
Project description:We created a multi-species microarray platform, containing probes to the whole genomes of seven different Saccharomyces species, with very dense coverage (one probe every ~500 bp) of the S. cerevisiae genome, including non-S288c regions, mitochondrial and 2 micron circle genomes, plus probes at fairly dense coverage (one probe every ~2,100 bp) for each of the genomes of six other Saccharomyces species: S. paradoxus, S. mikatae, S. kudriavzevii, S. bayanus, S. kluyveri and S. castellii. We performed array-Comparative Genomic Hybridization (aCGH) using this platform, examining 83 different Saccharomyces strains collected across a wide range of habitats; of these, 69 were widely used commercial S. cerevisiae wine strains, while the remaining 14 were from a wide range of other industrial and natural habitats. Thus, we were able to sample much of the pan-genome space of the Saccharomyces genus. We observed interspecific hybridization events, introgression events, and pervasive copy number variation (CNV) in all but a few of the strains. These CNVs were distributed throughout the strains such that they did not produce any clear phylogeny, suggesting extensive mating in both industrial and wild strains. To validate our results and to determine whether apparently similar introgressions and CNVs were identical by descent or recurrent, we also performed whole genome sequencing on nine of these strains. These data may help pinpoint genomic regions involved in adaptation to different industrial milieus, as well as shed light on the course of domestication of S. cerevisiae. Control arrays of the following types are included in the dataset: (1) "Self-self" hybridizations (called "Self cntrl" in the dataset), where the reference DNA mix was labeled with either Cy3 or Cy5 (in separate reactions) and then mixed and hybridized. (2) Reference sub-pool hybridizations: the mix of 6 non-cerevisiae strains' DNA was labeled with Cy5, mixed with the final Reference pool (labeled in Cy3) and hybridized (called "OtherSppCompletePool" control) or the mix of 42 S. cerevisiae strains' DNA was labeled with Cy5, mixed with the final Reference pool (labeled in Cy3) and hybridized (called "Scer only ref pool") (3) Single Saccharomyces species hybridizations, where each of the 6 non-cerevisiae species, and also the S288c S. cerevisiae lab strain, was labeled with Cy5, mixed with the final Reference pool (labeled in Cy3) and hybridized (these arrays are called by the strain number followed by the species nickname). Four separate replicates of the self-self hybridizations were performed, and duplicate arrays (done on different days using different DNA preparations) were performed for three of the wine strains: GSY2175, GSY2176 and GSY2196.
Project description:Hybrid progeny can enjoy increased fitness and stress tolerance relative to their ancestral species, a phenomenon known as hybrid vigor. Though this phenomenon has been documented throughout the Eukarya, evolution of hybrid populations has yet to be explored experimentally in the lab. To fill this knowledge gap we created a pool of Saccharomyces cerevisiae and S. bayanus homoploid and aneuploid hybrids, and then investigated how selection in the form of incrementally increased temperature or ethanol impacted hybrid genome structure and adaptation. During 500 generations of continuous ammonia-limited, glucose-sufficient culture, temperature was raised from 25C to 46??C. This selection invariably resulted in nearly-complete loss of the S. bayanus genome, although the dynamics of genome loss differed among independent replicates. Temperature-evolved isolates were significantly more thermal tolerant and exhibited greater phenotypic plasticity than parental species and founding hybrids. By contrast, when the same hybrid pool was subjected to increases in exogenous ethanol from 0% to 14%, selection favored euploid S. cerevisiae x S. bayanus hybrids. Ethanol-evolved isolates exhibited significantly greater ethanol tolerance relative only to S. bayanus and one of the founding hybrids tested. Adaptation to thermal and ethanol stress manifested as heritable changes in cell wall structure demonstrated by resistance to zymolyase or micafungin treatment. This is the first study to show experimentally that the fate of interspecific hybrids critically depends on the type of selection they encounter during the course of evolution. Array-CGH was performed on the S. cerevisiae parent strain CEN.PK (GSY2160), the S. bayanus parent strain CBS7001 (GSY2161) and on the F1 interspecific hybrid resulting from mating the 2 parents (GSY2168). Additionally, three rare viable spores obtained after sporulation of the F1 were assayed by array-CGH (F2a, F2b, F2c). A large pool of F2 spores (and probably some number of F1 hybrid cells) were subjected to gradually increasing temperatures, in three independent vessels, with populations sampled at various generation times. Likewise, the same pool was used to found populations in an additional three independent vessels, which were then subjected to gradually increasing ethanol concentrations (at constant temperature). Array-CGH was performed on three different clones from each of the three temperature vessels at the final 500 generation time point (T500 clones). Biological replicates of the T500 clones were performed (T500-new). Two self-self array-CGH hybridization controls were also performed (self-control). Array-CGH was performed on one clone from each of the three ethanol vessels taken at the 400 generation timepoint (EtOH400gen clones).
Project description:Array data from HeLa cells that remained Uninfected or were infected with wildtype S. flexneri serotype 2a strain 2457T. These groupings are further divided such that Uninfected or Infected cells remained untreated or were treated with the apopotosis inducer staurosporine (STS). The design of the experiment was a time course and samples were harvested at the indicated time points (hr). All hybridizations were conducted against a common reference sample. time_series_design