The Regulation And Function Of KLF4in Tumorigenesis And Development

The SI00protein family belongs to the largest subgroup of calcium binding EF-hand type proteins. These proteins have been reported to be involved in a range of biological functions that is related both to normal development and tumorigenesis. S100A14is a distinct member of the family involved in several functional and pathological processes. Our precious studies showed that the expression of S100A14was downregulated in esophageal squamous cell carcinomas (ESCC). S100A14induces cell cycle arrest, apoptosis or metastasis in ESCC, and regulates these processes in a receptor for advanced glycation end-products (RAGE)-dependent manner. On the contrary, we also found that S100A14promoted breast cancer cell invasion in a p53-dependent manner. And S100A14plays a critical role in HER2-induced cell growth. These results suggest that the expression and functional role of S100A14in malignant tumors is organ specific. However, the context-specific transcriptional regulation of S100A14varying in different tumors has not been fully understood. In the present study, we sought to dissect the mechanism and function of S100A14in breast cancer.First, we found that S100A14expression is significantly increased in metastatic breast cancer. Kaplan-Meier survival analyses in combined four data sets demonstrated that the expression of S100A14exquisitely and inversely correlated with metastasis-free survival.Next, RTCA (real-time cell analyzer) and wound-healing assay indicated that overexpression of S100A14promoted the migration of breast cancer cells. Furthermore, we found that upregulation of endogenous expression of S100A14could promote the metastatic behaviors. RT-qPCR and Western blotting indicated that TPA substantially invigorated S100A14mRNA and protein levels. However, we found that this regulation is indirect, implicating that other regulators are involved in regulating S100A14expression. With computer-aided transcription factor-binding site analysis, we identified two highly conserved CACCC motifs located at-511bp to+6bp upstream of the S100A14ATG codon. As known, the CACCC motif was generally regarded as the transcription factor KLF4functional binding elements. KLF4is a zinc finger protein of the kruppel-like factor family, which is a member of zinc finger transcription factors defined by containing a composition of three Cys2/His2zinc fingers. And then we employed three strategies to confirm S100A14as a downstream target of KLF4. We observed that the expression of S100A14was clearly upregulated with the overexpression of KLF4and downregulated in the absence of KLF4. Consistent with these results, ChIP and luciferase reporter assays indicate that KLF4can specifically bind to the CACCC motif in the promoter region of S100A14, which is required for KLF4to transactivate S100A14.Finally, knockdown of KLF4substantially attenuated the effect of TPA on the modulation of S100A14, and impaired TPA-mediated migration. Analyzing gene expression profiling and clinical outcome data from published studies elicited that the expression level of S100A14in human breast cancer is inversely correlated with metastasis-free survival and significantly associated with KLF4in both breast cancer cell lines and clinical breast cancer samples. Furthermore, by using the existing GEO database we show that high concomitant KLF4/S100A14is significantly associated with worse metastasis free-survival for patients of breast cancer.In summary, our results demonstrated that TPA modulates the expression level of S100A14via enhancing KLF4expression and increasing its binding directly to the two consensus CACCC motifs within the S100A14promoter, thereby resulting in the elevation of cell motility in breast cancer. Additionally, our results facilitate the development of innovative anticancer strategies and provide considerable insight into understanding the underlying molecular mechanisms by which KLF4and S100A14are involved in breast cancer cell biology. Peroxisome proliferator activated receptor y (PPARy), a subgroup of ligand-activated nuclear receptors, plays critical roles in cell cycle regulation, differentiation, apoptosis and invasion. PPARy is involved in tumorigenesis and is a potent target for cancer therapy.Previous studies showed that some PPARy agonists induce KLF4expression in a receptor-dependent manner in colorectal cancer cells. The Kruppel-like family of zinc-finger transcription factors (KLFs) regulates numerous biological processes including proliferation, differentiation, apoptosis, development, and inflammation. KLF4activation upregulates its target genes P21、GPA33、NAG-1,and inhibits the expression of CyclinDl, which inhibits cellular proliferation, promotes differentiation, induces apoptosis and inhibits angiogenesis. However, up to now, the mechanism of how PPARy agonists active the expression of KLF4is still unknown. This study provides further insights into PPARy signal transduction pathway as well as a novel cancer therapeutic strategy.We demonstrated that KLF4is a direct transcriptional target of PPARy. Computer-aided transcription factor binding site analysis identified one consensus PPAR response element (PPRE) at-1657bp to-1669bp upstream of the KLF4ATG codon. Further analysis by ChIP, EMSA and luciferase reporter assay revealed that PPARy can specifically bind to the PPRE in promoter region of KLF4, which was required for PPARy to transactivate KLF4. We found that stable silencing of KLF4obviously suppresses the G1/S arrest and antiproliferation effects induced by PPARy ligands. Analyzing gene expression profiling and clinical outcome data from published studies elicited that the expression level of KLF4is significantly associated with PPARy in both NCI-60cancer cell line panel and clinical colonrectal cancer samples. Furthermore, by using the existing GEO database we show that high concomitant PPARy and KLF4is significantly associated with worse survival for patients of colonrectal cancer.In conclusion, our data indicated that upregulating KLF4upon PPARy activation is mediated through PPRE in KLF4promoter, thus providing further insights into PPARy signal transduction pathway as well as a novel cancer therapeutic strategy.