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. 2022 Jan 15;22(1):23.
doi: 10.1186/s12935-022-02449-6.

The functions and prognostic value of Krüppel-like factors in breast cancer

Ke-Yun Zhu #  1   2 , Yao Tian #  3 , Ying-Xi Li #  4 , Qing-Xiang Meng  2   5 , Jie Ge  2   6 , Xu-Chen Cao  2   6 , Ti Zhang  7   8 , Yue Yu  9   10
Affiliations

The functions and prognostic value of Krüppel-like factors in breast cancer

Ke-Yun Zhu et al. Cancer Cell Int. .

Abstract

Background: Krüppel-like factors (KLFs) are zinc finger proteins which participate in transcriptional gene regulation. Although increasing evidence indicate that KLFs are involved in carcinogenesis and progression, its clinical significance and biological function in breast cancer are still limited.

Methods: We investigated all the expression of KLFs (KLF1-18) at transcriptional levels by using Oncomine and Gene Expression Profiling Interactive Analysis (GEPIA). The mRNA and protein expression levels of KLFs were also determined by using RT-qPCR and immunohistochemistry, respectively. CBioPortal, GeneMANIA and STRING were used to comprehensive analysis of the molecular characteristics of KLFs. The clinical value of prognostic prediction based on the expression of KLFs was determined by using the KM plotter. The relevant molecular pathways of KLFs were further analyzed by using Gene Set Enrichment Analysis (GSEA) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database. Finally, we investigated the effect of KLF2 and KLF15 on biological behavior of breast cancer cells in vitro.

Results: The expression of KLF2/4/6/8/9/11/15 was significantly down-regulated in breast cancer. The patients with high KLF2, KLF4 or KLF15 expression had a better outcome, while patients with high KLF8 or KLF11 had a poor prognosis. Furthermore, our results showed that KLF2 or KLF15 can be used as a prognostic factor independent on the other KLFs in patients with breast cancer. Overexpression of KLF2 or KLF15 inhibited cell proliferation and migration, and blocked cell cycle at G0/G1 phase, resulting in cell apoptosis.

Conclusions: KLF2 and KLF15 function as tumor suppressors in breast cancer and are potential biomarkers for prognostic prediction in patients with breast cancer.

Keywords: Bioinformatics; Breast cancer; Krüppel‐like factors; Prognosis.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
The transcription levels of KLFs in different types of cancers by Oncomine. Cancer vs. normal: up-regulated (red) or down-regulated (blue)
Fig. 2
Fig. 2
The expression levels of KLFs in breast cancer. A, B The expression of KLFs in breast cancer (T) and normal breast tissues (N) presented by scatter diagram (A) and box plot (B). C The relationship between KLFs expression levels and clinical stages of patients with breast cancer by GEPIA anslysis. D The relationship between KLFs expression levels and different molecular subtypes in breast cancer by UALCAN analysis. *P < 0.05, **P < 0.01, ***P < 0.001
Fig. 3
Fig. 3
The expression of KLFs in breast cancer. A The mRNA expression levels of KLFs determined by RT-PCR. B The protein expression levels of KLFs determined by immunohistochemistry. **P < 0.01, ***P < 0.001
Fig. 4
Fig. 4
Molecular characteristics of KLFs in breast cancer patients. A Gene mutation analysis of KLFs in patients with breast cancer by cBioPortal analysis. B Pearson’s correlation analysis of KLFs. C GeneMANIA analysis of relevant interactive genes of KLFs. D Molecular network for KLFs and most frequently altered neighbor genes by STRING analysis
Fig. 5
Fig. 5
Cross-analysis of the RFS in patients with breast cancer based on KLFs expression levels by using KM plotter. A The association between KLF2 expression and RFS in patients with different KLFs levels analyzed by KM-plotter. B The association between KLF15 expression and RFS in patients with different KLFs levels analyzed by KM-plotter
Fig. 6
Fig. 6
KLF2 and KLF15 inhibits breast cancer proliferation and migration in vitro. A The mRNA expression levels of KLF2 and KLF15 in breast cancer cell lines and normal breast cell line MCF10A determined by RT-qPCR. B The protein expression levels of KLF2 and KLF15 in breast cancer cell lines and normal breast cell line MCF10A determined by western blot. C The expression of KLF2 and KLF15 in KLF2 or KLF15 expressing plasmid transfected MDA-MB-231 cells determined by western blot. D Cell viability of KLF2-overexpressed (a) or KLF15-overexpressed (b) MDA-MB-231 cells, as well as control cells determined by MTT analysis. E Colony formation analysis of KLF2-overexpressed (a) or KLF15-overexpressed (b) MDA-MB-231 cells, as well as control cells. F, Edu analysis of KLF2-overexpressed (a) or KLF15-overexpressed (b) MDA-MB-231 cells, as well as control cells. G Transwell analysis of KLF2-overexpressed (a) or KLF15-overexpressed (b) MDA-MB-231 cells, as well as control cells. H Scratch analysis of KLF2-overexpressed or KLF15-overexpressed MDA-MB-231 cells, as well as control cells. ***P < 0.001
Fig. 7
Fig. 7
KLF2 and KLF15 induce cell apoptosis and cell cycle arrest at G1 phase. A GSEA analysis of KLF2/15 expression levels and relevant signaling pathways. B The cell cycle distribution of KLF2 or KLF15-overexpressed MDA-MB-231 and control cells determined by flow cytometry analysis. C The expression of Cyclin D1, survivin, p16, p21 and p27 determined by western blot. D The cell apoptosis analysis of KLF2 or KLF15-overexpressed MDA-MB-231 and control cells determined by flow cytometry analysis. ***P < 0.001
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