Gene expression analysis during heat shock in Drosophila melanogaster KC167 cells and 3rd instar dp cn bw larvae
ABSTRACT: Gene expression analysis of Drosophila melanogaster Kc167 cells and 3rd instar dp cn bw larvae, under heat shock and non-heat shock conditions (room temperature) using Nimblegen arrrays (GPL8443) Overall design: A 12 chip study using total RNA recovered from heat shocked or non-heat shocked (room temperature) samples in either cells or larvae, 3 biological replicates for each treatment
Project description:Gene expression analysis of Drosophila melanogaster 3rd instar hsf4 cn bw larvae, under heat shock and non-heat shock conditions (room temperature) using Nimblegen arrrays (GPL8443) Overall design: total RNA recovered from heat shocked or non-heat shocked (room temperature) samples in hsf4 larvae, 3 biological replicates for each treatment
Project description:Expression analysis of Heat Shock in Drosophila melanogaster KC167 Cells and 3rd instar dpcnbw larvae, and room temperature expression levels using Nimblegen arrrays (GPL8443) A 12 chip study using total RNA recovered from Heat shocked or non-heat shocked (room temperature) samples in either cells or larvae, 3 technical replicates for each treatment
Project description:ecdysone treated KC167 cells during Heat Shock and at Room Temperature Overall design: 2-colour microarray desgin using 9 spotted oligonucleotide microarrays printed at the Canadian Drosophila Microarray Centre. Total RNA from Ecdysone treated KC167 cells were compared to Total RNA from an EtOH (the solvent for ecdysone) control sample pool.
Project description:Whole-genome analysis of heat shock factor binding sites in Drosophila melanogaster. Heat shock factor IP DNA from non-shock (room temperature) Kc 167 cells compared to whole cell extract on Agilent 2x244k tiling arrays. Overall design: Heat Shock Factor IP vs whole cell extract from non-shock (room temperature) Kc cells.
Project description:In Drosophila larvae, acquired synaptic thermotolerance following heat shock has previously been shown to correlate with the induction of heat shock proteins (Hsps) including HSP70. We tested the hypothesis that synaptic thermotolerance would be significantly diminished in a temperature-sensitive strain (hsf4) which has been reported not to be able to produce inducible Hsps in response to heat shock. Contrary to our hypothesis, considerable thermoprotection was still observed at hsf4 larval synapses following heat shock. To investigate the cause of this thermoprotection, we conducted DNA microarray experiments to identify heat-induced transcript changes in these organisms. Transcripts of the hsp83, dnaJ-1(hsp40) and gstE1 genes were significantly up-regulated in hsf4 larvae after heat shock. In addition, increases in the levels of Hsp83 and DnaJ-1 proteins but not in the inducible form of Hsp70 were detected by Western blotting. The mode of heat shock administration differentially affected the relative transcript and translational changes for these chaperones. These results indicate that the compensatory up-regulation of constitutively expressed Hsps, in the absence of the synthesis of inducible Hsps including HSP70, could still provide substantial thermoprotection to both synapses and the whole organism. Two strains were used in this study; the mutant cn bw hsf4 strain, and the dp cn bw cl control strain from which the mutant was derived. For each strain, co-reared larvae were separated into a control group and a treatment group at random. RNA from the heat-shocked larvae and from the respective controls were directly compared on cDNA microarrays (GPL311). Two independent biological samples from each genotype were collected. In addition, an inter-strain comparison was made using the RNA from the untreated controls from each genotype. This was also performed in duplicate.
Project description:Circadian clocks are temporally aligned to the environment via signals, or Zeitgebers, such as daily light and temperature cycles, food availability, and social behavior. In this study, we show that genome-wide expression profiles from temperature-entrained flies show a dramatic difference in the presence or absence of a thermocycle. Whereas transcription appears to be modified globally by changes in temperature, there is a specific set of transcripts that continue to oscillate in constant conditions following temperature entrainment. These transcripts show a significant overlap with a previously defined set of transcripts oscillating in response to a photocycle. Further, these overlapping transcripts maintain the same mutual phase relationships after entrainment by temperature or light. Comparison of the collective temperature- and light-entrained circadian phases indicates that natural environmental light and temperature cycles cooperatively entrain the circadian clock. These findings suggest that a single transcriptional clock in the adult fly head is able to integrate information from both light and temperature. Keywords: circadian time course Overall design: cn bw flies that had been reared in constant darkness initially at 25 C and later in a 12-hr 18 C/ 12-hr 25 C thermocycle for more than 4 days were harvested every four hours during an additional two days at constant 25 C (also indicated as ambient/ambient or AA days 1 and 2) . Relative to time AA0 as the time of the onset of the subjective cryophase during constant 25 C, samples were collected in a AA2-AA6-AA10-AA14-AA18-AA22-AA26-AA30-AA34-AA38-AA42-AA46 schedule. Heads were isolated by breaking up frozen flies and passing them through a set of sieves. RNA was prepared using guanidine-thiocyanate extraction followed by purification over a CsCl gradient. Additional purification of the RNA samples was achieved by applying them to Rneasy columns (Qiagen). Biotin-labeled cRNA probe was generated from 25 μg of purified RNA and hybridized as described previously (Wijnen H, Naef F, and Young MW, Methods Enzymol. 2005; 393: 341-365). For more information see also http://biorhythm.rockefeller.edu
Project description:Whole-genome analysis of heat shock factor binding sites in Drosophila melanogaster. Heat shock factor IP DNA or Mock IP DNA from heat shocked Kc 167 cells compared to whole cell extract on Agilent 2x244k tiling arrays. Overall design: Heat Shock Factor IP or mock IP vs whole cell extract from heat shocked Kc cells.
Project description:Growth and transcriptional profiles of the barophilic methanarchaeon Methanocaldococcus jannaschii were studied at temperatures up to 98C and pressures up to 500 atm. Application of 500 atm of hyperbaric pressure shifted the optimal growth temperature upwards, and heat shock from 88C to 98C at 500 atm resulted in termination of growth. Pressure shock of M. jannaschii from 7.8 to 500 atm over 15-min, the first pressure upshift reported for a barophile, did not accelerate growth. Transcriptional profiles indicated a similar pressure response under growth and heat shock at 500 atm and pressure shock to 500 atm suggesting that the commonly affected genes are important for high-pressure adaptation. Factorial microarray design allowed de-convolution of the interacting effect of elevated pressure and heat shock on expression profiles, thus suggesting genes that may contribute to the organism’s survival in the turbulent in situ conditions of deep-sea hydrothermal vents. Keywords: stress response, time course, high pressure, heat shock, pressure shock Overall design: The cDNA arrays that were used contained three technical replicate spots for each ORF in the genome of M. jannaschii. Twenty-four sub-arrays were hybridized with the following pairs of samples: four biological-replicate pairs of samples extracted from 7.8 atm and 500 atm, both without heat shock, four biological-replicate pairs of samples extracted from 7.8 atm without heat shock and 7.8 atm with heat shock (0.5 h and 1.5h), four biological-replicate pairs of samples extracted from 500 atm without heat shock and 500 atm with heat shock (0.5 h and 1.5h), and four biological-replicate pairs of samples extracted from 7 atm and 500 atm, both heat shocked (0.5 h). Each of the four pairs of biological replicates included two pairs of dye-swapped samples. Six sub-arrays were hybridized with the following pairs of samples: three biological-replicate pairs of samples extracted from cells pressure shocked to 500 atm (0.5 h and 1.5 h) and from cells grown at 7 atm without heat shock.