A novel miR-375-HOXB3-CDCA3/DNMT3B regulatory circuitry contributes to leukemogenesis in acute myeloid leukemia

Abstract

BackgroundAcute myeloid leukemia (AML) is a heterogeneous group of hematopoietic malignancies due to sophisticated genetic mutations and epigenetic dysregulation. MicroRNAs (miRNAs), a class of small non-coding RNAs, are important regulators of gene expression in all biological processes, including leukemogenesis. Recently, miR-375 has been reported to be a suppressive miRNA in multiple types of cancers, but its underlying anti-leukemia activity in AML is largely unknown.MethodsQuantitative reverse transcriptase PCR (qRT-PCR) was used to measure the expression of miR-375 and HOXB3 in leukemic cells and normal controls. Targets of miR-375 were confirmed by western blot and luciferase assay. Phenotypic effects of miR-375 overexpression and HOXB3 knockdown were assessed using viability (trypan blue exclusion assay), colony formation/replating, as well as tumor xenograft assays in vivo.ResultsThe expression of miR-375 was substantially decreased in leukemic cell lines and primary AML blasts compared with normal controls, because DNA hypermethylation of precursor-miR-375 (pre-miR-375) promoter was discovered in leukemic cells but not in normal controls. Lower expression of miR-375 predicted poor outcome in AML patients. Furthermore, forced expression of miR-375 not only decreased proliferation and colony formation in leukemic cells but also reduced xenograft tumor size and prolonged the survival time in a leukemia xenograft mouse model. Mechanistically, overexpression of miR-375 reduced HOXB3 expression and repressed the activity of a luciferase reporter through binding 3′-untranslated regions (3’-UTR) of HOXB3 mRNA. Overexpression of HOXB3 partially blocked miR-375-induced arrest of proliferation and reduction of colony number, suggesting that HOXB3 plays an important role in miR-375-induced anti-leukemia activity. Knockdown of HOXB3 by short hairpin RNAs reduced the expression of cell division cycle associated 3 (CDCA3), which decreased cell proliferation. Furthermore, HOXB3 induced DNA methyltransferase 3B (DNMT3B) expression to bind in the pre-miR-375 promoter and enhanced DNA hypermethylation of pre-miR-375, leading to the lower expression of miR-375.ConclusionsCollectively, we have identified a miR-375-HOXB3-CDCA3/DNMT3B regulatory circuitry which contributes to leukemogenesis and suggests a therapeutic strategy of restoring miR-375 expression in AML.Electronic supplementary materialThe online version of this article (10.1186/s12885-018-4097-z) contains supplementary material, which is available to authorized users.

Role of microRNAs, circRNAs and long noncoding RNAs in acute myeloid leukemia

“…For example, miR-375 could suppress HOXB3 expression and cause AML cell proliferation arrest and colony reduction. In return, HOXB3 enhanced DNMT3B's binding to the promoter of miR-375, leading to DNA hypermethylation and lower expression of miR-375 [56].…”

Identification of Acute Myeloid Leukemia Bone Marrow Circulating MicroRNAs

“…For example, miR-34a is strongly expressed in AML patient bone marrow fluid and is known to be downregulated in leukemia cells in the marrow and peripheral blood serum of patients [ 37 , 38 ] and appears to play an important role as a tumor suppressor and immunotherapeutic agent [ 39 , 40 ]. The expression of miR-375 was also downregulated in leukemic cell lines and primary AML blasts compared to normal controls, and its overexpression decreased proliferation and colony formation, reduced xenograft tumor size and prolonged the survival time in a leukemia xenograft mouse model [ 41 ]. MiR-576-3p was shown to be downregulated in colorectal [ 42 ] and bladder cancer [ 43 ], to inhibit the migration and invasion of lung adenocarcinoma [ 44 ] and to significantly inhibit the migration and pro-angiogenic abilities of glioma cells under hypoxic conditions [ 45 ].…”

Liquiritin represses proliferation , migration and invasion of colorectal cancer cells through inhibition of the miR-671/HOXB3 signaling pathway

“…In contrast to normal colonic epithelial cells NCM460, miR-671 level was elevated, and the HOXB3 level was downregulated in colorectal cancer cells ( Figure S1C and 1D). On the other hand, according to previous studies, miR-671 and HOXB3 levels are abnormally expressed and exhibit important effects on the growth and metastasis in the pathogenesis of cancer [16][17][18][32][33][34]. Therefore, we further examined the dose-dependent effect of liquiritin on the expression levels of miR-671 and HOXB3 in vitro and in vivo.…”