Stepwise GATA1 and SMC3 mutations alter megakaryocyte differentiation in a Down syndrome leukemia model

Abstract

Acute megakaryoblastic leukemia of Down syndrome (DS-AMKL) is a model of clonal evolution from a preleukemic transient myeloproliferative disorder requiring both a trisomy 21 (T21) and a GATA1 s mutation to a leukemia driven by additional driver mutations. We modeled the megakaryocyte differentiation defect through stepwise gene editing of GATA1s , SMC3 +/– , and MPL W515K , providing 20 different T21 or disomy 21 (D21) induced pluripotent stem cell (iPSC) clones. GATA1s profoundly reshaped iPSC-derived hematopoietic architecture with gradual myeloid-to-megakaryocyte shift and megakaryocyte differentiation alteration upon addition of SMC3 and MPL mutations. Transcriptional, chromatin accessibility, and GATA1-binding data showed alteration of essential megakaryocyte differentiation genes, including NFE2 downregulation that was associated with loss of GATA1s binding and functionally involved in megakaryocyte differentiation blockage. T21 enhanced the proliferative phenotype, reproducing the cellular and molecular abnormalities of DS-AMKL. Our study provides an array of human cell–based models revealing individual contributions of different mutations to DS-AMKL differentiation blockage, a major determinant of leukemic progression.

RUNX1 isoform disequilibrium promotes the development of trisomy 21–associated myeloid leukemia

“…37 In contrast, the 12 RUNX1A-specific cofactors were involved in active transcription/replication and G1-S transition, including the megakaryocytic transcription factor NFE2, which is mutated in a subset of patients with myeloid neoplasms and is functionally involved in the megakaryocyte differentiation blockage of GATA1s pluripotent stem cells (Figure 4B). [38][39][40] Importantly, MAX, a crucial cofactor of MYC, was among the RUNX1Aspecific interacting proteins, as verified via western blot (Figure 4C). GATA1s coimmunoprecipitated with RUNX1A and RUNX1C, albeit not at a significant enrichment level (data not shown).…”

Advances in molecular characterization of myeloid proliferations associated with Down syndrome

“…GATA1s mutation alone disrupts differentiation of megakaryocytes and promotes expansion of myeloid and megakaryocytic progenitors, while production of aberrant megakaryoblasts is strengthened on the background of trisomy 21 ( Banno et al, 2016 ; Juban et al, 2021 ; Matsuo et al, 2021 ). TAM requires the synergy between trisomy 21 and GATA1s but leukemic transformation may be independent of trisomy 21 ( Wagenblast et al, 2021 ; Arkoun et al, 2022 ). In contrast, synergy between GATA1s and subsequent “tertiary” molecular alterations is critical for progression of TAM to ML-DS.…”

“…GATA1s and STAG2 knockout cooperatively increased the megakaryocytic population and induced the ML-DS immunophenotype ( Barwe et al, 2022 ). In another study, trisomic 21 iPSC line ( Chou et al, 2012 ) was edited to introduce GATA1s followed by heterozygous inactivation of SMC3 ( SMC3 +/− ) and then, introduction of a gain-of-function MPL mutation (MPLW515K) ( Arkoun et al, 2022 ). It was found that GATA1s impaired megakaryocyte differentiation and that SMC3 +/− enhanced this effect independent of trisomy 21.…”

“…MPLW515K further increased the megakaryocyte output in this model, including through the induction of growth factor independence. Low expression of NFE2 was critical for the induction of megakaryocyte dysplasia by GATA1s ( Arkoun et al, 2022 ). These novel iPSC-based models are likely to rapidly advance our understanding of ML-DS pathogenesis, and assist therapy development.…”

Dissecting stepwise mutational impairment of megakaryopoiesis in a model of Down syndrome–associated leukemia

“…To help understand the evolution of TMD to DS-AMKL, it is necessary to develop human-relevant models that can recapitulate the DS-AMKL mutations and allow examination of progressive perturbations of megakaryocytic differentiation and other disease phenotypes. In this issue of the JCI , Arkoun and colleagues accomplish this objective using a stepwise technique to introduce GATA1 , MPL , and SMC3 mutants into induced pluripotent stem cells (iPSCs) from humans with or without DS ( 9 ). The researchers uncovered the individual contributions of each variant and how they could cooperate with T21 to lead to DS-AMKL.…”