Fig 1: IRS1 and KLF10 are targets of miRNA-126. (A) The predicted binding sites between miRNA-126 and IRS1, and KLF10, and relative luciferase activity in IRS1 and KLF10 3′UTR (wild-type/mutant) and miRNA-126. (B) mRNA levels of IRS1 and KLF10 in MDSCs. (C) Protein expressions of IRS1 and KLF10 in MDSCs. Data are shown as the means ± SD. *P < 0.05, **P < 0.01.
Fig 2: Overexpression of KLF10 inhibits proliferation and induces apoptosis of melanoma cells. (A) Western blotting and (B) reverse transcription-quantitative PCR were used to detected the expression levels of KLF10. (C) A Cell Counting Kit-8 assay was used to detect the levels of cell proliferation. (D) A colony formation assay was used to detect the cell cloning levels. Scale bar, 250 µm. (E) Cell apoptosis was detected by TUNEL staining. Scale bar, 50 µm. (F) Expression levels of apoptosis-related proteins (Bax, Bcl-2 and cleaved caspase 3) were examined by western blotting. ***P<0.001 vs. Ov-NC. KLF10, Kruppel-like factor 10; NC, negative control; Ov, overexpression.
Fig 3: KLF10 is downregulated in melanoma cells. (A) The Gene Expression Profiling Interactive Analysis website was used to investigate KLF10 expression in the tissues of patients with melanoma. (B) Western blotting and (C) reverse transcription-quantitative PCR were used to detect the expression levels of KLF10 in melanoma cells. *P<0.05, ***P<0.001 vs. HEM. KLF10, Kruppel-like factor 10; N, normal; SKCM, skin cutaneous melanoma; T, tumor.
Fig 4: KLF10 binds to ACSM3 and promotes the transcription of ACSM3. (A) The Gene Expression Profiling Interactive Analysis website revealed the expression levels of ACSM3. (B) The association between ACSM3 expression and overall survival rate in melanoma patients, (C) the association of ACSM3 and disease-free survival rate in patients with melanoma, and (D) the association between KLF10 expression and ACSM3 expression in patients with melanoma were examined. (E) Western blotting and (F) RT-qPCR were used to detect the expression levels of ACSM3 in melanoma cells. (G) JASPAR predicted the binding sites of KLF10 and the ACSM3 promoter. (H) Luciferase reporter gene assay was used to detect the ACSM3 promoter activity. (I) Immunoprecipitation further indicated that KLF10 could bind with ACSM3. (J) Western blotting and (K) RT-qPCR were used to detect the expression levels of ACSM3 after KLF10 overexpression. *P<0.05, ***P<0.001 vs. HEM, ACSM3 + Ov-NC, IgG or Ov-NC. ACSM3, acyl-CoA medium-chain synthetase 3; KLF10, Kruppel-like factor 10; MUT, mutant; N, normal; NC, negative control; Ov, overexpression; RT-qPCR, reverse transcription-quantitative PCR; SKCM, skin cutaneous melanoma; T, tumor; TPM, transcripts per million; WT, wild-type.
Fig 5: Overexpression of KLF10 inhibits the invasion and migration of melanoma cells. (A) Wound-healing and (B) Transwell assays were used to detect the levels of migration and invasion of melanoma cells. Scale bar, 100 µm. (C) Expression levels of metastasis-related proteins (MMP2 and MMP9) were detected by western blotting. ***P<0.001 vs. Ov-NC. KLF10, Kruppel-like factor 10; NC, negative control; Ov, overexpression.
Supplier Page from Abcam for Anti-KLF10 antibody [EPR12102(2)]