PML-RARA can increase hematopoietic self-renewal without causing a myeloproliferative disease in mice

Abstract

Acute promyelocytic leukemia (APL) is characterized by the t(15;17) translocation that generates the fusion protein promyelocytic leukemia-retinoic acid receptor α (PML-RARA) in nearly all cases. Multiple prior mouse models of APL constitutively express PML-RARA from a variety of non-Pml loci. Typically, all animals develop a myeloproliferative disease, followed by leukemia in a subset of animals after a long latent period. In contrast, human APL is not associated with an antecedent stage of myeloproliferation. To address this discrepancy, we have generated a system whereby PML-RARA expression is somatically acquired from the mouse Pml locus in the context of Pml haploinsufficiency. We found that physiologic PML-RARA expression was sufficient to direct a hematopoietic progenitor self-renewal program in vitro and in vivo. However, this expansion was not associated with evidence of myeloproliferation, more accurately reflecting the clinical presentation of human APL. Thus, at physiologic doses, PML-RARA primarily acts to increase hematopoietic progenitor self-renewal, expanding a population of cells that are susceptible to acquiring secondary mutations that cause progression to leukemia. This mouse model provides a platform for more accurately dissecting the early events in APL pathogenesis. IntroductionMore than 95% of acute promyelocytic leukemia (APL; FAB-M3) cases are associated with the recurrent t(15;17)(q22:q11) translocation, which results in the expression of a fusion transcript promyelocytic leukemia-retinoic acid receptor α (PML-RARA), inconsistent expression of a reciprocal fusion product (RARA-PML), and haploinsufficiency for both PML and RARA (1, 2). Although APL has served as an important model disease for understanding leukemogenic pathways directed by a specific translocation, the precise mechanisms by which t(15;17) initiates leukemia remain unknown.Several mouse models have been used to study the mechanisms by which PML-RARA causes transformation (reviewed in refs. 3, 4). These models have used viral transduction, transgenic, and knockin strategies; regulatory sequences from the MRP8 gene (also known as S100 calcium binding protein A8 [S100A8]) and human and murine cathepsin G (CTSG and Ctsg, respectively) genes have been used to direct the expression of PML-RARA to the myeloid compartment (5-8), where can cause a myeloproliferative disease and APL after a long latent period.However, there are many issues with existing mouse models of APL that must be addressed to fully understand how PML-RARA works. (a) The dose of PML-RARA is known to be important for AML penetrance, but no model to our knowledge has yet expressed PML-RARA from the endogenous Pml locus. This has not previously been attempted, since Pml is ubiquitously expressed, and since widespread expression of PML-RARA during development causes embryonic lethality (9, 10). (b) t(15;17) is a somatically acquired event, but all prior models have used constitutive PML-RARA expression, raising the possibility that hematopoietic cells may have a...

Transcription and methylation analysis of preleukemic promyelocytes indicate a dual role for PML/RARA in leukemia initiation

“…Our data extend previous studies linking PML/RARA to altered proliferation, [42][43][44] providing the novel finding that altered cell-cycle kinetics is a feature of PML/RARAexpressing cells at very early stages of the leukemogenic process. We found that the enhanced capacity for promyelocytes to proliferate depends on the PML part of the fusion and correlates with the ability to initiate leukemia, in line with previous findings.…”

“…Finally, since we observed putative PML/RARA binding sites in our most down-regulated gene Ltf, we investigated binding of the fusion protein using chromatin immunoprecipitation. We confirmed binding and enrichment of PML/RARA at the Ltf locus (Online Supplementary Figure S7C), providing a mechanistic explanation for the strong deregulation observed.Our data extend previous studies linking PML/RARA to altered proliferation, [42][43][44] providing the novel finding that altered cell-cycle kinetics is a feature of PML/RARAexpressing cells at very early stages of the leukemogenic process. We found that the enhanced capacity for promyelocytes to proliferate depends on the PML part of the fusion and correlates with the ability to initiate leukemia, in line with previous findings.…”

PML-RARA requires DNA methyltransferase 3A to initiate acute promyelocytic leukemia

“…In contrast, PR +/--derived BM cells have a competitive transplantation advantage, as demonstrated by an increased ability to contribute to peripheral blood lineages (especially myeloid) over time (39). To determine whether Dnmt3a is required for this competitive advantage, we mixed whole BM cells from mice with the indicated genotypes ( Figure 7A) at a ratio equal to that of WT competitor BM cells and transplanted the cells into lethally irradiated WT mice.…”

Dynamic HoxB4-regulatory network during embryonic stem cell differentiation to hematopoietic cells

“…19 In total, we identified 31 TF genes that were both targeted differentially by HoxB4 and exhibited differential expression between adjacent time points ( Figure 4A) 32,33 Stat5, 34 Tcf7, 35 and Pml-RARA. 36 Direct regulation of multiple hematopoiesis TFs by HoxB4 strongly implies that it is a master regulator positioned high in the hierarchy of the hematopoiesisregulatory network. Most of the TFs targeted by HoxB4 were either up-regulated or down-regulated; however, several TFs showed more complicated HoxB4 regulation ( Figure 4A).…”