Abstract

Acute promyelocytic leukemia (APL), a distinct subtype of acute myeloid leukemia, is characterized by an oncogenic fusion protein of a translocation between chromosomes 15 and 17, promyelocytic leukemia/retinoic acid receptor a (PML/RARα). PML/RARα plays an essential role in the leukemogenesis of APL by interfering with its target genes, eventually leading to a differentiation block at the promyelocytic stage and a hyperproliferation of blocked promyelocytes; both are the hallmarks of APL. It has been well-documented that cell differentiation and proliferation are two fundamental but distinct, even mutually exclusive, processes. In APL, the leukemic differentiation block is generally attributed to deregulated transcription of hematopoietic differentiation related transcription factors, RARα and PU.1, which is disrupted by PML/RARα. On the other hand, altered proliferation is usually caused by the impairment of the formation of functional PML nuclear bodies that act as a growth suppressor. There is emerging evidence to suggest that cell differentiation and proliferation can be controlled by the same regulator or pathway. Several studies have demonstrated that cell cycle regulators, especially CDK inhibitors (CKIs), are involved in the regulation of differentiation in addition to their well-documented function of controlling the cell cycle. Based on the exploration of verified PML/RARα binding sites derived from several genome-wide screening, of particular interest is CDKN2D, one of the INK4 family CKIs encoding p19INK4D. To understand the mechanisms coordinating these two processes in APL, we investigated the regulatory mechanism of the negative cell cycle regulator CDKN2D by the PML/RARα fusion protein and its role in differentiation and proliferation. We found that CDKN2D expression in APL cells was significantly lower than that in normal promyelocytes. Chromatin immunoprecipitation and luciferase reporter assays showed that PML/RARα directly bound to and inhibited the transactivation of the CDKN2D promoter. Further evidence by the truncated CDKN2D promoters revealed the localization of PML/RARa binding, which resided on the ER8 sites of the promoter. Moreover, ectopic expression of CDKN2D induced G0/G1 phase arrest and a partial granulocytic differentiation in APL-derived NB4 cells, suggesting its role in regulating both cell proliferation and granulocytic differentiation. Finally, we found that ATRA could significantly induce CDKN2D expression in APL cells. Knockdown of CDKN2D expression during ATRA treatment partially blocked the ATRA induced-differentiation and cell cycle arrest, further supporting our findings that CDKN2D expression was required for these two processes. Collectively, our data indicate that CDKN2D repression by PML/RARα disrupts both proliferation and differentiation to facilitate the pathogenesis of APL, and induced-expression of CDKN2D by ATRA alleviates the disruption of both processes to ensure treatment efficiency. This study suggests a mechanism for coupling proliferation and differentiation in leukemic cells through the action of CDKN2D.

Disclosures:

No relevant conflicts of interest to declare.

Author notes

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Asterisk with author names denotes non-ASH members.