To investigate the effect of zinc finger aspartate-histidine-histidine-cysteine-type palmitoyltransferase 9 (ZDHHC9) expression on proliferation, migration and apoptosis of breast cancer and its mechanism.

The expression of ZDHHC9 in breast cancer and its relationship with prognosis were analyzed using The Cancer Genome Atlas (TCGA) database. Lentivirus infection was used to transfer short hairpin RNA (shRNA) into HCC1937 cells, MCF7 cells, and SKBR3 cells to knock down ZDHHC9. One control group (shNC) and two knockdown groups (sh1, sh2) were set for each cell line. The effects of ZDHHC9 on the proliferation of breast cancer cells were determined by cell counting kit-8 (CCK-8) cell proliferation assay, 5-ethynyl-2′-deoxyuridine-555 (EdU) cell proliferation assay and plate clone formation assay. The effect of ZDHHC9 on migration and invasion ability was verified by scratch assay and Transwell cell invasion assay. The effect of ZDHHC9 on apoptosis of cancer cells was detected by flow cytometry. Western blotting assay was used to observe the effect of down-regulated ZDHHC9 on phosphatidylinositol 3 kinase/protein kinase B pathway (PI3K/Akt pathway) and extracellular signal-regulated kinase pathway (ras/raf/MEK/Erk pathway). The t test was used for comparison between groups.

TCGA database analysis showed ZDHHC9 was highly expressed in breast cancer and was associated with poor prognosis. CCK-8 assay showed that the proliferation of breast cancer cells was inhibited after down-regulating ZDHHC9 (HCC1937-shNC vs. HCC1937-sh1: 2.577±0.012 vs. 1.887±0.077, t=15.319, P<0.01; MCF7-shNC vs. MCF7-sh1: 0.905±0.016 vs. 0.398±0.053, t=16.034, P<0.01; SKBR3-shNC vs. SKBR3-sh1: 2.322±0.090 vs. 1.786±0.022, t=10.067, P<0.01). The results of plate cloning experiments suggested that ZDHHC9 reduced colony formation (HCC1937-shNC vs. HCC1937-sh1: 472.333±48.211 vs. 362.667±43.108, t=7.637, P<0.01; MCF7-shNC vs. MCF7-sh1: 176.667±37.448 vs. 79.667±27.227, t=3.639, P<0.01; SKBR3-shNC vs. SKBR3-sh1: 296.333±19.553 vs. 210.667±11.676, t=6.515, P<0.01). Cell scratch test confirmed that Down-regulated ZDHHC9 Inhibited the migration of breast cancer cells (HCC1937-shNC vs. HCC1937-sh1: 0.536±0.046 vs. 0.235±0.010, t=10.969, P<0.01; SKBR3-shNC vs. SKBR3-sh1: 0.429±0.021 vs. 0.231±0.060, t=5.381, P<0.01). Transwell invasion assay showed that the invasion ability of breast cancer cells decreased after ZDHHC9 was down-regulated [HCC1937-shNC vs. HCC1937-sh1: (211.333±11.930) vs. 73.667±5.508, t=18.146, P<0.01; SKBR3-shNC vs. SKBR3-sh1: 159.333±11.590 vs. 45.333±7.024, t=14.570, P<0.01]. Flow cytometry indicated that down-regulation of ZDHHC9 increased apoptosis (HCC1937-shNC vs. HCC1937-sh1: 0.121±0.007 vs. 0.157±0.004, t=6.634, P<0.01; SKBR3-shNC vs. SKBR3-sh1: 0.124±0.005 vs. 0.142±0.005, t=6.218, P<0.01). Western blotting showed that ZDHHC9 knockdown inhibited the activation of PI3K/Akt and ras/raf/MEK/Erk pathways (for relative expression of phosphorylated protein kinase B (p-Akt), MCF7-shNC vs. MCF7-sh1: 0.931±0.005 vs. 0.707±0.002, t=24.193, P<0.01; for relative expression of phosphorylated extracellular signal-regulated kinase (p-ERK1/2), MCF7-shNC vs. MCF7-sh1: 1.418±0.010 vs. 1.206±0.053, t=6.748, P<0.01).

ZDHHC9 knockdown inhibited the proliferation, migration and invasion of breast cancer cells, and promoted cell apoptosis.

Breast cancer; Zinc finger aspartate-histidine-histidine-cysteine-type palmitoyltransferase 9; Proliferation; Migration; Apoptosis