| Accession | GMS-175-3
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| Title | Localization of genome-organizing proteins in the archaeon Thermococcus kodakarensis |
| Submit date | 2025-05-14 12:12:06 |
| Last update date | 2025-05-16 17:45:00 |
| Contact | Naomichi TAKEMATA takemata AT fc.ritsumei.ac.jp Department of Biotechnology, College of Life Sciences, Ritsumeikan University |
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| Total file size | 6.11 MB |
| Keywords | Nuclear-scale SMC Archaea Stiffness Asynchronous Cell culture ChIP-seq Genome DNA |
| Experiment type | ChIP-seq |
| Summary | In eukaryotes, SMC complexes form TADs by extruding DNA loops and being stalled by roadblock proteins. It remains unclear whether a similar mechanism of domain formation exists in prokaryotes. Using 3C-seq, we show that an archaeal homolog of the bacterial Smc-ScpAB complex organizes the genome of Thermococcus kodakarensis into TAD-like domains. We find that TrmBL2, a DNA-binding protein that forms a stiff nucleoprotein filament, stalls the T. kodakarensis SMC complex and establishes a boundary at the site-specific recombination site dif. TrmBL2 stalls the SMC complex at tens of additional non-boundary loci with lower efficiency. Intriguingly, the stalling efficiency is correlated with structural properties of underlying DNA sequences. Our study illuminates a eukaryotic-like mechanism of domain formation in archaea and a role of intrinsic DNA structure in large-scale genome organization. |
| Citation(s) | Yamaura, K., Takemata, N., Kariya, M. et al. Chromosomal domain formation by archaeal SMC, a roadblock protein, and DNA structure. Nat Commun 16, 1312 (2025). https://doi.org/10.1038/s41467-025-56197-y |
| Organism | OTHERs |
| Cell (Tissue) | not applicable |
| Protocol | Crosslinked cells were lysed by sonication, and the extract was clarified by centrifugation. Protein-DNA complexes were purified from the extract with antisera. After reversal of crosslinks, the DNA was extracted with phenol:chloroform:isoamyl alcohol and precipitated in ethanol. Libraries were constructed using NEBNext Ultra II DNA Library Prep Kit for Illumina with Sample Purification Beads. |
| Data processing | ChIP-seq data were mapped using Bowtie 2 version 2.3.5.1 (Langmead & Salzberg, 2012). Low-quality alignments (MAPQ < 30) were removed using SAMtools version 1.9 (Li et al., 2009). Generated BAM files were processed using the bamCoverage function (version 3.5.1) of deepTools (Ramirez et al., 2016) to calculate Reads Per Kilobase region per Million mapped reads (RPKM) for genomic bins. The IP/input ratio was calculated for each bin. |
| [1042] TrmBL2_fig8.png
TrmBL2-mediated regulation of Smc-ScpAB dynamics in T. kodakarensis image/png 657.07 KB MD5: d1f1dd62aa00d7b222c7e13dc7babc1c |
| [1003] T_kodakarensis_trmBL2_Smc_enrichment.txt
Smc enrichment in a trmBL2 deletion mutant of T. kodakarensis text/plain 1.81 MB MD5: 575d514976d476b7545b063066ad6fa5 |
| [1004] T_kodakarensis_WT_Smc_enrichment.txt
Smc enrichment in a wild-type strain of T. kodakarensis text/plain 1.82 MB MD5: e1a4e7fd5bfca51b5033ec7cde1cc92c |
| [1005] T_kodakarensis_WT_TrmBL2_enrichment.txt
TrmBL2 enrichment in a wild-type strain of T. kodakarensis text/plain 1.82 MB MD5: 868e6c7d1688f37c3a9ffcc893d79301 |
