Enhanced FLI1 accessibility mediates STAG2-mutant leukemogenesis
Bibliographic record
Abstract
Abstract Hematopoietic stem cells (HSCs) are multipotent cells that can give rise to all blood cell types. In leukemia, HSCs are often the cell of origin that carry the founding mutation during clonal evolution. The molecular mechanisms that regulate HSC lineage specification are of great interest for understanding hematopoiesis and its dysregulation in diseases. One of the key factors that influence HSC differentiation is the chromatin structure and accessibility of the genome, which modulates the expression of lineage-specific genes and transcription factors (TFs). STAG2 is a member of cohesin complex that is recurrently mutated in >10 cancers and is essential in maintaining the integrity of 3D genome partitioning structures known as topologically associated domains (TADs). Our previous work has demonstrated that depletion of various cohesin factors, including Stag2, leads to increased hematopoietic stem and progenitor population self-renewal and myeloid-biased differentiation. Loss of Stag2 leads to impaired sub-TADs and affects key hematopoietic TFs, such as PU.1, to access and engage their target genes. In addition to defective lineage priming, Stag2 loss also associates with promoting leukemogenesis and establishes clonal dominance in MDS, prior to AML transformation. However, the temporal and spatial patterns of chromatin changes during stem cell to myeloid commitment and the role of STAG2-mutation in leukemogenesis are not fully elucidated. To describe the 3D chromatin structure stem to myeloid lineage transition, we performed low-input Hi-C on LSK and GMP population. During normal myeloid cell fate specification, we observed an overall shift towards a more repressive and restricted chromatin state with loss of DNA loops associated with stem cell specific genes, such as Mecom and Hoxa9. When Stag2 is lost, we observed loss of proximity at the promoter and upstream enhancer associated with Mecom in the LSK KO cells, which suggests Stag2 is important for stem cell related enhancer-promoter interactions. Next, we assessed chromatin accessibility change between WT LSK vs WT GMP and Stag2 mutant LSK vs WT LSK. We found that Fli1 is the top motif among the accessible region of WT LSK cells comparing to WT GMPs. Furthermore, the accessible regions of Fli1 were increased in Stag2 mutant LSK cells. To determine the impact of STAG2 mutation in leukemogenesis, we generated STAG2 KO (refer to as KO) OCI-AML3 cells. Comparing to isogeneic non-targeting (refer to as NT) control cells, KO cells have an increased cell surface expression of stem cell associated markers, such as CD34 and ESAM. To obtain chromatin signature, we performed ATACseq and found an increased chromatin accessibility in the KO compared to NT, especially at the HOXA locus. Among the more accessible regions, motif analysis identified FLI1 as the predicted top transcription factor (TF) to bind. We then performed FLI1 Cut&Run and showed FLI1 binds to cis-regulatory elements and more accessible region of the KO cells. Functionally, FLI1 knock down led to decreased CD34 cell surface markers in the KO cells, which confirms that FLI1 mediates phenotype changes. We next generated dual Stag2ΔNpm1c/+ murine models with tamoxifen inducible UbcCreERT2 to establish the functional effects on hematopoiesis. After 4 weeks, LSK cells (Lin-Sca1+Kit+) are increased in Stag2ΔNpm1c/+double mutant mice. Within LSK cells, Stag2ΔNpm1c/+ has marked expansion of the myeloid biased MPP3 (LSK+Flk2-Cd150-Cd48+) cells. Molecularly, we found Stag2ΔNpm1c/+ MPP3 cells have increased accessibility compared to Npm1c/+ cells. Motif analysis identified Fli1 as the top TF in Stag2ΔNpm1c/+, which confirms the observation in STAG2 KO OCI-AML3 cell lines. When aged, mice carrying Stag2ΔNpm1c/+ developed highly penetrant acute leukemia with hypolobated megakaryocytes and dyserythropoiesis in the bone marrow, which suggests myelodysplastic related changes. Overall, we examined the 3D genome changes during myeloid fate specification and established both in vitro and in vivo system of cohesin mutant leukemia model. We determined that the abnormal chromatin remodeling driven by STAG2 mutation occurs in HSPCs and hyperactivity of FLI1 is the key dysfunction in Stag2/Npm1c co-mutant. Further studies include determining the phenotypic and chromatin response of FLI1 inhibition in STAG2 mutant models, which will pave the way to highlight new therapeutic potentials of cohesin mutant leukemia.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.004 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".