Figure 4.

ZDHHC6 is the predominant PAT for FLT3. (A) ZDHHC6 interacts with all FLT3 variants. 293T cells transfected with HA-GST (control) or HA-ZDHHC6, along with the indicated FLT3 constructs, were immunoprecipitated by HA-EZ agarose beads, followed by western blot (WB) analysis. (B) Examination of endogenous FLT3-ITD palmitoylation in MV4;11 cells depleted of ZDHHC2 or ZDHHC6 with 2 independent guide RNAs (gRNAs), by the APE assay. (C) Quantification of surface FLT3-ITD levels in MV4;11 cells depleted of ZDHHC6 by 2 different gRNAs in comparison with control (Ctrl) gRNA (n = 3). (D) WB analysis of signal transduction in MV4;11 cells depleted of ZDHHC6, in comparison with Ctrl gRNA. (E) Colony-forming capacity of MV4;11 cells depleted of ZDHHC6 in triplicate. (F) Bioluminescence imaging of NSG mouse recipients of MV4;11 cell transplants depleted of ZDHHC6 with 2 different gRNAs along with the control gRNA. (G) Quantification of bioluminescence signals from panel F. Single guide (sg) Ctrl (n = 7), sgZDHHC6-#1 (n = 6), or sgZDHHC6-#2 (n = 6). Each symbol represents an individual mouse. Means ± standard error of the mean are presented as vertical lines. (C,E) Data are presented as means ± standard deviation. In all relevant panels, *P < .05; **P < .01; ***P < .001; ns, not significant, as determined by 2-tailed Student t tests.