IRCM Researchers

BTB/POZ domain protein and c-Myc co-factor Miz-1 is a new target for cancer treatment
Deletion of the BTB/POZ domain of Miz-1 (Myc interacting zinc finger protein 1), an important c-Myc co-factor, significantly delays T- and B- ALL/lymphoma in mice and interferes with the oncogenic activity of c-Myc and the activation of p53. The genetic ablation of the Miz-1 POZ domain also enhances the anti-leukemic effect of cytarabine in mice, suggesting that targeting Miz-1 could render current chemotherapies more effective with a better outcome for patients. These discoveries were the basis for two publications (PNAS, 2014; Cancer Research, 2019) and for three reports of invention (IRCM PTM 1901, -1902, -1903). Impact: These findings created a unique opportunity to set up the “Miz-1 POZ domain drug discovery platform” and to explore Miz-1 as a therapeutic target in cancers that depend on c-Myc.

GFI1 is a biomarker candidate for myeloid leukemia
We discovered a variant allele of the human gene for the zinc finger transcription factor GFI1 (Growth factor independence 1). This GFI1 variant has reduced function and is associated with AML (acute lymphoblastic leukemia). The progression from Myelodysplastic Syndrome (MDS) to AML, the development of de novo AML and myelo-proliferative diseases were accelerated in mice expressing this variant or reduced GFI1 levels. The GFI1 variant or GFI1 knockdown also altered the response of leukemic cells to drugs affecting histone acetylation (Exp. Hematology, 2016, Leukemia, 2016 and Leukemia, 2019). Impact: The GFI1 variant and GFI1 expression levels are prognostic factors and can predict the response to epigenetic therapies.

Methylation of DNA repair factor MRE11 is facilitated by GFI1
We identified the DNA repair proteins MRE11 (Meiotic Recombination 11 homolog 1), 53BP1 (p53 binding protein 1), and the methyltransferase PRMT1 as new GFI1 binding partners. GFI1 recruits PRMT1 to MRE11 or 53BP1 and enables their post-translational modification via methylation. This indicated that GFI1 also has non-transcriptional roles in which it regulates proteins involved in DNA repair. GFI1 deficiency delays DNA repair in lymphocytes and renders them highly sensitive to DNA damage (Nature Communications, 2018). Impact: The cellular response against DNA damaging drugs or irradiation depends on the activity and expression level of GFI1 and overexpressing of GFI1 may be linked to therapy resistance.

GFI1 regulates a p53 dependent DNA damage response pathway
Genetic ablation of the transcription factor GFI1 delays lymphomagenesis by the activation of a p53 DNA damage response pathway, indicating that GFI1 is an “onco- requisite” factor. A reciprocal regulation between GFI1 and p53 exists that controls the induction of apoptosis in T cells. Here, GFI1 prevents induction of p53 dependent apoptosis by recruiting the Lysine demethylase KDM1A (LSD1) to p53, which leads to the demethylation of its C-terminal domain and changes its regulatory activity (Cancer Cell, 2013, Scientific Reports, 2019). Impact: These observations support that low GFI1 expression could render patients with lymphoid leukemia more susceptible to therapeutics that work through the DNA damage response.

GFI1B controls hematopoietic stem cells numbers and platelet production
Mice deficient for GFI1B (Growth factor independence 1B, a GFI1 paralog) expand hematopoietic stem cells (HSCs) and megakaryocyte (MK) precursors, but lack platelets. GFI1B controls integrin-dependent cytoskeleton organization, spreading and migration of MKs and α- tubulin dependent formation of pro-platelets (Haematologica, 2017). Using BioiD, we discovered that GFI1B forms complexes with β-catenin, some of its co-factors and LSD1. ChIP-Seq analyses showed that a tripartite β- catenin/GFI1B/LSD1 complex exists and occupies and regulates Wnt/β-catenin target genes. This work suggested that GFI1B controls both the cellularity and functional integrity of both HSCs and MKs precursors by regulating the Wnt/β-catenin signaling pathway (Nature Communications, 2019). Impact: 1.) On the understanding how HSC cell numbers are controlled, which is critical, since the use of human cord blood stem cells in stem cell transplantation therapy depends on the ability to expand HSCs in vitro. 2.) Clear evidence that platelet production requires GFI1 providing a mechanistic link to the inherited bleeding disorder “GFI1B-related thrombocytopenia”.

Mouse model for GFI1B-related thrombocytopenia (GFI1B-RT)
GFI1B-related thrombocytopenia (GFI1B-RT) is a rare inherited bleeding disorder caused by the presence of allelic variants of the GFI1B gene that produce truncated GFI1B proteins with dominant-negative (DN) properties. Affected family members show low platelet counts, an expansion of megakaryocytes and mild erythroid defects. We have used CRISPR-Cas9 to generate mice that carry such dominant negative (DN) Gfi1b alleles and showed that these animals have reduced platelet counts and megakaryocyte expansion similar as GFI1B-RT patients. Gfi1b-DN mice also had a delayed recovery from platelet depletion, indicating a defect in stress thrombopoiesis. Injecting Gfi1b-DN mice with romiplostim, a thrombopoietin receptor super agonist used in the clinic, increased platelet numbers even beyond normal levels. Impact: Our data prove a causal link between DN mutations in GFI1B and inherited thrombocytopenia and suggest that patients with GFI1B-RT could be treated successfully with thrombopoietin agonists (Haematologica, 2020).

Role of GFI1 in inflammation
We have used knockin mice harboring a P2A mutation in GFI1 coding region that renders it unable to bind LSD1 and associated histone modifying enzymes such as HDACs to investigate the consequences of the disruption of these complex. We observed that GFI1P2A mice die prematurely and show increased numbers of memory effector and regulatory T cells in the spleen accompanied by a severe systemic inflammation with high serum levels of IL-6, TNFa and IL-1b and overexpression of the gene encoding the cytokine Oncostatin M (OSM). We identified lung alveolar macrophages, CD8 T cell from the spleen and thymic eosinophils and monocytes as the sources of these cytokines in GFI1P2A mice. Impact: We demonstrate that histone modification by LSD1 that are facilitated by GFI1 are critical to control inflammatory pathways. Since LSD1 inhibitors are used in experimental anti-cancer therapies in clinical trials, our findings point to possible complications with these drugs (J. Immunol., 2021, in press).    

GFI1 as a component of the nucleosome remodeling and histone deacetylase (NuRD) complex
Using BioId and mass spectrometry techniques we showed that GFI1 interacts with the chromodomain helicase CHD4 and other components of the “Nucleosome remodeling and deacetylase” (NuRD) complex. In granulo-monocytic precursors, GFI1, CHD4 or GFI1/CHD4 complexes occupy sites of open chromatin enriched for histone marks associated with active transcription suggesting that GFI1 recruits the NuRD complex to target genes that are regulated by active or bivalent promoters and active enhancers. We demonstrate that GFI1/CHD4 complexes regulate chromatin openness and histone modifications differentially to enable regulation of target genes affecting the signaling pathways of the immune response or nucleosome organization or cellular metabolic processes. Impact: Our data suggest that GFI1 does not exclusively occupies sites of closed chromatin but may also take part in the regulation of poised enhancers and active promoters during the differentiation of early hematopoietic precursors (BioRXiv, 2021).