ABSTRACT
Genes associated with induced pluripotent stem cells (iPS genes) are several pivotal transcriptional factors, which are used to induce pluripotent stem cells from some adult somatic cells. The roles of these iPS genes and especially the signature for these iPS genes in colorectal cancer (CRC) are still unclear. Overexpressed Oct4 and Lin28 but down-regulated Nanog were found in tumor tissues compared with that in their paired normal counterparts of CRC patients. Interestingly, we found that Oct4, Lin28 and Nanog were highly overexpressed in some patients. And the signature for iPS genes was correlated with tumor site (P = 0.012), lymph node status (P = 0.033), Dukes classification (P = 0.033) of CRC patients. Moreover, an independent public expression profiling data showed signature for the four iPS genes could successfully be used to predict the survival of CRC patients with Dukes stages B and C. Immunofluorescent staining of fresh CRC tissues from patients showed that strong co-expressions of Oct4 and Nanog proteins or Sox2 and Lin28 were present in some CRC cells. Then, CRC cell subclone with four iPS genes overexpression were establish by a mixed retroviral system. We found that iPS genes promote sphere-formation, proliferation, colony formation, migration of human CRC cells in vitro and tumor growth in vivo. Our study first shows the clinical significance of iPS signature in CRC patients.
Subject(s)
Colorectal Neoplasms/genetics , Homeodomain Proteins/genetics , Octamer Transcription Factor-3/genetics , RNA-Binding Proteins/genetics , Transcriptome , Blotting, Western , Colorectal Neoplasms/mortality , Colorectal Neoplasms/pathology , Disease-Free Survival , Female , Fluorescent Antibody Technique , Humans , Immunohistochemistry , Male , Middle Aged , Nanog Homeobox Protein , Neoplasm Staging , Pluripotent Stem Cells , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
The regulators of a key metastasis gene PRL-3 in colorectal cancer (CRC) are still largely unknown. We found three potential binding sites of Snail, a key transcriptional factor involved in the epithelial-mesenchymal transition (EMT), in the region of PRL-3 promoter (located at -642 to -383). Moreover, our results showed that one of the Snail binding sites (located at -624 to -619) was the key element to maintain promoter activity of human PRL-3 gene. The transcriptional activity of PRL-3 promoter was abolished after the Snail binding site (located at -624 to -619) was mutated. Both promoter activity and protein expression of PRL-3 in CRC cell lines could be regulated by Snail. In clinical samples of CRC and metastatic lymph node of CRC, expression of PRL-3 protein was correlated with expression of Snail protein. Functional studies using gene over-expression and knockdown methods indicated that Snail promoted proliferation, cell adhesion and migration of human CRC cells. In SW480 cells with PRL-3 stable knockdown, cell proliferation increased after Snail was up-regulated. Our data first reveal transcriptional factor Snail as a key regulator of PRL-3 in CRC. The link between Snail and PRL-3 suggests a new potential mechanism of Snail contributing to progression and metastasis of CRC.
Subject(s)
Colorectal Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Neoplasm Proteins/genetics , Protein Tyrosine Phosphatases/genetics , Transcription Factors/metabolism , Binding Sites , Cell Adhesion/genetics , Cell Growth Processes/genetics , Cell Line, Tumor , Cell Movement/genetics , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Female , Gene Knockdown Techniques , Humans , Male , Neoplasm Proteins/metabolism , Promoter Regions, Genetic , Protein Tyrosine Phosphatases/metabolism , Regulatory Elements, Transcriptional , Snail Family Transcription Factors , Transcription, Genetic , Zinc FingersABSTRACT
To understand the molecular mechanisms of metastasis and prognosis of colorectal cancer (CRC), we isolated single cell-derived progenies (SCPs) from SW480 cells in vitro and compared their metastatic potential in an orthotopic CRC tumour model in vivo. Two groups of SCPs with the capability of high and low metastasis, respectively, were obtained. By analysing the gene expression profiles of high (SCP51), low (SCP58) metastatic SCPs, and their parental cell line (SW480/EGFP), we demonstrated that 143 genes were differentially expressed either between SCP51 and SCP58 or between SCP58 and SW480/EGFP. Gene-annotation enrichment analysis of DAVID revealed 80 genes in the top ten clusters of the analysis (gene enrichment score > 1). Of the 80-gene set, 32 genes are potentially involved in metastasis, as revealed by Geneclip. Five putative metastatic genes (LYN, SDCBP, MAP4K4, DKK1, and MID1) were selected for further validations. Immunohistochemical analysis in a cohort of 181 CRC clinical samples showed that the individual expression of LYN, MAP4K4, and MID1, as well as the five-gene signature, was closely correlated with lymph node metastasis in CRC patients. More importantly, the individual expression of LYN, MAP4K4, SDCBP, and MID1, as well as the five-gene signature, was significantly correlated with overall survival in CRC patients. Thus, our five-gene signature may be able to predict metastasis and survival of CRC in the clinic, and opens new perspectives on the biology of CRC.
Subject(s)
Colorectal Neoplasms/genetics , Neoplasm Metastasis/genetics , Animals , Cell Proliferation , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Female , Gene Expression Profiling/methods , Humans , Male , Mice , Mice, Nude , Neoplasm Metastasis/pathology , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Neoplasm Transplantation , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/physiology , Oligonucleotide Array Sequence Analysis/methods , Prognosis , Reverse Transcriptase Polymerase Chain Reaction/methods , Survival Analysis , Tumor Cells, CulturedABSTRACT
We studied the expression and regulatory effects of ESC self-renewal molecule Nanog in colorectal cancer (CRC). Immunohistochemical analysis of 175 colorectal tumor samples showed that overexpression of Nanog was strongly correlated with poor prognosis, lymph node metastasis and Dukes classification for CRC. Univariate and multivariate survival analyses further indicated that Nanog expression was a potential prognostic factor for CRC. Gain-of-function analysis revealed that lentivirus-mediated Nanog overexpression promoted proliferation, motility and migration of human CRC cells. Interestingly, we found that Nanog played as both an inducer and a receipt of epithelial-mesenchymal transition (EMT) related signals. Nanog induced expression of Slug and Snail, two major regulator of EMT. Meanwhile, Nanog could also be regulated by Snail and initiated by TGF-ß1. Our data demonstrate self-renewal gene Nanog has a prognostic role in CRC, which functions in progression of CRC by promoting proliferation, invasion, and motility of human CRC cells, and participates EMT process during CRC progression.
