Low Autophagy (ATG) Gene Expression Is Associated with an Immature AML Blast Cell Phenotype and Can Be Restored during AML Differentiation Therapy

Figure 6

miR-106a targets ULK1 and attenuates ATRA-induced AML differentiation. (a) Inhibition of ULK1 attenuates neutrophil differentiation of APL cells. NB4 cells stably expressing a scramble control shRNA (Ctrl) or shRNAs targeting ULK1 (shULK1 #1 and shULK1 #2) were treated with ATRA for 4 days. qPCR for ULK1 and CEBPE as well as FACS analysis of CD11b is shown. Data are mean ± s.e.m. of three independent experiments. (b) ULK1, CEBPE, and p21^CIP1 expressions were detected by qPCR in NB4 cells stably transduced with a scrambled control (NB4) or a lentiviral vector expressing miR-106a precursors (NB4 106a). Cells were treated with 1 μM ATRA for 4 days. Results were normalized to HMBS and are shown as -fold change relative to the corresponding untreated control cell line. (e) qPCR analysis of miR-106a expression in NB4 control and miR-106a overexpressing NB4 cells. (f) Western blot analysis of ULK1 expression in control, anti-miR-106a, and miR-106a-expressing NB4 cells upon ATRA treatment for 4 days. GAPDH was used as a loading control. (g) Total RNA was isolated from 16 AML patients, and qPCR was performed to determine miR-106a and ULK1 mRNA levels, respectively. SNORA38B and 5s rRNA were used as reference RNAs for the microRNA qPCR and HMBS and ABL1 as reference mRNAs for the mRNA qPCR. ΔCt values of ULK1 and miR-106a were calculated and plotted against each other. Linear regression and Pearson were calculated using Prism software. Mann–Whitney U test, , , .

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