Mice expressing a transgene encoding the transcription factor NF-E2 in hematopoietic cells exhibit features of myeloproliferative neoplasms, including thrombocytosis, Epo-independent colony formation, stem and progenitor cell overabundance, leukocytosis, and progression to acute myeloid leukemia.

We have previously described novel histone acetyltransferase (HAT) inhibitors that block neuroblastoma cell growth in vitro. Here we show that two selected pyridoisothiazolone HAT inhibitors, PU139 and PU141, induce cellular histone hypoacetylation and inhibit growth of several neoplastic cell lines originating from different tissues. Broader in vitro selectivity profiling shows that PU139 blocks the HATs Gcn5, p300/CBP-associated factor (PCAF), CREB (cAMP response element-binding) protein (CBP) and p300, whereas PU141 is selective toward CBP and p300. The pan-inhibitor PU139 triggers caspase-independent cell death in cell culture. Both inhibitors block growth of SK-N-SH neuroblastoma xenografts in mice and the PU139 was shown to synergize with doxorubicin in vivo. The latter also reduces histone lysine acetylation in vivo at concentrations that block neoplastic xenograft growth. This is one of the very few reports on hypoacetylating agents with in vivo anticancer activity.

Mutations in the transcription factor NF-E2 in patients with myeloproliferative neoplasms result in a truncated protein that enhances the function of wild-type NF-E2 and causes erythrocytosis and throbocytosis in a mouse model.

The transcription factor "nuclear factor erythroid 2" (NFE2) is overexpressed in the majority of patients with myeloproliferative neoplasms (MPNs). In murine models, elevated NFE2 levels cause an MPN phenotype with spontaneous leukemic transformation. However, both the molecular mechanisms leading to NFE2 overexpression and its downstream targets remain incompletely understood. Here, we show that the histone demethylase constitutes a novel NFE2 target gene. JMJD1C levels are significantly elevated in polycythemia vera (PV) and primary myelofibrosis patients; concomitantly, global H3K9me1 and H3K9me2 levels are significantly decreased. JMJD1C binding to the promoter is increased in PV patients, decreasing both H3K9me2 levels and binding of the repressive heterochromatin protein-1α (HP1α). Hence, JMJD1C and NFE2 participate in a novel autoregulatory loop. Depleting JMJD1C expression significantly reduced cytokine-independent growth of an MPN cell line. Independently, NFE2 is regulated through the epigenetic JAK2 pathway by phosphorylation of H3Y41. This likewise inhibits HP1α binding. Treatment with decitabine lowered H3Y41ph and augmented H3K9me2 levels at the locus in HEL cells, thereby increasing HP1α binding, which normalized NFE2 expression selectively in JAK2-positive cell lines.

No abstract

© Ferrata Storti Foundationorganelles as well as the cell nucleus. 4 Elimination of organelles like ribosomes and mitochondria, called autophagy in general, or more specifically ribophagy and mitophagy, respectively, is an essential step in the maturation of reticulocytes to erythrocytes.

No abstract

Expression of the transcription factor and epigenetic modulator “nuclear factor erythroid-2” (NF-E2) is aberrantly elevated in patients with Myeloproliferative Neoplasms (MPN). We have recently shown that NF-E2 overexpression in a murine model causes a phenotype similar to MPN in humans. This includes thrombocytosis, leukocytosis, expansion of the stem- and progenitor cell compartments as well as spontaneous transformation to acute leukemia. However, both the downstream pathways by which this transcription factor exerts its effects as well as the mechanisms leading to its overexpression in MPN patients remain incompletely characterized. We show here that the histone demethylases JMJD1C and JMJD2C constitute novel NF-E2 target genes. Using chromatin immunoprecipitation (ChIP) we demonstrate NF-E2 binding to the JMJD1C and JMJD2C loci. Both JMJD1C and JMJD2C protein levels are statistically significantly elevated in patients with polycythemia vera (PV) compared to healthy controls. JMJD1C and 2C proteins mediate the demethylation of histone H3K9, converting H3K9me2 via H3K9me to unmethylated H3K9. Consistent with increased demethylase activity, we observed statistically significantly decreased levels of global H3K9me and H3K9me2 in PV patients compared to healthy controls. We subsequently investigated whether the NF-E2 gene is regulated by epigenetic mechanisms and whether these are altered in PV patients. Indeed, using ChIP we demonstrate presence of the repressive H3K9me2 mark at three sites within the NF-E2 locus in healthy control granulocytes, where NF-E2 is expressed at low levels. These repressive histone marks are significantly reduced or completely absent in PV patients, which overexpress NF-E2. At the same time, binding of the repressive heterochromatin protein 1-alpha (HP1a) to the NF-E2 gene, while present in healthy controls, is significantly reduced in PV patients. HP1a chromatin binding is dependent on the presence of the H3K9me2 modification. Decreased levels of H3K9me2 and decreased HP1a binding are therefore both consistent with elevated NF-E2 expression in PV. We hypothesized that increased levels of the H3K9me2 demethylase JMJD1C in PV patients are responsible for the loss of this histone mark in the NF-E2 gene. Consistent with this model, we demonstrate significantly increased binding of JMJD1C to the NF-E2 locus in purified primary PV granulocytes. Our data demonstrate that the NF-E2 gene is regulated by epigenetic mechanisms and that these histone modifications are perturbed in PV patients. Moreover, NF-E2 participates in an auto-regulatory loop, by directly regulating transcription of the histone demethylase JMJD1C. Increased levels of NF-E2 thus cause increased JMJD1C expression. The JMJD1C protein, in turn, demethylates H3K9me2 residues in the NF-E2 locus, thereby further augmenting NF-E2 expression. Besides sustaining this novel autoregulatory loop, increased JMJD1C activity may contribute to MPN pathology by altering gene expression at additional loci. We sought to determine whether elevated NF-E2 levels could be normalized by pharmacological intervention. Treatment with the nucleoside analogue Decitabine reduced NF-E2 expression, and restored both the H3K9me2 marks and HP1a binding to the NF-E2 locus. These data provide a molecular rational for pre-clinical investigation of the effects of histone demethylase inhibitors and Decitabine on MPN cell biology and, subsequently, for a phase I trial investigating a combination of Decitabine and histone demethylase inhibitors in MPN patients. Disclosures: No relevant conflicts of interest to declare.