SRA

Phase separation serves an important role in the three-dimensional chromosome organization and remodeling in eukaryotes. Whether this process is involved in archaeal chromosome organization is unknown. Here we demonstrate that archaeal DNA condensing protein1 (aDCP1) from the hyperthermophilic crenarchaeon Sulfolobus islandicus is able to bridge DNA efficiently and form large protein-DNA condensates with a droplet- or gel-like morphology in vitro. Within the condensates, aDCP1 exhibits a fast dynamic while the DNA appears in a solid-like state. At the single-molecule level, aDCP1 efficiently compacts DNA through a three-step mechanism, which presumably entails the clustering of aDCP1 on the DNA and the subsequent fusion of the clusters. Deletion of the aDCP1 gene results in noticeable changes in chromosome conformation in S. islandicus, which are characterized by enhanced interactions between the A and B compartments and reduced interactions within the self-interacting domains as well as between domains in the same compartment. Taken together, our results indicate that aDCP1 is capable of inducing DNA bridging-induced phase separation and serves a role in chromosome organization in the organism. Overall design: Chromatin immunoprecipitation (ChIP-Seq) was performed according to Takemata et al.(39) with slight modifications. Briefly, the cells were collected , cross-linked by adding 1% formaldehyde for 15 minutes, and quenched with a final concentration of 125 mM glycine. The cells were pelleted by centrifugation at 5,000 g for 10 minutes and washed with PBS. The cells were then resuspended in TBS-TT buffer (20 mM Tris, 150 mM NaCl, 0.1% Tween-20, 0.1% Triton X-100, pH 7.5) and fragmented by sonication until the DNA fragments were of 200-500 bp. After centrifugation (10,000 g for 15 minutes), a 100 µl aliquot of the DNA-containing supernatant was kept apart to be uses as an input control and the remaining sample was divided into two aliquots. One aliquot was incubated with anti-aCcr1 antibody-coated protein A beads (Cytiva) and the other was incubated with pre-immune serum-coated protein A beads, which served as a nonspecific binding control (Mock control). After incubation at 60 °C for 10 hrs, the samples were collected and the captured DNA was purified by using the DNA Cycle-Pure Kit (Omega) according to the manufacturer's instruction. The input samples were treated as above without the addition of antiserum and beads. The purified DNA was used for ChIP-Seq library preparation. The library was constructed by Novogene Corporation (Beijing, China). Subsequently, pair-end sequencing of sample was performed on Illumina platform (Illumina, CA, USA). Library quality was assessed on the Agilent Bioanalyzer 2100 system