The cohesin subunit Rad21 is a negative regulator of hematopoietic self-renewal through epigenetic repression of Hoxa7 and Hoxa9

Abstract

Acute myelogenous leukemia (AML) is a high-risk hematopoietic malignancy caused by a variety of mutations, including genes encoding the cohesin complex. Recent studies have demonstrated that reduction in cohesin complex levels leads to enhanced self-renewal in hematopoietic stem and progenitors (HSPCs). We sought to delineate the molecular mechanisms by which cohesin mutations promote enhanced HSPC self-renewal since this represents a critical initial step during leukemic transformation. We verified that RNAi against the cohesin subunit Rad21 causes enhanced self-renewal of HSPCs in vitro through derepression of Polycomb Repressive Complex 2 (PRC2) target genes, including Hoxa7 and Hoxa9. Importantly, knockdown of either Hoxa7 or Hoxa9 suppressed self-renewal, implying both are critical downstream effectors of reduced cohesin levels. We further demonstrate that the cohesin and PRC2 complexes interact and are bound in close proximity to Hoxa7 and Hoxa9. Rad21 depletion resulted in decreased levels of H3K27me3 at the Hoxa7 and Hoxa9 promoters, consistent with Rad21 being critical to proper gene silencing by recruiting the PRC2 complex. Our data demonstrates that the cohesin complex regulates PRC2 targeting to silence Hoxa7 and Hoxa9 and negatively regulate self-renewal. Our studies identify a novel epigenetic mechanism underlying leukemogenesis in AML patients with cohesin mutations.

Cohesin <b><i>RAD21</i></b> Gene Promoter Methylation in Patients with Chronic Lymphocytic Leukemia

“…Further stratification of patients according to the type of the detected chromo- 130 some abnormalities revealed that 0% of patients with numerical aberrations, 27.78% of patients with structural changes, and 25.00% of patients with both numerical and structural aberrations (10/40) had a RAD21 promoter methylation ( Table 2 ). The fact that none of the patients carrying only numerical and not structural aberrations was methylated is consistent with a recent study which reported that RAD21 depletion does not induce aneuploidy but increases Hoxa9 expression and enhances self-renewal of primary HSPCs [Fisher et al, 2017]. Our results are reinforced by 2 other recent studies showing that downregulation of elevated cohesin leads to copy number-associated gene expression changes without disturbing chromosomal segregation and that reduced cohesin destabilizes high-level gene amplification by disrupting pre-replication complex bindings in human cancers with chromosomal instability [Yun et al, 2016].…”

“…The presence of RAD21 methylation only in CLL patients and not in healthy donors is an interesting observation indicating a possible implication of RAD21 inactivation in CLL pathogenesis. According to a recent study, RAD21 depletion enhances self-renewal of HSPCs [Fisher et al, 2017]. Therefore, based on our results combined with the above mentioned roles of RAD21, we suggest that its inactivation via methylation may disturb DNA repair mechanisms and enhances self-renewal of CLL cells contributing to the development of the disease.…”

“…More specifically, whole-exome sequencing showed that its ablation in human cell lines produces chromosomal rearrangements such as translocations, duplications, or deletions and leads to the formation of chromosome fusions which synergistically with replication stress generate distant single-ended DNA double-strand breaks that can produce potentially deleterious rearrangements [Gelot et al, 2016]. It is worth mentioning that most recently it has been demonstrated that RAD21 acts as a negative regulator of hematopoietic selfrenewal through epigenetic repression of HoxA7 and HoxA9 genes, indicating its possible implication in leukemogenesis [Fisher et al, 2017]. However, the methylation status of RAD21 has never been studied in hematological malignancies including CLL.…”

Cohesin Mutations in Myeloid Malignancies

“…In addition, a unique sub-type of AML, acute megakaryoblastic leukemia associated with Down Syndrome (DS-AMKL) exhibits cohesin mutations in almost 50% of patients [15]. A number of groups have investigated mechanisms by which cohesin mutations may underlie AML (Figure 1, key figure), which will be the focus of this review [16–20]. …”

“…Our lab and others have investigated this question using in vivo mouse models and cultured cells[16–20]. The common finding is that cohesin mutations are capable of conferring enhanced self-renewal on hematopoietic stem and progenitor cells (HSPCs).…”