FAT1 and FAT4 somatic mutations in FLT3-ITD AML at DX and REL. (A) Schematic of FAT1 and FAT4 genes with the locations of alterations. Black or red triangles show either missense or stop-gain, respectively. Red arrow represents missense mutation present in AML TCGA data set. Conserved domains are displayed by using UniProt (http://www.uniprot.org/). (B) IGV (http://www.broadinstitute.org/igv) heat map of 4q35.2 shows deletional peak with FAT1 deletion culled from 200 AML patients (TCGA consortium; http://cancergenome.nih.gov). (C) Real-time and protein blot display knockdown of FAT1 in MV4-11 cells. Quantitative PCR data represent means ± SD; n = 3. **P ≤ .01. (D) Cell growth showed increased proliferation with FAT1 knockdown in MV4-11 cells (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay; quadruplicate experiments). Data represent means ± SD. *P ≤ .05. (E) Cell counting assay: 0.5 × 10⁵ cells were plated in 6-well plates in quadruplets, and cell proliferation was measured by counting cells over a 5-day period. Results are shown as means ± SD; n = 4. (F) Methylcellulose colony assay of MV4-11 cells transfected with either scramble control or FAT1 siRNA. Data represent means ± SD; n = 3. *P ≤ .05.