Understanding normal and cancer stem cells should provide insights into the origin of prostate cancer and their mechanisms of resistance to current treatment strategies. In this study, we isolated and characterized stem-like cells present in the immortalized human prostate cell line, RWPE-1. We used a reporter system with green fluorescent protein (GFP) driven by the promoter of s-SHIP (for stem-SH2-domain-containing 5¢-inositol phosphatase) whose stem cell-specific expression has been previously shown. We observed that s-SHIP-GFPexpressing RWPE-1 cells showed stem cell characteristics such as increased expression of stem cell surface markers (CD44, CD166, TROP2) and pluripotency transcription factors (Oct4, Sox2), and enhanced sphereforming capacity and resistance to arsenite-induced cell death. Concomitant increased expression of the long noncoding RNA H19 was observed, which prompted us to investigate a putative role in stemness for this oncofetal gene. Targeted suppression of H19 with siRNA decreased Oct4 and Sox2 gene expression and colonyforming potential in RWPE-1 cells. Conversely, overexpression of H19 significantly increased gene expression of these two transcription factors and the sphere-forming capacity of RWPE-1 cells. Analysis of H19 expression in various prostate and mammary human cell lines revealed similarities with Sox2 expression, suggesting that a functional relationship may exist between H19 and Sox2. Collectively, we provide the first evidence that s-SHIP-GFP promoter reporter offers a unique marker for the enrichment of human stem-like cell populations and highlight a role in stemness for the long noncoding RNA H19.

Epstein-Barr virus (EBV) is a common human herpesvirus. Infection with EBV is associated with several human malignancies in which the virus expresses a set of latent proteins, among which is latent membrane protein 1 (LMP1). LMP1 is able to transform numerous cell types and is considered the main oncogenic protein of EBV. The mechanism of action is based on mimicry of activated members of the tumor necrosis factor (TNF) receptor superfamily, through the ability of LMP1 to bind similar adapters and to activate signaling pathways. We previously generated two unique models: a monocytic cell line and a lymphocytic (NC5) cell line immortalized by EBV that expresses the type II latency program. Here we generated LMP1 dominant negative forms (DNs), based on fusion between green fluorescent protein (GFP) and transformation effector site 1 (TES1) or TES2 of LMP1. Then we generated cell lines conditionally expressing these DNs. These DNs inhibit NF-B and Akt pathways, resulting in the impairment of survival processes and increased apoptosis in these cell lines. This proapoptotic effect is due to reduced interaction of LMP1 with specific adapters and the recruitment of these adapters to DNs, which enable the generation of an apoptotic complex involving TRADD, FADD, and caspase 8. Similar results were obtained with cell lines displaying a latency III program in which LMP1-DNs decrease cell viability. Finally, we prove that synthetic peptides display similar inhibitory effects in EBV-infected cells. DNs derived from LMP1 could be used to develop therapeutic approaches for malignant diseases associated with EBV. E pstein-Barr virus (EBV) is a human herpesvirus involved in infectious mononucleosis and the development of several human malignancies, such as B and T lymphomas and several carcinomas (35). EBV can infect B cells and transform them into lymphoblastoid cell lines (LCLs) (19). EBV may also infect T cells and monocytes (27) or epithelial cells (35). In transformed cells, EBV is found in a latent state, and several viral genes are expressed. Latent membrane protein 1 (LMP1) is derived from one of these genes and has been shown to be essential for B-cell immortalization and the proliferation of monocytes transformed by EBV (19,27). LMP1 can be regarded as an oncogene product per se, since it can transform rodent fibroblasts (46) and sensitize transgenic mice to lymphomas (22). LMP1 is a 63-kDa plasma membrane protein with six transmembrane domains and can mimic a constitutively activated cell surface receptor that belongs to the tumor necrosis factor receptor (TNFR) superfamily (6). In contrast to the mechanism of action of TNFR1, spontaneous oligomerization mediated by the transmembrane and cytoplasmic N-terminal domains of LMP1 induces the activation of several signaling pathways (26). Several signaling domains have been identified so far in the cytoplasmic C-terminal region of the protein (8,15,16). These domains bind various adapters and induce specific signaling pathways. The critical residues responsible for adapte...