Figure 4.
ZDHHC21-induced AML proliferation, stemness, and differentiation blockage depends on its palmitoyl acyltransferase activity. (A) The protein levels of mZDHHC21 in shZDHHC21-transfected HL60 cells re-expressing mZDHHC21-HA or mZDHHC21(C120S)-HA. (B) Cell proliferation of shZDHHC21-transfected AML cells re-expressing mZDHHC21-HA or mZDHHC21(C120S)-HA in HL60 cells. (C) The colony-forming ability of HL60 cells after culturing in soft agar. (D-E) Differentiation analysis of shZDHHC21-transfected HL60 cells re-expressing mZDHHC21-HA or mZDHHC21-C120S-HA. (D) NBT-reducing activity. (E) Cell morphological analysis after Wright-Giemsa staining. (F-G) mZDHHC21 overexpression but not mZDHHC21 (C120S) rescued shZDHHC21-induced tumor growth arrest in the subcutaneous xenograft model. NB4 cells infected with vector, mZDHHC21, or mZDHHC21 (C120S) were injected subcutaneously into 4- to 5-week-old female NSG mice and then received an intratumoral injection of virus containing shCtrl or shZDHHC21. (F) Tumor growth of NB4 xenografts. Tumor volume growth curves are presented as the mean ± SE, n = 5. The red arrow indicates the time of intratumoral injection. (G) Tumor weight of each group. (H-I) Cell differentiation analysis of AML cells treated with 2BP for 3 days. (H) CD11b expression of HL60 cells and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). (I) CD11b expression of other AML cell lines treated with 2BP at a single concentration. MV411 (40 μM), OCI-AML3 (60 μM), THP-1 (60 μM), and Kasumi-1 (60 μM). (J) CD11b expression of AML cells treated with TM by a single concentration. HL60 (1 μg/mL), NB4 (0.03125 μg/mL), MV411 (0.25 μg/mL), OCI-AML3 (1 μg/mL), THP-1 (0.125 μg/mL), and Kasumi-1 (0.0625 μg/mL). (K) CD14 expression of HL60 and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). (L) NBT-reducing activity of HL60 cells and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). Data are shown as mean ± SD, n = 3. n.s., P > .05; ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. The significance analysis was conducted by a two-tailed unpaired t test or a one-way analysis of variance. WT, wild-type.

ZDHHC21-induced AML proliferation, stemness, and differentiation blockage depends on its palmitoyl acyltransferase activity. (A) The protein levels of mZDHHC21 in shZDHHC21-transfected HL60 cells re-expressing mZDHHC21-HA or mZDHHC21(C120S)-HA. (B) Cell proliferation of shZDHHC21-transfected AML cells re-expressing mZDHHC21-HA or mZDHHC21(C120S)-HA in HL60 cells. (C) The colony-forming ability of HL60 cells after culturing in soft agar. (D-E) Differentiation analysis of shZDHHC21-transfected HL60 cells re-expressing mZDHHC21-HA or mZDHHC21-C120S-HA. (D) NBT-reducing activity. (E) Cell morphological analysis after Wright-Giemsa staining. (F-G) mZDHHC21 overexpression but not mZDHHC21 (C120S) rescued shZDHHC21-induced tumor growth arrest in the subcutaneous xenograft model. NB4 cells infected with vector, mZDHHC21, or mZDHHC21 (C120S) were injected subcutaneously into 4- to 5-week-old female NSG mice and then received an intratumoral injection of virus containing shCtrl or shZDHHC21. (F) Tumor growth of NB4 xenografts. Tumor volume growth curves are presented as the mean ± SE, n = 5. The red arrow indicates the time of intratumoral injection. (G) Tumor weight of each group. (H-I) Cell differentiation analysis of AML cells treated with 2BP for 3 days. (H) CD11b expression of HL60 cells and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). (I) CD11b expression of other AML cell lines treated with 2BP at a single concentration. MV411 (40 μM), OCI-AML3 (60 μM), THP-1 (60 μM), and Kasumi-1 (60 μM). (J) CD11b expression of AML cells treated with TM by a single concentration. HL60 (1 μg/mL), NB4 (0.03125 μg/mL), MV411 (0.25 μg/mL), OCI-AML3 (1 μg/mL), THP-1 (0.125 μg/mL), and Kasumi-1 (0.0625 μg/mL). (K) CD14 expression of HL60 and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). (L) NBT-reducing activity of HL60 cells and NB4 cells treated with 2BP at a series of concentrations (0, 10, 20, 40, 60 μM). Data are shown as mean ± SD, n = 3. n.s., P > .05; ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. The significance analysis was conducted by a two-tailed unpaired t test or a one-way analysis of variance. WT, wild-type.

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