Genes Involved in Differentiation, Stem Cell Renewal, and Tumorigenesis Are Modulated in Telomerase-Immortalized Human Urothelial Cells

Abstract

The expression of hTERT, the catalytic subunit of telomerase, immortalizes normal human urothelial cells (NHUC). Expression of a modified hTERT, without the ability to act in telomere maintenance, did not immortalize NHUC, confirming that effects at telomeres are required for urothelial immortalization. Previous studies indicate that inhibition of telomerase has an immediate effect on urothelial carcinoma (UC) cell line viability, before sufficient divisions to account for telomere attrition, implicating non -telomere effects of telomerase in UC. We analyzed the effects of telomerase on gene expression in isogenic mortal and hTERT-transduced NHUC. hTERT expression led to consistent alterations in the expression of genes predicted to be of phenotypic significance in tumorigenesis. A subset of expression changes were detected soon after transduction with hTERT and persisted with continued culture. These genes (NME5, PSCA, TSPYL5, LY75, IGFBP2, IGF2, CEACAM6, XG, NOX5, KAL1, and HPGD) include eight previously identified as polycomb group targets. TERT-NHUC showed overexpression of the polycomb repressor complex (PRC1 and PRC4) components, BMI1 and SIRT1, and down-regulation of multiple PRC targets and genes associated with differentiation. TERT-NHUC at 100 population doublings, but not soon after transduction, showed increased saturation density and an attenuated differentiation response, indicating that these are not acute effects of telomerase expression. Some of the changes in gene expression identified may contribute to tumorigenesis. Expression of NME5 and NDN was down-regulated in UC cell lines and tumors. Our data supports the concept of both telomere-based and non -telomere effects of telomerase and provides further rationale for the use of telomerase inhibitors in UC.

Immortalisation of Normal Human Urothelial Cells Compromises Differentiation Capacity

“…Although the precise underlying mechanisms for this remain unclear, hTERT overexpression can progressively attenuate the molecular differentiation of NHU cells (shown in this study and reported elsewhere [31]) and diminish their capacity to form a functional urothelial barrier. A mechanism for hTERT-mediated suppression of differentiation would be in keeping with the stem cell maintenance function of hTERT, whose activity is known to extend beyond telomerase activation and includes chromatin reorganisation, indicating a potential role in epigenetic regulation of gene transcription [32].…”

Clinical Implications of Gene Dosage and Gene Expression Patterns in Diploid Breast Carcinoma

“…Interestingly, our data suggest the consistent deregulation of 12 genes, i.e., UBE2C, S100A8, CBX2, LOC389033, STC2, DNALI1, SCUBE2, NME5, SUSD3, SERPINA11, AZGP1, and PIP, in breast carcinomas with a malignant phenotype. Functional studies have implicated several of these genes in a role in tumor cell growth, motility, and progression in multiple cancer types (37)(38)(39)(40)(41)(42)(43)(44). Here, we could associate all 12 genes with progesterone expression and 11 with estrogen expression (UBE2C Response to estradiol stimulus, regulation of cytokine biosynthetic process, regulation of inflammatory response, DNA replication checkpoint, positive regulation of interleukin-4 and 8 production, positive regulation of exit from mitosis, anti-apoptosis, cell-cell signaling, mammary gland development, regulation of cell proliferation, negative regulation of tumor necrosis factor production excluded).…”

Integrated genomic and transcriptional analysis of the in vitro evolution of telomerase‐immortalized urothelial cells (TERT‐NHUC)

“…At the four analysis time-points, microarray expression and Ingenuity pathway analyses, www.ingenuity.com, (Ingenuity Systems V R , Redwood City, California) were performed as described (Chapman et al, 2008). aCGH was performed as described previously (Hurst et al, 2004).…”