YTHDC1 regulates proliferation and survival of human AML cells in an m6A-dependent manner. (A) Comparison of the expression level of YTHDC1 in patients with primary AML vs healthy donors from The Cancer Genome Atlas database. (B) Comparison of the expression level of YTHDC1 in patients with primary AML bearing various chromosomal translocations vs those in BM HSCs collected from healthy donors. The expression values (detected by using Affymetrix exon arrays) were log2-transformed. The P value was detected by 1-way analysis of variance followed by multiple comparisons vs the HSC group. Normal: AML with normal karyotype; Complex: AML with complex karyotype. (C) Western blot showing protein level of YTHDC1 in AML patient cells, AML cell lines, and CD34+ cells from healthy donor. (D) Western blot indicating KD efficiency of YTHDC1 by specific shRNAs in NB-4, KASUMI-1, and MOLM-13 cells. Growth curve of MOLM-13 (E), KASUMI-1 (F), and NB-4 (G) cells expressed Scramble shRNA (Scr), YTHDC1 shRNA1# (Ysh1#), and YTHDC1 shRNA2# (Ysh2#); cells were counted every 2 days. (H) Flow cytometric analysis of apoptosis frequency of MOLM-13, KASUMI-1, and NB-4 cells expressed Scramble shRNA (Scr), YTHDC1 shRNA1# (Ysh1#), and YTHDC1 shRNA2# (Ysh2#). Annexin V+ indicated the apoptotic cells. (I) Colony-forming units of MOLM-13, KASUMI-1, and NB-4 cells expressed Scramble shRNA (Scr), YTHDC1 shRNA1# (Ysh1#), and YTHDC1 shRNA2# (Ysh2#). Colony number was counted 7 days after plating; 2000 cells per well for MOLM-13 and KASUMI-1 input; 1000 cells per well for NB-4 input. Flow cytometric analysis of differentiated cell frequency (CD11b+only, CD14+ only, and total) of MOLM-13 (J), KASUMI-1 (K), and NB-4 (L) cells expressed Scramble shRNA (Scr), YTHDC1 shRNA1# (Ysh1#), and YTHDC1 shRNA2# (Ysh2#). Data are presented as mean ± standard deviation; Student t test. *P < .05; **P < .01; ***P < .001.