The DNA-binding protein HTa from Thermoplasma acidophilum is an archaeal histone analog
- MRC London Institute of Medical Sciences (LMS), United Kingdom
- Institute of Clinical Sciences (ICS), Faculty of Medicine, Imperial College, United Kingdom
Figures
Figure 1
Predicted structure and measured abundance of HTa.
(a) Predicted secondary structures of HTa (T. acidophilum), the bacterial HU protein HupA (E. coli), and the archaeal histone protein HmfA (M. fervidus). (b) Predicted quaternary structure of the …
Figure 2 with 1 supplement
Phylogenetic relationships of HU family proteins from bacteria, eukaryotes, and archaea.
(a) Protein-level phylogenetic tree of HU proteins including HTa (see Materials and methods for details on phylogenetic reconstruction). The tree is midpoint-rooted. Reported domain-level membership …
Figure 2—figure supplement 1
Phylogenetic placement of HU proteins attributed to halophilic archaea.
The phylogenetic tree shown is an excerpt of the protein-level HU family tree shown in Figure 2, focussing on sequences from halophilic archaea (orange), which cluster mainly with sequences of …
Figure 3 with 4 supplements
HTa-mediated primary chromatin architecture in T. acidophilum mapped by MNase-Seq.
(a) Growth curve of T. acidophilum as determined using optical density (OD600). Time points used for downstream experiments are indicated (means and ± SEM across four biological replicates). (b) …
Figure 3—figure supplement 1
Agarose gel (3%) of MNase digestion products from T. acidophilum (day 2) along with digestion products of E. coli ectopically expressing either HTa, HupA, YFP, HupA (E38K,V42L), HU from T. composti or HU from L. floricola, from the same plasmid backbone.
HupA (E38K,V42L) is a mutant that had previously been shown to induce extreme compaction of the E. coli nucleoid (Kar et al., 2005).
Figure 3—figure supplement 2
Distribution of the lengths of fragments mapped to the T. acidophilum genome for all replicates across the growth cycle.
Figure 3—figure supplement 3
Heat maps indicating MNase-seq coverage by fragment length relative to the center of broad peaks in T. acidophilum, for the same sample (day1, replicate 3), digested for either 15 or 30 min.
Figure 3—figure supplement 4
Multiscale analysis of MNase signal.
(a) Chromosome-wide MNase-Seq coverage along the T. acidophilum chromosome (day2, replicate 2), normalized using sonicated DNA to remove replication-associated coverage bias. (b) Multiscale analysis …
Figure 4 with 3 supplements
Asymmetric coverage signals around peaks in T. acidophilum and M. fervidus that track underlying nucleotide content.
(a) Empirical example and (b) schematic describing our approach to re-orienting coverage signals at broad peaks based on the coverage of small fragments around the dyad axis. (c, d) Heat maps …
Figure 4—figure supplement 1
Weblogos of bitscores and nucleotide occurrence probabilities at (a) narrow and (b) broad peaks detected during exponential phase in T. acidophilum.
Information content is so low that the bitscore plots appear empty when using the common 0–2 bit visualization range. Logos are only visible when zooming in on the 0–0.02 range.
Figure 4—figure supplement 2
Normalized MNase-Seq coverage relative to the center of narrow peaks oriented according to the abundance of (a) 87–97 bp fragments in M. fervidus and (b) 70–100 bp fragments in T. acidophilum.
Middle and right panels are focused on peaks where 87–97 bp (70–100 bp) fragments are common or rare, respectively. Lower panels display the proportion of SS (=CC|CG|GC|GG) and WW (=AA|AT|TA|TT) …
Figure 4—figure supplement 3
As in Figure 4—figure supplement 2 but for 87–97 bp peaks scored according to 117–127 bp fragments and oriented according to 60–70 bp fragments.
Note the increase in WW content flanking the smaller-sized peaks that do not get extended further.
Figure 5 with 2 supplements
Comparison and predictive power of nucleotide enrichment patterns associated with HTa and archaeal histones.
(a) Proportion of SS (=CC|CG|GC|GG) dinucleotides, (b) A|T mononucleotides, and (c) RR (=purine/purine)|YY (=pyrimidine/pyrimidine) dinucleotides relative to the centers of reads of defined length …
Figure 5—figure supplement 1
Proportion of SS (=CC|CG|GC|GG) dinucleotides relative to the centers of reads of defined length (41–53 bp) in T. acidophilum.
Figure 5—figure supplement 2
Predicting in vivo HTa occupancy.
(a) In vivo occupancy in T. acidophilum is poorly predicted by a Lasso model trained on a T. acidophilum naked DNA digest (rho = 0.07, p<2.2×10−16). (b) In contrast, in vivo occupancy in T. …
Figure 6 with 4 supplements
In vitro experiments to assess HTa binding preferences.
(a) Occupancy of small fragments across the T. acidophilum genome in vivo (day 2) correlates with occupancy following in vitro reconstitution and with (b) occupancy predicted by a Lasso model …
Figure 6—figure supplement 1
In vitro reconstitution of HTa:DNA nucleoprotein complexes.
(a) 16% TTS protein gel (Biorad) showing different concentrations of BSA (Biorad) and purified untagged HTa. (b) Bioanalyzer trace of in vitro chromatin reconstitution. Two replicates are …
Figure 6—figure supplement 2
EMSA backbone sequences.
(a) In vivo occupancy (day 2) at five 100 bp regions detailed in (b) is correlated with in vitro occupancy. Randomized dinucleotides are highlighted in green. (c) The proportion Pslow of diversified …
Figure 6—figure supplement 3
The relationship between GC content of an oligo and Pslow.
Only oligos represented by at least 200 sequenced reads are considered. This analysis shows that results in Figure 6e are not driven by few highly abundant oligos but represent the cumulative effect …
Figure 6—figure supplement 4
The relationship between GpC dinucleotide content of an oligo and Pslow.
Only oligos represented by at least 200 sequenced reads are considered. This analysis shows that results in Figure 6f are not driven by few highly abundant oligos but represent the cumulative effect …
Figure 7 with 1 supplement
Broad peaks are associated with heterogeneous GC content in exponential but not stationary phase.
(a) Average GC content at broad peaks (day 2), separated into deciles based on the relative abundance of small fragments and (b) the corresponding relative coverage for large and small fragments …
Figure 7—figure supplement 1
Small fragment abundance at narrow peaks.
(a) Average GC content at narrow peaks (day 2), separated into deciles based on the relative abundance of small fragments. (b) corresponding relative coverage for large and small fragments during …
Figure 8 with 1 supplement
MNase-Seq coverage around transcriptional start sites in T. acidophilum and histone-encoding archaea in the context of dynamic transcription.
(a) Broad peaks associated with low abundance of small fragments are enriched in intergenic regions. (b) Left and central panel: Heat maps indicating MNase-seq coverage by fragment length relative …
Figure 8—figure supplement 1
HTa and histone occupancy around transcription end sites.
(a) Median normalized MNase-seq coverage across fragment sizes relative to the distance from TESs or stop codons in different species. To ensure that the stop codons constitute a reasonable proxy …
Author response image 1
Nucleotide periodicities in the T. acidophilum genome.
Additional files
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Supplementary file 1
Representation of HU homologs across bacterial phyla.
- https://cdn.elifesciences.org/articles/52542/elife-52542-supp1-v2.docx
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Supplementary file 2
Examples of putative archaeal and eukaryotic homologs that likely represent contamination during genome assembly.
- https://cdn.elifesciences.org/articles/52542/elife-52542-supp2-v2.docx
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Supplementary file 3
Fourier filtering parameters.
- https://cdn.elifesciences.org/articles/52542/elife-52542-supp3-v2.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/52542/elife-52542-transrepform-v2.docx
