haloferax volcanii ds2 hv  (ATCC)

Journal: bioRxiv

Article Title: Rapid rewiring of an archaeal transcription factor function via flexible cis-trans interactions

doi: 10.1101/2024.11.20.624553

Figure Lengend Snippet: Haloarchaeal species exhibit differential susceptibility to oxidative stress conditions. Growth is plotted as the log10 optical density at 600 nm (OD600) over time (hours). Each curve represents the generalized additive model (GAM) fit to the raw data from at least 3 biological replicate experiments (seeded from starter cultures inoculated with separate colonies), each with 3 technical replicates (aliquots from the same starter culture). Shaded grey ribbons indicate the standard error. Error is low where ribbons are not visible. Concentrations are written above the corresponding growth curve in each panel. Concentrations surrounded by a box is that condition chosen for testing Δ rosR growth in subsequent experiments. (A) Hfx. volcanii growth under a titration of peroxide (H 2 O 2 ) concentrations. (B) Hfx. mediterranei growth under H 2 O 2 . (C) Hfx. volcanii growth under varying paraquat (PQ) concentrations. (D) Hfx. mediterranei growth under PQ.

Article Snippet: Wild-type strains Halobacterium salinarum NRC-1 ( Hs) , Haloferax volcanii strain DS2 ( Hv) , and Haloferax mediterranei ( Hm) ATCC 33500 were used in this study.

Techniques: Titration

Journal: bioRxiv

Article Title: Rapid rewiring of an archaeal transcription factor function via flexible cis-trans interactions

doi: 10.1101/2024.11.20.624553

Figure Lengend Snippet: hv RosR activates arlA1 and arlA2 , encoding structural components of the archaellum in Hfx. volcanii . (A) Top, genomic region encoding the archaellum and related motility functions; middle, ChIP-seq data for the region highlighted in teal (read depth y-axis is shown at right); bottom, chromosomal coordinates for genes in the zoomed region, with genes on the forward strand depicted on top of the line and reverse strand below the line. (B) Scatterplot of normalized counts from RNA-seq data, with parental control strain counts on the X-axis and hv Δ rosR counts on the y-axis. Genes passing the significance threshold of p < 0.05 and log2 fold change >= |1| are shown in blue, those significant genes also bound in ChIP-seq data in pink (see legend). (C) ChIP-qPCR validation of ChIP-seq data. Bar height represents the mean enrichment of hv RosR binding each site relative to a control region. Error bars represent the standard deviation from the mean of triplicate samples. hv RosR-HA enrichment is shown in salmon and PyrE2-HA parent strain control in grey. Amplicons tested are shown in orange below the bar graph (“peak 1, peak 2, peak 3”) and are set relative to chromosomal position of the corresponding genes. Asterisks indicate the statistical significance of enrichment for each peak (salmon bars) relative to the parent control (grey bars) by two-sided unpaired Student’s t-test, * p < 1.02 x 10 -3 , ** p < 4.71 x 10 -4 , *** p < 6.31 x 10 -5 . (D) Logo of the consensus binding motif detected computationally from hv RosR ChIP-seq binding site sequences. Motif position in nucleotides is given on the x-axis and bit score of per-base representation in the position weight matrix is given on the y-axis. See also Supplementary Table S4 for detailed RNA-seq, ChIP-seq, and motif data.

Article Snippet: Wild-type strains Halobacterium salinarum NRC-1 ( Hs) , Haloferax volcanii strain DS2 ( Hv) , and Haloferax mediterranei ( Hm) ATCC 33500 were used in this study.

Techniques: ChIP-sequencing, RNA Sequencing, Control, ChIP-qPCR, Biomarker Discovery, Binding Assay, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: MinD2 modulates cell shape and motility in the archaeon Haloferax volcanii

doi: 10.3389/fmicb.2024.1474570

Figure Lengend Snippet: Proposed model for MinD2 function in Haloferax volcanii. H. volcanii cells show growth-dependent cell shape morphology transitioning from rods to plate-shaped cells. We show that MinD2 has a strong effect on shape, with ∆minD2 mutant cells predominantly adopting a plate shape. We propose that the shape phenotype impacts the proper positioning of the archaellum (archaellum motor: yellow square and ArlD: orange ovals) and chemotactic machinery (dark green lines). According to this model, in ∆minD2 cells, the localization of both machineries is diffused (light green) with a very few cells showing the ability to localize the chemosensory arrays in early log phase. As the growth stage progresses, only chemosensory arrays remain diffused (dark green) and the archaellum motor is able to localize at a pole. Previous studies on MinD4 ( ; ), shows the ∆minD4 mutant cells has an effect on the positioning on archaellum and chemosensory arrays. Together, we postulate that deleting both MinD homologues ( ∆minD2minD4 ) has a synergistic effect, resulting in these cells being unable to localize both archaellum and chemosensory arrays to the pole at any stage of the growth phase.

Article Snippet: To ensure the removal of ribosomal RNA, 2 μg of input RNA was treated with a Ribopool designed specifically for Haloferax volcanii (siTOOLs) according to the manufacturer’s instructions.

Techniques: Mutagenesis

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 1 Archaeal ATAC-seq and the open chromatin landscape of H. volcanii. A, B Adaptation and optimization of the ATAC-seq assay to the archaeal context. A Distribution of TSS ratio scores (see the “Methods” section for details) for native, 0.1%- and 1%-formaldehyde ATAC-seq libraries. C Fragment length distribution in H. volcanii ATAC-seq datasets. D Estimated relative copy number of H. volcanii chromosomes. Genomic DNA was tagmented and amplified (n = 4) and normalized read coverage was estimated for each chromosome/ plasmid. The average ratios are shown. E Distribution of MACS2 ATAC-seq peaks relative to TSSs. F, G Representative browser snapshots of ATAC-seq profiles along the H. volcanii genome. F High reproducibility of H. volcanii chromatin accessibility measurements using ATAC-seq. Shown is the between-replicate correlation over TSSs in RPM (reads per million) units. I Global ATAC-seq profile over each of the five H. volcanii chromosomes. The number in brackets corresponds to the magnification of the true proportional size of plasmids relative to the main chromosome

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: Amplification, Plasmid Preparation

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 2 Absolute DNA occupancy/protection levels in H. volcanii. A–C TSS metaprofiles in different conditions (two replicates of an exponentially dividing culture, and a stationary culture). D Single-molecule map (250-bp) around a main-chromosome TSS. Black indicates unmethylated and therefore protected sites, and gray indicates methylated and thus accessible sites. E, F Single-molecule maps over the pHV2 plasmid: 250-bp window map (E) and very high coverage (≥ 1200 single molecules) 200-bp window map (F)

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: Methylation, Plasmid Preparation

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 3 The ssDNA and active transcription landscape in the H. volcanii genome as measured by KASseq. A Global KAS-seq profiles over each of the five H. volcanii chromosomes in an exponential culture. B High reproducibility of active transcription measurements using KAS-seq. C–D Representative browser snapshots of KAS-seq profiles along the H. volcanii genome. E KAS-seq metaprofile along H. volcanii gene bodies

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques:

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 4 Abundant ssDNA structures associated with some H. volcanii CRISPR arrays in specific conditions. A Global KAS-seq profiles over each of the five H. volcanii chromosomes in a long-standing culture (∼3 months) reveal an extremely strong ssDNA peak associated with one of the CRISPR arrays on the pHV4 plasmid. B KAS-seq signal levels around pHV4 plasmid CRISPR arrays in different conditions. C KAS-seq and ATAC-seq levels around all three H. volcanii CRISPR arrays in different conditions

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: CRISPR, Plasmid Preparation

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 5 Coordination between chromatin accessibility and transcriptional activity within H. volcanii operons. The black bar shows the operon boundaries. A Ribosomal RNA operon. Note that the tracks shown here were generated by including multimapping reads (see the “Methods” section for details). B A-type ATP synthase subunits A, A, B, C, D, E, F, I, K, and H. C RNA polymerase II subunits. D NADH dehydrogenase-like complex subunits A, B, CD, H, I, J1, J2, K, L, M, and N

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: Activity Assay, Generated

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 6 Chromatin accessibility does not correlate with transcriptional activity in H. volcanii. A Differential chromatin accessibility between exponential and stationary conditions. B Differential KAS-seq levels between exponential and stationary conditions. C–E Lack of correlation between KAS and ATAC signals in exponential and stationary conditions. F Lack of correlation between changes in chromatin accessibility and changes in transcriptional activity

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: Activity Assay

Journal: Genome biology

Article Title: The chromatin landscape of the euryarchaeon Haloferax volcanii.

doi: 10.1186/s13059-023-03095-5

Figure Lengend Snippet: Fig. 7 Chromatin accessibility does not correlate with transcriptional activity in H. volcanii. A Genome-wide heatmaps of ATAC-seq and KAS-seq signals around H. volcanii TSSs, sorted by KAS-seq levels in the exponential condition. B, C Representative snapshots of genes with significantly altered transcriptional activity between the exponential and standing conditions, but no corresponding changes in chromatin accessibility

Article Snippet: Haloferax volcanii cell culture H. volcanii cells were obtained from the DSMZ German Collection of Microorganisms and Cell Cultures GmbH (Cat # 3757) and cultured in Halobacterium media [70, 71], prepared as follows: 7.50 g casamino acids, 10.00 g yeast extract, 3.00 g sodium citrate, 2.00 g KCl, 20.00 g MgSO4 × 7 H2O, 0.05 g FeSO4 × 7 H2O, 0.20 mg MnSO4 × H2O, and 250.00 g NaCl were mixed with distilled water in a total volume of 1 L. The media were then autoclaved and allowed to cool.

Techniques: Activity Assay, Genome Wide