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ZBTB7A links tumor metabolism to myeloid differentiation

2020; Elsevier BV; Volume: 87; Linguagem: Inglês

10.1016/j.exphem.2020.05.010

1873-2399

Enric Redondo Monte, Paul Kerbs, Philipp A. Greif,

Cancer, Hypoxia, and Metabolism

•Metabolism and cell differentiation are two interconnected processes that influence each other.•ZBTB7A may act as a link between linage commitment and metabolism in the hematopoietic system.•Metabolic treatments could induce blast differentiation in patients with hematologic malignancies with a reduced toxicity. ZBTB7A is a transcription factor that regulates all three branches of hematopoietic differentiation while repressing the expression of key glycolytic enzymes and glucose transporters. Here, we propose that ZBTB7A acts as a link between differentiation and metabolism, two interconnected cellular processes. In particular, ZBTB7A can activate or repress metabolic programs necessary for the differentiation of specific cell lineages while controlling key pathways such as Notch signaling. Finally, the dual role of ZBTB7A has implications for the treatment of myeloid malignancies, where the block of differentiation could potentially be overcome by metabolic therapies with low toxicity. ZBTB7A is a transcription factor that regulates all three branches of hematopoietic differentiation while repressing the expression of key glycolytic enzymes and glucose transporters. Here, we propose that ZBTB7A acts as a link between differentiation and metabolism, two interconnected cellular processes. In particular, ZBTB7A can activate or repress metabolic programs necessary for the differentiation of specific cell lineages while controlling key pathways such as Notch signaling. Finally, the dual role of ZBTB7A has implications for the treatment of myeloid malignancies, where the block of differentiation could potentially be overcome by metabolic therapies with low toxicity. ZBTB7A is a transcription factor with an important role in the regulation of lineage fate decisions [1Lunardi A Guarnerio J Wang G Maeda T Pandolfi PP Role of LRF/Pokemon in lineage fate decisions.Blood. 2013; 121: 2845-2853Crossref PubMed Scopus (52) Google Scholar] and glycolysis [2Liu XS Haines JE Mehanna EK et al.ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis.Genes Dev. 2014; 28: 1917-1928Crossref PubMed Scopus (68) Google Scholar]. Pandolfi and colleagues performed most of the early research on Zbtb7a, also known as leukemia-related factor (Lrf) [reviewed in 1]. Using both Zbtb7a complete knockout (KO) and conditional hematopoietic KO mice, they found that Zbtb7a influences several branches of hematopoietic differentiation. First, Zbtb7a expression is indispensable for erythroid development, where it represses Bim-mediated apoptosis [3Maeda T Ito K Merghoub T et al.LRF is an essential downstream target of GATA1 in erythroid development and regulates BIM-dependent apoptosis.Dev Cell. 2009; 17: 527-540Abstract Full Text Full Text PDF PubMed Scopus (83) Google Scholar] and binds gene targets of the key erythroid transcription factor Gata1 [4Yu M Riva L Xie H et al.Insights into GATA-1-mediated gene activation versus repression via genome-wide chromatin occupancy analysis.Mol Cell. 2009; 36: 682-695Abstract Full Text Full Text PDF PubMed Scopus (242) Google Scholar]. Second, Zbtb7a promotes B-cell differentiation to the detriment of T cells through repression of the key lineage regulator Notch [5Maeda T Merghoub T Hobbs RM et al.Regulation of B versus T lymphoid lineage fate decision by the proto-oncogene LRF.Science. 2007; 316: 860-866Crossref PubMed Scopus (174) Google Scholar]. Third, Zbtb7a conditional and complete KO mice exhibited a reduced number of myeloid progenitors through a yet unknown mechanism [5Maeda T Merghoub T Hobbs RM et al.Regulation of B versus T lymphoid lineage fate decision by the proto-oncogene LRF.Science. 2007; 316: 860-866Crossref PubMed Scopus (174) Google Scholar,6Lee SU Maeda M Ishikawa Y et al.LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance.Blood. 2013; 121: 918-929Crossref PubMed Scopus (42) Google Scholar]. Finally, Zbtb7a expression in erythroblasts represses transcription of the Notch ligand Delta-like 4 (Dll4), playing an extrinsic role in hematopoietic stem cell (HSC) maintenance by avoiding their premature lymphoid differentiation [6Lee SU Maeda M Ishikawa Y et al.LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance.Blood. 2013; 121: 918-929Crossref PubMed Scopus (42) Google Scholar]. Interestingly, the role of ZBTB7A is not limited to lineage fate decisions, and it has been extensively linked to cancer because of a diversity of functions [7Constantinou C Spella M Chondrou V Patrinos GP Papachatzopoulou A Sgourou A The multi-faceted functioning portrait of LRF/ZBTB7A.Hum Genomics. 2019; 13: 66Crossref PubMed Scopus (15) Google Scholar]. Deletions in 19p13.3, the chromosomal region containing ZBTB7A, were described to be a common event in cancer progression [8Beroukhim R Mermel CH Porter D et al.The landscape of somatic copy-number alteration across human cancers.Nature. 2010; 463: 899-905Crossref PubMed Scopus (2681) Google Scholar,9Zack TI Schumacher SE Carter SL et al.Pan-cancer patterns of somatic copy number alteration.Nat Genet. 2013; 45: 1134-1140Crossref PubMed Scopus (1170) Google Scholar]. What is more, Liu et al. [10Liu XS Liu Z Gerarduzzi C et al.Somatic human ZBTB7A zinc finger mutations promote cancer progression.Oncogene. 2016; 35: 3071-3078Crossref PubMed Scopus (25) Google Scholar] described mutations in ZBTB7A in the context of solid cancers with a prevalence ranging from 1.1 to 4.2%. In the context of colon cancer, they could demonstrate that ZBTB7A directly represses the transcription of key genes for the glycolytic pathway (such as PFKP, PKM, and SLC2A3), thus acting as a tumor suppressor [2Liu XS Haines JE Mehanna EK et al.ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis.Genes Dev. 2014; 28: 1917-1928Crossref PubMed Scopus (68) Google Scholar]. ZBTB7A came into the spotlight of our research with the discovery of its frequent alterations in patients with core binding factor (CBF) leukemia (Figure 1) [11Hartmann L Dutta S Opatz S et al.ZBTB7A mutations inacute myeloid leukaemia with t(8;21) translocation.Nat Commun. 2016; 7: 11733Crossref PubMed Scopus (41) Google Scholar, 12Lavallee VP Lemieux S Boucher G et al.RNA-sequencing analysis of core binding factor AML identifies recurrent ZBTB7A mutations and defines RUNX1–CBFA2T3 fusion signature.Blood. 2016; 127: 2498-2501Crossref PubMed Scopus (48) Google Scholar, 13Faber ZJ Chen X Gedman AL et al.The genomic landscape of core-binding factor acute myeloid leukemias.Nat Genet. 2016; 48: 1551-1556Crossref PubMed Scopus (161) Google Scholar, 14Kawashima N Akashi A Nagata Y et al.Clinical significance of ASXL2 and ZBTB7A mutations and C-terminally truncated RUNX1–RUNX1T1 expression in AML patients with t(8;21) enrolled in the JALSG AML201 study.Ann Hematol. 2019; 98: 83-91Crossref PubMed Scopus (16) Google Scholar]. This subtype of acute myeloid leukemia (AML) is characterized by disruption of the CBF complex, which is crucial for normal hematopoiesis. The disruption can be caused by a translocation between chromosomes 8 and 21 [t(8;21)], an inversion of chromosome 16 [inv(16)], or an internal translocation of chromosome 16 [t(16;16)] [15Speck NA Gilliland DG Core-binding factors in haematopoiesis and leukaemia.Nat Rev Cancer. 2002; 2: 502-513Crossref PubMed Scopus (462) Google Scholar]. Curiously enough, there is no evidence of mutations in any other kind of leukemia, and mutations are an order of magnitude more prevalent in AML with t(8;21) translocation than in AML with inversion inv(16)/t(16;16) [16Opatz S Bamopoulos SA Metzeler KH et al.The clinical mutatome of core binding factor leukemia.Leukemia. 2020; 34: 1553-1562Crossref PubMed Scopus (44) Google Scholar]. This mutational pattern points toward an oncogenic collaboration between RUNX1–RUNX1T1 (the fusion gene resulting from t(8;21)) and ZBTB7A deficiency. Our observation that ZBTB7A overexpression is incompatible with RUNX1–RUNX1T1-directed stem cell expansion because of a block of cell cycle progression might explain this specific association of genetic lesions [17Redondo Monte E Wilding A Leubolt G et al.ZBTB7A prevents RUNX1–RUNX1T1-dependent clonal expansion of human hematopoietic stem and progenitor cells.Oncogene. 2020; 39: 3195-3205Crossref PubMed Scopus (11) Google Scholar]. Moreover, leukemia-specific ZBTB7A mutations present a loss of function phenotype, leading to upregulation of glycolysis. Finally, ZBTB7A blocks monocytic and HSC differentiation while promoting granulocytic and erythroid differentiation. Our K562 cell line model of ZBTB7A inactivation indicates that, indeed, ZBTB7A regulates glycolysis in the myeloid compartment (Figure 2A,C; Supplementary Figure E1, online only, available at: www.exphem.org). At the same time, ZBTB7A KO has a major impact on the regulation of differentiation, specifically through the Notch signaling pathway (Figure 2B,C; Supplementary Figure 1). In this perspective, we propose that the dual role of ZBTB7A in regulation of both glycolysis and differentiation is not a coincidence but rather a necessity. We suggest that ZBTB7A may regulate and allow for normal hematopoietic differentiation through alteration of metabolism. MYC and HIF1 are well-known regulators of glycolysis, and their corresponding pathways are responsive to stimuli such as lack of oxygen and proliferation signals, among others [18Dang CV MYC on the path to cancer.Cell. 2012; 149: 22-35Abstract Full Text Full Text PDF PubMed Scopus (2106) Google Scholar,19Denko NC Hypoxia, HIF1 and glucose metabolism in the solid tumour.Nat Rev Cancer. 2008; 8: 705-713Crossref PubMed Scopus (1272) Google Scholar]. The fact that ZBTB7A directly represses the expression of key glycolytic genes in an MYC- and HIF1-independent manner [2Liu XS Haines JE Mehanna EK et al.ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis.Genes Dev. 2014; 28: 1917-1928Crossref PubMed Scopus (68) Google Scholar] hints at the idea that regulation of metabolism through ZBTB7A goes beyond the response to an immediate need for energy. Rather, ZBTB7A may direct metabolic programs necessary for the differentiation of specific cell lineages.Supplementary Figure 1ZBTB7A controls the expression of key glycolytic and differentiation genes. K562 ZBTB7A knockout (KO) cells were produced using CRISPR/Cas9 technology and RNA-Seq was performed in cell lines derived from single cell clones. Box-plots show expression of indicated genes in wild type (CTRL) vs KO cells. TMM: trimmed mean of M values , ns: non-significant, ***: p-value<0.001, ****: p-value<0.0001.View Large Image Figure ViewerDownload Hi-res image Download (PPT) Different cell types require the activation of distinct metabolic programs to maintain their differentiation status or further differentiate [20McGraw TE Mittal V Stem cells: metabolism regulates differentiation.Nat Chem Biol. 2010; 6: 176-177Crossref PubMed Scopus (25) Google Scholar]. A well-known example is the reliance of human HSCs on glycolysis to maintain their stemness phenotype [21Simsek T Kocabas F Zheng J et al.The distinct metabolic profile of hematopoietic stem cells reflects their location in a hypoxic niche.Cell Stem Cell. 2010; 7: 380-390Abstract Full Text Full Text PDF PubMed Scopus (761) Google Scholar], while depending on mitochondrial oxidation to undergo differentiation [22Takubo K Nagamatsu G Kobayashi CI et al.Regulation of glycolysis by Pdk functions as a metabolic checkpoint for cell cycle quiescence in hematopoietic stem cells.Cell Stem Cell. 2013; 12: 49-61Abstract Full Text Full Text PDF PubMed Scopus (511) Google Scholar]. Furthermore, mutations in genes that enhance glycolysis block stem cell differentiation [23Yu WM Liu X Shen J et al.Metabolic regulation by the mitochondrial phosphatase PTPMT1 is required for hematopoietic stem cell differentiation.Cell Stem Cell. 2013; 12: 62-74Abstract Full Text Full Text PDF PubMed Scopus (218) Google Scholar]. Fatty acid oxidation (FAO) also plays a role in HSC maintenance and self-renewal, whereas FAO inhibition leads to linage commitment [24Ito K Carracedo A Weiss D et al.A PML-PPAR-delta pathway for fatty acid oxidation regulates hematopoietic stem cell maintenance.Nat Med. 2012; 18: 1350-1358Crossref PubMed Scopus (485) Google Scholar]. A similar effect is also observed in T cells, where regulatory T cells depend on FAO for their differentiation and maintenance, while effector T cells depend on glycolysis [25Michalek RD Gerriets VA Jacobs SR et al.Cutting edge: distinct glycolytic and lipid oxidative metabolic programs are essential for effector and regulatory CD4+ T cell subsets.J Immunol. 2011; 186: 3299-3303Crossref PubMed Scopus (1312) Google Scholar,26Shi LZ Wang R Huang G et al.HIF1alpha-dependent glycolytic pathway orchestrates a metabolic checkpoint for the differentiation of TH17 and Treg cells.J Exp Med. 2011; 208: 1367-1376Crossref PubMed Scopus (1191) Google Scholar]. Oburoglu et al. [27Oburoglu L Tardito S Fritz V et al.Glucose and glutamine metabolism regulate human hematopoietic stem cell lineage specification.Cell Stem Cell. 2014; 15: 169-184Abstract Full Text Full Text PDF PubMed Scopus (180) Google Scholar] further investigated the link between differentiation and metabolism in HSCs. They reported that erythroid differentiation depends on glutamine metabolism-dependent nucleotide synthesis. When they blocked glutamate metabolism with the glutamine analogue 6-diazo-5-oxo-L-norleucine (DON), HSCs were redirected into myeloid differentiation. Additionally, block of glycolysis using 2-deoxy-D-glucose (2DG) directed HSCs into the erythroid linage. Interestingly, this mechanism is not exclusive to the hematopoietic tissue. Reports have linked metabolism and differentiation in human pluripotent stem cells [28Zhang J Khvorostov I Hong JS et al.UCP2 regulates energy metabolism and differentiation potential of human pluripotent stem cells.EMBO J. 2011; 30: 4860-4873Crossref PubMed Scopus (382) Google Scholar], cardiac stem cells [29Chung S Dzeja PP Faustino RS Perez-Terzic C Behfar A Terzic A Mitochondrial oxidative metabolism is required for the cardiac differentiation of stem cells.Nat Clin Pract Cardiovasc Med. 2007; 4: S60-S67Crossref PubMed Scopus (384) Google Scholar], and embryonic stem cells [30Agathocleous M Love NK Randlett O et al.Metabolic differentiation in the embryonic retina.Nat Cell Biol. 2012; 14: 859-864Crossref PubMed Scopus (117) Google Scholar,31Yanes O Clark J Wong DM et al.Metabolic oxidation regulates embryonic stem cell differentiation.Nat Chem Biol. 2010; 6: 411-417Crossref PubMed Scopus (404) Google Scholar]. The fact that ZBTB7A is essential for terminal erythroid differentiation [3Maeda T Ito K Merghoub T et al.LRF is an essential downstream target of GATA1 in erythroid development and regulates BIM-dependent apoptosis.Dev Cell. 2009; 17: 527-540Abstract Full Text Full Text PDF PubMed Scopus (83) Google Scholar], while also blocking glycolysis [2Liu XS Haines JE Mehanna EK et al.ZBTB7A acts as a tumor suppressor through the transcriptional repression of glycolysis.Genes Dev. 2014; 28: 1917-1928Crossref PubMed Scopus (68) Google Scholar], directly correlates with the observation that 2DG promotes erythrocytic differentiation of HSCs [27Oburoglu L Tardito S Fritz V et al.Glucose and glutamine metabolism regulate human hematopoietic stem cell lineage specification.Cell Stem Cell. 2014; 15: 169-184Abstract Full Text Full Text PDF PubMed Scopus (180) Google Scholar]. In this context, we propose that ZBTB7A allows for erythroid differentiation because of BIM downregulation, while activating the metabolic program necessary for erythroid survival (Figure 2D). A similar example is the block of monocytic differentiation through ZBTB7A overexpression [17Redondo Monte E Wilding A Leubolt G et al.ZBTB7A prevents RUNX1–RUNX1T1-dependent clonal expansion of human hematopoietic stem and progenitor cells.Oncogene. 2020; 39: 3195-3205Crossref PubMed Scopus (11) Google Scholar]. Accordingly, inhibition of glycolysis by 2DG impairs monocyte-to-macrophage maturation because of insufficient pyruvate supply [32Suzuki H Hisamatsu T Chiba S et al.Glycolytic pathway affects differentiation of human monocytes to regulatory macrophages.Immunol Lett. 2016; 176: 18-27Crossref PubMed Scopus (51) Google Scholar]. At the same time, monocyte function highly depends on glycolysis, which is specifically mediated by upregulation of the glucose transporter SLC2A1 [33Lee MKS Al-Sharea A Shihata WA et al.Glycolysis is required for LPS-induced activation and adhesion of human CD14(+)CD16(–) monocytes.Front Immunol. 2019; 10: 2054Crossref PubMed Scopus (29) Google Scholar], under transcriptional control of ZBTB7A [17Redondo Monte E Wilding A Leubolt G et al.ZBTB7A prevents RUNX1–RUNX1T1-dependent clonal expansion of human hematopoietic stem and progenitor cells.Oncogene. 2020; 39: 3195-3205Crossref PubMed Scopus (11) Google Scholar]. However, other specific steps of myeloid development do not seem to follow this pattern. For example, ectopic expression of ZBTB7A promotes granulocytic differentiation [17Redondo Monte E Wilding A Leubolt G et al.ZBTB7A prevents RUNX1–RUNX1T1-dependent clonal expansion of human hematopoietic stem and progenitor cells.Oncogene. 2020; 39: 3195-3205Crossref PubMed Scopus (11) Google Scholar], a cell type with a highly glycolytic metabolism [34Kramer PA Ravi S Chacko B Johnson MS Darley-Usmar VM A review of the mitochondrial and glycolytic metabolism in human platelets and leukocytes: implications for their use as bioenergetic biomarkers.Redox Biol. 2014; 2: 206-210Crossref PubMed Scopus (255) Google Scholar]. This apparent contradiction could be explained by the fact that a metabolic program (i.e., repression of glycolysis) may only be needed during the differentiation process, and the resulting cell may still have other metabolic needs. In conclusion, the discovery of other genes mutated specifically in AML t(8;21) but not in AML inv(16), such as GATA2 [16Opatz S Bamopoulos SA Metzeler KH et al.The clinical mutatome of core binding factor leukemia.Leukemia. 2020; 34: 1553-1562Crossref PubMed Scopus (44) Google Scholar], forecasts that major discoveries are still to come in the field of oncogenic collaborations underlying myeloid malignancy. Our model of ZBTB7A inactivation in the erythroleukemia-like cell line K562 might be adequate for studying effects in the erythroid linage. However, further models of ZBTB7A mutation are required to study the effect on myeloid differentiation and metabolism beyond erythropoiesis. Altogether, there is increasing evidence that ZBTB7A may play a role as a hub between differentiation and metabolism, two interconnected mechanisms with translational implications in the field of hematology–oncology. Moreover, the mechanisms described in this article are likely to have an impact on solid tumor differentiation, as metabolism and differentiation are linked in healthy tissues other than blood [35Agathocleous M Harris WA Metabolism in physiological cell proliferation and differentiation.Trends Cell Biol. 2013; 23: 484-492Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar]. Understanding the interdependency of these two cellular processes may lead to the development of new therapies with low toxicity, for example, induction of blast differentiation through block or promotion of specific metabolic pathways. The authors declare no competing interests. PAG acknowledges support by the Wilhelm Sander-Stiftung (Förderantrag No. 2014.162.2), the Deutsche Forschungsgemeinschaft (Collaborative Research Centre 1243 "Cancer Evolution", Project A08) and the Munich Clinician Scientist Program (MCSP).