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PLK1 has tumor-suppressive potential in APC-truncated colon cancer cells.

ABSTRACT: The spindle assembly checkpoint (SAC) acts as a molecular safeguard in ensuring faithful chromosome transmission during mitosis, which is regulated by a complex interplay between phosphatases and kinases including PLK1. Adenomatous polyposis coli (APC) germline mutations cause aneuploidy and are responsible for familial adenomatous polyposis (FAP). Here we study the role of PLK1 in colon cancer cells with chromosomal instability promoted by APC truncation (APC-ΔC). The expression of APC-ΔC in colon cells reduces the accumulation of mitotic cells upon PLK1 inhibition, accelerates mitotic exit and increases the survival of cells with enhanced chromosomal abnormalities. The inhibition of PLK1 in mitotic, APC-∆C-expressing cells reduces the kinetochore levels of Aurora B and hampers the recruitment of SAC component suggesting a compromised mitotic checkpoint. Furthermore, Plk1 inhibition (RNAi, pharmacological compounds) promotes the development of adenomatous polyps in two independent Apc ^Min/+ mouse models. High PLK1 expression increases the survival of colon cancer patients expressing a truncated APC significantly.

SUBMITTER: Raab M

PROVIDER: S-EPMC5856809 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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PLK1 has tumor-suppressive potential in APC-truncated colon cancer cells.

Raab Monika M Sanhaji Mourad M Matthess Yves Y Hörlin Albrecht A Lorenz Ioana I Dötsch Christina C Habbe Nils N Waidmann Oliver O Kurunci-Csacsko Elisabeth E Firestein Ron R Becker Sven S Strebhardt Klaus K

Nature communications 20180316 1

The spindle assembly checkpoint (SAC) acts as a molecular safeguard in ensuring faithful chromosome transmission during mitosis, which is regulated by a complex interplay between phosphatases and kinases including PLK1. Adenomatous polyposis coli (APC) germline mutations cause aneuploidy and are responsible for familial adenomatous polyposis (FAP). Here we study the role of PLK1 in colon cancer cells with chromosomal instability promoted by APC truncation (APC-ΔC). The expression of APC-ΔC in co ...[more]

PMID: 29549256

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Project description:BACKGROUND: Several in vitro assays have been used to identify “cancer stem cells” (CSC), including expression of cell surface markers and Hoechst dye efflux properties. However, each of these methods has potential pitfalls that complicate interpretation of the results. Focusing on colon cancers (CC), the CD133 antigen has been proposed as a marker of colon CSC. However, conflicting results have been reported in the literature indicating the need of a systematic analysis of CSC within CC and a complete validation of markers for the isolation of these cells. AIMS: Aim of this study was to confirm that CD133 expression is a valid method for isolating CSC in CC and verify if other antigens can increase the specificity of this marker for isolating CSC in CC. METHODS: CD133+ and CD133- cells were isolated from different human CC lines (CaCo-2, HT29, LOVO, HCT-116) by FACS sorter and the tumor-initiating potential of CD133+ cells was assessed in vitro, by soft-agar colony formation assay, and in vivo, upon transplantation into nude mice. Furthermore, the gene expression profile of CD133+ versus CD133- CaCo-2 cells was compared by the means of microarray analysis. Then, in the effort to identify a common “tumor stem cell” signature for CC, the most relevant transcripts resulting from gene expression profiling on CD133+ cells was assessed by real-time PCR on SP-fraction isolated by FACS sorter from the same CC cell lines. Finally, we deplete CD133 expression in the CaCo-2 cell line by the means of siRNA and verified by Western Blot analysis whether there was a functional correlation between CD133 and the target genes. Moreover, CaCo-2 and HCT116 cells were exposed to sodium butyrate (NaBu) for 72h. Colon cells differentiation was assessed by Alkaline phosphatase activity and expression of CD133 and target genes was tested by western blot. RESULTS: We confirmed that only CD133+ cells have a tumor-initiating potential in vitro and in vivo. Furthermore, microarray analysis of CD133+ versus CD133- CaCo-2 cells revealed a significant overexpression of various transcripts involved in cell proliferation, invasion and stemness in CD133+ cell fraction. Comparison of the transcripts by real-time PCR revealed that the genes of Endothelin-1 (END-1) and NR4A2 are highly expressed in both CD133 + cells and in SP fractions. Finally, when we deplete CD133 expression in Caco-2cells by siRNA, we observed a significant attenuation of END-1 and NR4A2 expression, thus demonstrating that CD133 is involved in the transcriptional regulation of these genes. Interestingly, we also showed that the expression of all three genes was inversely correlated with cell differentiation status as demonstrated by the fact that their expression decreases in a time- and dose-dependent manner after differentiation induced by NaBu. CONCLUSION: Overall, this study confirms the role of CD133antigen as CSC marker and showed for the first time the existence of a functional relationship between CD133, END-1 and NR4A2 expression, hypothesizing that CD133 is involved in the transcriptional regulation of these gene. Microarray analysis was performed on CD133+ and CD133- sorted CACO-2 cells. For both fractions, cells were sorted three independent times. Sample preparation was performed according to Affymetrix recommendations. A total of 6 arrays were hybridized, including 3 CD133+ replicates and 3 CD133- replicates.

Dataset Information

PLK1 has tumor-suppressive potential in APC-truncated colon cancer cells.

Publications

PLK1 has tumor-suppressive potential in APC-truncated colon cancer cells.

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