Project description:SecDF is a highly conserved accessory protein of the Sec-translocase located in the cytoplasmic membrane. The deletion mutant (delta Bc4405) of Bacillus cereus ATCC14579 shows multiple phenotypic changes, including aberrent cell division starting at transitionary phase. To understand the underlying processes genotypic profiling was carried out at 3h and 4h after inoculation. The morphology of the mutant seems to be more severe if glucose is added to the LB medium, thus all cultures contained 1% glucose.
Project description:Transcriptional profiling comparing Escherichia coli simultaneously exposed to tellurite and CTX with untreated control cells; Tellurite with control; CTX with control Three-condition experiment, antibacterial (tellurite; CTX or tellurite/CTX) vs. Untreated control cells. Biological replicates: 3 control, 3 toxicants exposed cells, independently grown and harvested. One replicate per array.
Project description:E. coli cultures were exposed to tellurite 0.5 µg/ml during 15 min. Total RNA was extracted and cDNA labeled probes were generated by reverse transcription using Alexa 555 and Alexa 647 fluorophores. These probes were used to hybridize genomic slides containing genomic arrays to determine global transcriptional changes. Two-conditions experiment, antibacterial vs. Untreated control cells. Biological replicates: 2 control, 2 toxicants exposed cells, independently grown and harvested. One replicate per array. Dye swap conditions.
Project description:Bacillus subtilis encodes seven extracytoplasmic function (ECF) sigma factors. Three (sigma M, sigma W and simga X) mediate responses to cell envelope active antibiotics. The functions of sigma Y, sigma Z, sigma V, and YlaC remain largely unknown, and strong inducers of these sigma factors and their regulons have yet to be defined. Here, we define transcriptomic and phenotypic differences under non-stress conditions between strains carrying deletions in all seven ECF sigma factor genes (Δ7ECF), a sigMWX triple mutant (∆MWX), and the parental 168 strain. Our results identify >80 genes as at least partially dependent on ECF sigma factors and, as expected, most of these are dependent on sigma M, sigma W or sigma X which are active at a significant basal level during growth. Several genes, including the eps operon encoding enzymes for exopolysaccharide (EPS) production, were decreased in expression in Δ7ECF but affected little if at all in ΔMWX. Consistent with this observation, Δ7ECF (but not ∆MWX) showed reduced biofilm formation. Extending previous observations, we also note that ∆MWX is sensitive to a variety of antibiotics and Δ7ECF is either as sensitive as, or slightly more sensitive than, the ΔMWX strain to these stressors. These findings emphasize the overlapping nature of the seven ECF s factor regulons in B. subtilis, confirm that three of these (sigma M, W or X) play the dominant role in conferring intrinsic resistance to antibiotics, and provide initial insights into the roles of the remaining ECF sigma factors. Strains WT vs. ΔMWX, WT vs. Δ7ECF, Δ7ECF vs. ΔMWX. Each experiment was conducted three times using three independent total RNA preparations (biological triplicates). For each paried comparison, one sample was was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647. For each comparison, one replicate was performed with dyewap with the same RNA preparation.
Project description:To define the ECF sigma sigV - regulated genes during log growth phase in LB media under induction conditions for sigV The seven extracytoplasmic function (ECF) sigma (σ) factors of Bacillus subtilis are broadly implicated in resistance to antibiotics and other cell envelope stressors mediated, in part, by regulation of cell envelope synthesis and modification enzymes. We here define the regulon of σV as including at least 20 operons many of which are also regulated by σM, σX, or σW. The σV regulon is strongly and specifically induced by lysozyme and this induction is key to the intrinsic resistance of B. subtilis to lysozyme. Strains with null mutations in either sigV or in all seven ECF σ factor genes (Δ7ECF) have essentially equal increases in sensitivity to lysozyme. Induction of σV in the Δ7ECF background restores lysozyme resistance, whereas induction of σM, σX or σW does not. Lysozyme resistance results from the ability of σV to activate the transcription of two operons: the autoregulated sigV-rsiV-oatA-yrhK operon and dltABCDE. Genetic analyses reveal that oatA and dlt are largely redundant with respect to lysozyme sensitivity: single mutants are not affected in lysozyme sensitivity whereas a double oatA dltA mutant is as sensitive as a sigV null strain. Moreover, the triple sigV oatA dltA mutant is no more sensitive than the oatA dltA double mutant, indicating that there are no other σV-dependent genes necessary for lysozyme resistance. Thus, σV confers lysozyme resistance by activation of two cell wall modification pathways: O-acetylation of peptidoglycan catalyzed by OatA and D-alanylation of teichoic acids by DltABCDE. Strains Δ7Pxyl-sigV + xylose vs. Δ7Pxyl-sigV - xylose, 168 + lysozyme vs. 168 - lysozyme. Each experiment was conducted 6 times using three independent total RNA preparations (biologlical triplicates). For each paired comparison, one sample was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647. For each comparison, three replicates were performed with dyeswap with the same RNA preparation.
Project description:Transcriptional profile of the rpoC G1122D mutant during log growth phase in LB medium. Bacillus subtilis W168, WT vs. rpoC (G1122D). The experiment was conducted two times using three independent total RNA preparations (biological triplicates). Each comparison was performed in dye-swap with the same RNA preparation. For each paried comparison, one sample was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647.
Project description:Transcriptional profile of the rocG gudB double null mutant during log growth phase in LB medium. Bacillus subtilis W168, WT vs. delta-rocG delta-gudB. The experiment was conducted two times using three independent total RNA preparations (biological triplicates). For each paried comparison, WT was labeled with Alexa Fluor 555 and the rocG gudB double mutant was with Alexa Fluor 647.
Project description:Transcriptional profile of the uppS-RBS (A to C) mutant during log growth phase in LB medium. Bacillus subtilis W168, WT vs. uppS-RBS (A to C). The experiment was conducted two times using three independent total RNA preparations (biological triplicates). Each comparison was performed in dye-swap with the same RNA preparation. For each paried comparison, one sample was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647.
Project description:B. cenocepacia J2315 was exposed to heat stress and to stress form reactive oxygen species. <br>To expose the cultures to heat stress, cells were grown at 37ºC to an O.D. of 0.4 to 0.45 and then transferred into a different shaking incubator at 42.5ºC, incubated for 1 hour at 150 rpm and harvested.<br>To expose cultures to oxidative stress by reactive oxygen species, cells were grown at 37ºC to an OD of 0.5. Then t butyl hydroperoxide or hydrogen peroxide solution were added to the culture at 0.001% and 0.15% final concentration. The culture was further incubated for 15 min and then harvested. <br>The expression profiles were compared to cells grown in LB medium without exposure to stress.<br>
Project description:Transcriptional profile of the sigA (G336C) mutant during log growth phase in LB medium. Bacillus subtilis W168, WT vs. sigA (G336C). The experiment was conducted two times using three independent total RNA preparations (biological triplicates). Each comparison was performed in dye-swap with the same RNA preparation. For each paried comparison, one sample was labeled with Alexa Fluor 555 and the other was with Alexa Fluor 647.