PU.1 is linking the glycolytic enzyme HK3 in neutrophil differentiation and survival of APL cells

Abstract

The transcription factor PU.1 is a master regulator of myeloid differentiation and function. On the other hand, only scarce information is available on PU.1-regulated genes involved in cell survival. We now identified the glycolytic enzyme hexokinase 3 (HK3), a gene with cytoprotective functions, as transcriptional target of PU.1. Interestingly, HK3 expression is highly associated with the myeloid lineage and was significantly decreased in acute myeloid leukemia patients compared with normal granulocytes. Moreover, HK3 expression was significantly lower in acute promyelocytic leukemia (APL) compared with non-APL patient samples. In line with the observations in primary APL patient samples, we observed significantly higher HK3 expression during neutrophil differentiation of APL cell lines. Moreover, knocking down PU.1 impaired HK3 induction during neutrophil differentiation. In vivo binding of PU.1 and PML-RARA to the HK3 promoter was found, and PML-RARA attenuated PU.1 activation of the HK3 promoter. Next, inhibiting HK3 in APL cell lines resulted in significantly reduced neutrophil differentiation and viability compared with control cells. Our findings strongly suggest that HK3 is: (1) directly activated by PU.1, IntroductionAcute promyelocytic leukemia (APL or AML-M3) accounts for 5% to 8% of acute myeloid leukemia (AML) subtypes. 1 APL is characterized by a balanced chromosomal translocation involving the promyelocytic leukemia (PML) gene on chromosome 15 and the retinoic acid receptor␣ (RARA) on chromosome 17 that results in expression of the oncogenic fusion gene PML-RARA. 2 PML-RARA is a gain-of-function protein that promotes leukemic transformation by impairing the formation of functional PML nuclear bodies 3 and repressing RARA target genes in a dominantnegative manner. 4 This repression interferes with gene expression programs involved in differentiation, apoptosis, and self-renewal and leads to increased cell survival and inhibition of terminal differentiation, with an accumulation of promyelocytes. The block in differentiation that is observed in APL cells can be reverted with pharmacologic doses of ATRA by triggering PML-RARA degradation and thus restoring normal myeloid differentiation (reviewed by de Thé and Chen 5 ).The Ets-family member PU.1 is a master transcriptional regulator of myeloid differentiation, 6,7 as evidenced in PU.1 null mice that die at birth because of the lack of functional myeloid cells. 7 Inhibition of PU.1-induced myeloid differentiation may represent a critical step in APL leukemogenesis. On the one hand, PU.1 is inhibited at the transcriptional level by PML-RARA binding to its promoter region 8 ; on the other hand, a recent study identified PML-RARA as a binding partner of PU.1, 4 resulting in repression of PU.1-activated genes. A majority of PU.1 target genes are directly involved in myeloid differentiation and function, such as CD11b, CD45, the granulocyte/macrophage colonystimulating factor receptor (GM-CSFR), myeloperoxidase, lysozyme, and neutrophil elastase (reviewed...

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