Figure 4. METTL14 blocks myeloid differentiation of human AML cells.
(A,B) Effect of METTL14 knockdown (A) or overexpression (B) on AML cell growth/proliferation was analyzed. The knockdown efficiency of shRNAs were confirmed by western blot and shown on the left (A).
(C,D) Flow cytometric analysis of CD11b+ and/or CD14+ cell populations (C) or Wright-Giemsa staining of cytospin slides (D). Arrows indicate differentiated cells. Bar= 20 µm.
(E,F) Effects of METTL14 knockdown on cell proliferation (E) and differentiation (F) of leukemia blasts (CD34+) from 3 primary AML patients.
(G–I) Effects of METTL14 knockdown on progression of human AML cells in NSGS (Wunderlich et al., 2010) mice. MM6 cells were transduced with shMT14-#2 or scramble shRNA. After selected with puromycin for two passages, cells were injected into NSGS mice (0.3×106 cells/mouse) via tail vein. Kaplan–Meier curves were shown in (G), engraftment of MM6 cells in BM and PB of NSGS mice was shown in (H), while representative images of liver tissues were shown in (I).
(J) MM6 and NB4 cells with or without METTL14 knockdown were treated with DMSO or ATRA (100 nM) for 72 hours and subjected to flow cytometric analysis. The percentages of each cell population were summarized on right.
(K) AML cells were transduced with empty vector or METTL14 overexpression vectors and treated with ATRA (20 nmol/L for 24 hours for NB4, 200 nmol/L for 48 hours for U937) before subjected to flow cytometric analysis. Percentages of cells with CD11b staining in each group were shown.
*, p < 0.05; **, p <0.01; ***, p < 0.001; t-test (for Figures 4A, 4B, 4C, 4E, 4F, and 4K; Mean±SD values are shown) or log-rank test (for Figure 4G). See also Figure S4.