Analysis of an alternative human CD133 promoter reveals the implication of Ras/ERK pathway in tumor stem-like hallmarks

Abstract

BackgroundAn increasing number of studies support the presence of stem-like cells in human malignancies. These cells are primarily responsible for tumor initiation and thus considered as a potential target to eradicate tumors. CD133 has been identified as an important cell surface marker to enrich the stem-like population in various human tumors. To reveal the molecular machinery underlying the stem-like features in tumor cells, we analyzed a promoter of CD133 gene using human colon carcinoma Caco-2 and synovial sarcoma Fuji cells, which endogenously express CD133 gene.ResultsA reporter analysis revealed that P5 promoter, located far upstream in a human CD133 gene locus, exhibits the highest activity among the five putative promoters (P1 to P5). Deletion and mutation analysis identified two ETS binding sites in the P5 region as being essential for its promoter activity. Electrophoretic mobility shift assays demonstrated the specific binding between nuclear factors and the ETS binding sequence. Overexpression of dominant-negative forms of Ets2 and Elk1 resulted in the significant decrease of P5 activity. Furthermore, treatment of Fuji cells with a specific MEK/ERK inhibitor, U0126, also markedly decreased CD133 expression, but there was no significant effect in Caco-2 cells, suggesting cell type-specific regulation of CD133 expression. Instead, the side population, another hallmark of TSLCs, was dramatically diminished in Caco-2 cells by U0126. Finally, Ras-mediated oncogenic transformation in normal human astrocytes conferred the stem-like capability to form neurosphere-like colonies with the increase of CD133 mRNA expression.ConclusionsIn conclusion, the Ras/ERK pathway at least in part contributes to the maintenance and the acquisition of stem-like hallmarks, although the extent of its contribution is varied in a cell type-specific manner. These findings could help our comprehensive understanding of tumor stemness, and also improve the development of eradicative therapies against human malignancies.

Mutations in the Ras–Raf Axis Underlie the Prognostic Value of CD133 in Colorectal Cancer

“…Mutation analysis indicated that high CD133 expression was related to mutations in K-Ras or B-Raf. Interestingly, it was previously shown that activation of the Ras-Raf-MEK-ERK pathway can induce the activation of CD133 promoter 5 via its nuclear target Ets, thereby inducing CD133 expression (47). We confirmed this Ras dependency of CD133 mRNA and protein expression either by disrupting the mutant K-Ras allele in CRC cell lines or by inhibiting Ras downstream signaling.…”

BMP4 Administration Induces Differentiation of CD133+ Hepatic Cancer Stem Cells, Blocking Their Contributions to Hepatocellular Carcinoma

“…Our results showed that short-term treatment with BMP4 could induce Erk1/2 activation in a time-and dose-dependent manner. In addition, the inhibition of Erk1/2 phosphorylation by a MEK1/ 2 inhibitor may lead to CD133 protein overexpression in HCC cell lines, which is distinct from the observation of CSCs in colon carcinoma (39). The MEK inhibitor nearly abolished the CD133 expression reduction resulting from the BMP4 treatment, and a short-term exposure to a high level of Erk1/2 phosphorylation plays an important role in BMP4 prodifferentiation effects.…”

Hypoxia induces CD133 expression in human lung cancer cells by up-regulation of OCT3/4 and SOX2

“…moreover, the translational efficiency and stability of CD133 transcripts varied between 5' UTR sequences containing exon 1 (1A, 1B, 1C, 1D, or 1E). The 5' UTR region in CD133 is involved in the regulation of well-known proteins that regulate growth and differentiation of normal stem cells, and in the progression of specific types of cancers, such as leukemia and prostate cancer (23,36,37). Therefore, the role of exon1 in CD133 expression needs to be determined.…”