Almost 30% of all acute myeloid leukemias (AML) are associated with an internal tandem duplication (ITD) in the juxtamembrane domain of FMS-like tyrosine kinase 3 receptor (FLT3). Patients with FLT3-ITD mutations tend to have a poor prognosis. MicroRNAs (miRNAs) have a pivotal role in myeloid differentiation and leukemia. MiRNA-155 (MiR-155) was found to be upregulated in FLT3-ITD-associated AMLs. In this study, we discovered that FLT3-ITD signaling induces the oncogenic miR-155. We show in vitro and in vivo that miR-155 expression is regulated by FLT3-ITD downstream targets nuclear factor-κB (p65) and signal transducer and activator of transcription 5 (STAT5). Further, we demonstrate that miR-155 targets the myeloid transcription factor PU.1. Knockdown of miR-155 or overexpression of PU.1 blocks proliferation and induces apoptosis of FLT3-ITD-associated leukemic cells. Our data demonstrate a novel network in which FLT3-ITD signaling induces oncogenic miR-155 by p65 and STAT5 in AML, thereby targeting transcription factor PU.1.

Key Points miR-30c is a direct target of C/EBPα and upregulated by C/EBPα-p42. NOTCH1 is a direct target of miR-30c and regulated by C/EBPα and miR-30c.

Transcription factor C/EBPα is a master regulator of myelopoiesis and its inactivation is associated with acute myeloid leukemia. Deregulation of C/EBPα by microRNAs during granulopoiesis or acute myeloid leukemia development has not been studied. Here we show that oncogenic miR-182 is a strong regulator of C/EBPα. Moreover, we identify a regulatory loop between C/EBPα and miR-182. While C/EBPα blocks miR-182 expression by direct promoter binding during myeloid differentiation, enforced expression of miR-182 reduces C/EBPα protein level and impairs granulopoiesis in vitro and in vivo. In addition, miR-182 expression is highly elevated particularly in acute myeloid leukemia patients with C-terminal CEBPA mutations, thereby depicting a mechanism by which C/EBPα blocks miR-182 expression. Furthermore, we present miR-182 expression as a prognostic marker in cytogenetically high-risk acute myeloid leukemia patients. Our data demonstrate the importance of a controlled balance between C/EBPα and miR-182 for the maintenance of healthy granulopoiesis.