ABSTRACT: The Cohesin complex has recently been described to regulate gene expression. We wanted to determine the gene expression profile specific in mouse ES cells after depletion of the Cohesin subunit Rad21. We used microarrays to detail the global programme of gene expression underlying depletion of Rad21 and identified distinct early development related genes up-regulated and many pluripotency related genes downregulated.
Project description:The Cohesin complex has recently been described to regulate gene expression. We wanted to determine the gene expression profile specific in mouse ES cells after depletion of the Cohesin subunit Rad21. We used microarrays to detail the global programme of gene expression underlying depletion of Rad21 and identified distinct early development related genes up-regulated and many pluripotency related genes downregulated. Rad21 was depleted in R1/E ES cells for 48h using esiRNAs against Rad21. An esiRNA against non-targeting Luciferase was used as a negative control
Project description:MTD project_description Inflammation and decreased stem cell function characterize organism aging, yet the relationship between these factors remains incompletely understood. This study shows that aged hematopoietic stem and progenitor cells exhibit increased ground-stage NF-κB activity, which enhances their responsiveness to undergo differentiation and loss of self-renewal in response to inflammation. The study identifies Rad21/cohesin as a critical mediator of NF-κB signals, by increasing chromatin accessibility of inter-/intra-genic and enhancer regions. Rad21/NF-κB are required for normal differentiation, but limit self-renewal of hematopoietic stem cells (HSCs) during aging and inflammation in an NF-κB dependent manner. HSCs from aged mice fail to downregulate Rad21/cohesin and inflammation/differentiation inducing signals in the resolution phase after acute inflammation. and The inhibition of cohesin/NF-κB is sufficient to revert the hypersensitivity of aged HSPCs to inflammation-induced differentiation. During aging, myeloid-biased HSCs with disrupted and naturally occurring reduced expression of Rad21/cohesin are increasingly selected over lymphoid-biased HSCs. Together, Rad21/cohesin mediated NF-κB signaling limits HSPC function during aging and selects for cohesin deficient HSCs with myeloid skewed differentiation.
Project description:Cohesin loss-of-function mutations are frequently observed in tumors, but the mechanism is unclear. Here, we found that depletion of RAD21, a core subunit of cohesin, leads to massive genome-wide DNA breaks, up to five-fold, and 147 translocation hotspot genes that are co-mutated with cohesin in multiple cancers. Increased DNA damages are independent of RAD21-loss-induced transcription alteration and loop anchor elimination. However, damage-induced chromosomal translocations coincide with the asymmetrically distributed Okazaki fragments of DNA replication, suggesting that RAD21 depletion causes replication stresses evidenced by the slower replication speed and increased stalled forks. Mechanistically, approximately 30% of the human genome exhibits an earlier replication timing after RAD21 depletion, caused by the early initiation of >900 extra dormant origins. Correspondingly, most translocation hotspot genes lie in timing-altered regions. Therefore, we conclude that cohesin dysfunction causes replication stresses induced by excessive DNA replication initiation, resulting in gross DNA damages that may promote tumorigenesis.
Project description:We recently identified recurrent mutations of cohesin complex in myeloid neoplasms through whole-exome sequencing analysis. RAD21 is one of the main components of the cohesin complex. In this study, to investigate the biological impact of wild-type RAD21 on Kasumi1 cells harboring RAD21 mutation, Kasumi1 cells were retrovirally transduced with either mock or wild-type RAD21, and expression array was performed. Expression analysis was performed for mock- or wild-type RAD21-transduced Kasumi-1 cells in triplicate. The experiment was performed twice independently.
Project description:We recently identified recurrent mutations of cohesin complex in myeloid neoplasms through whole-exome sequencing analysis. RAD21 is one of the main components of the cohesin complex. In this study, to investigate the biological impact of wild-type RAD21 on Kasumi1 cells harboring RAD21 mutation, Kasumi1 cells were retrovirally transduced with either mock or wild-type RAD21, and expression array was performed.
Project description:Rad21 is a subunit of cohesin. The main function of cohesin is to hold replicated chromosomes together until cells divide, but it also plays a role in gene expression. To find out which genes might be regulated by cohesin, a study was conducted to look for global changes in gene expression in zebrafish embryos lacking cohesin component Rad21. The zebrafish Rad21 mutant used for expression analysis was rad21nz171, an allele isolated in a forward genetic screen for regulators of runx1. Experiment Overall Design: RNA from rad21nz171 mutant and wild type zebrafish embryos collected at 24 hours post-fertilization (h.p.f.) and 48 h.p.f. was hybridized to Affymetrix microarrays (Gene Chip zebrafish genome arrays cat. no. 900488). Four pools of 50 embryos for each genotype and time point were used as the RNA source, and RNA from each pool was hybridized independently such that the experiment had four biological replicates.
Project description:Rad21 is a subunit of cohesin. The main function of cohesin is to hold replicated chromosomes together until cells divide, but it also plays a role in gene expression. To find out which genes might be regulated by cohesin, a study was conducted to look for global changes in gene expression in zebrafish embryos lacking cohesin component Rad21. The zebrafish Rad21 mutant used for expression analysis was rad21nz171, an allele isolated in a forward genetic screen for regulators of runx1.
Project description:In eukaryotes, the nuclear chromatin in the interphase is a hierarchical organization. Previous studies have suggested that cohesin-DNA interactions play important architectural functions for chromatin structure in the interphase. In this study, we applied super-resolution imaging, single-molecule imaging to measure the distribution of cohesin subunits. Hi-C and RNA-seq were used to reveal the 3D genome structures alterations and the relationship with breast cancer upon Rad21 up-regulation. Hi-C experiment showed that the contact frequency of short distance was increased while that of long distance was decreased in the absence of Rad21 up-regulation. Chromatin interactions between A and B compartments increased and compartmentalization strength was reduced significantly upon Rad21 up-regulation. Correspondingly, accumulated contacts were shown at TAD corners and inter-TAD interactions increased. These observations support cohesin loop extrusion model and lead us to propose the unique role of Rad21-loader interaction in cohesin loading and further cohesin extrusion. Moreover, we combined transcriptome changes and chromatin structure alterations upon Rad21 up-regulation and revealed the correlation between Rad21 and breast cancer by affecting chromatin architecture.