The functional roles of SNPs within the 8q24 gene desert in the cancer phenotype are not yet well understood. Here, we report that CCAT2, a novel long noncoding RNA transcript (lncRNA) encompassing the rs6983267 SNP, is highly overexpressed in microsatellite-stable colorectal cancer and promotes tumor growth, metastasis, and chromosomal instability. We demonstrate that MYC, miR-17-5p, and miR-20a are up-regulated by CCAT2 through TCF7L2-mediated transcriptional regulation. We further identify the physical interaction between CCAT2 and TCF7L2 resulting in an enhancement of WNT signaling activity. We show that CCAT2 is itself a WNT downstream target, which suggests the existence of a feedback loop. Finally, we demonstrate that the SNP status affects CCAT2 expression and the risk allele G produces more CCAT2 transcript. Our results support a new mechanism of MYC and WNT regulation by the novel lncRNA CCAT2 in colorectal cancer pathogenesis, and provide an alternative explanation of the SNP-conferred cancer risk.[Supplemental material is available for this article.]Notwithstanding the considerable advancements in our understanding of the molecular genetic basis of cancer, in the majority of cancer-associated genomic regions, the responsible protein-coding genes have not been identified yet. The discovery of short (19-22 nt), noncoding RNAs (ncRNAs)-called microRNAs (miRNAs) (Ambros 2001)-not only revealed a novel mechanism of gene regulation but also led to the identification of miRNAs directly involved in cancer development (Spizzo et al. 2009). It is therefore plausible that as-yet-unidentified members of the broader category of ncRNA mapping to cancer-associated genomic regions play ratelimiting roles in tumor initiation and/or progression (Rinn and Chang 2012). For instance, we previously reported that highly conserved genomic regions (ultraconserved regions, or UCRs) (Bejerano et al. 2004) are frequently transcribed as long (>200 bp) ncRNAs (lncRNAs) in both normal and tumor tissues (Calin et al. 2007). Furthermore, germline mutations, as well as single nucleotide polymorphisms (SNPs) in ultraconserved ncRNAs, were found to occur more frequently in patients with colon cancer and chronic leukemia than in the general population (Wojcik et al. 2010).The rs6983267 SNP, mapping to the 8q24.21 chromosomal region, has been consistently associated with an increased risk of colorectal cancer (CRC) (Haiman et al. 2007): The G allele was associated with greater predisposition to CRC than the T allele (odds ratios of 1.27 and 1.47 for heterozygotes and homozygotes, respectively; P = 1.27 3 10 À14 ) (Tomlinson et al. 2007). The increased cancer risk from this SNP variant was also observed in other cancer types, including prostate, ovarian, and inflammatory breast cancer (Ghoussaini et al. 2008;Bertucci et al. 2012). Despite the consistent association between rs6983267 and cancer risk, the underlying molecular and cellular mechanisms remain largely unknown. The genomic region spanning rs6983267 was found to contain DNA (Pom...

Three SUMO (small ubiquitin-related modifier) genes have been identified in humans, which tag proteins to modulate subcellular localization and/or enhance protein stability and activity. We report the identification of a novel intronless SUMO gene, SUMO-4, that encodes a 95-amino acid protein having an 86% amino acid homology with SUMO-2. In contrast to SUMO-2, which is highly expressed in all of the tissues examined, SUMO-4 mRNA was detected mainly in the kidney. A single nucleotide polymorphism was detected in SUMO-4, substituting a highly conserved methionine with a valine residue (M55V). In HepG2 (liver carcinoma) cells transiently transfected with SUMO-4 expression vectors, Met-55 was associated with the elevated levels of activated heat shock factor transcription factors as compared with Val-55, whereas the levels of NF-B were suppressed to an identical degree. The SUMO-4M (Met) variant is associated with type I diabetes mellitus susceptibility in families (p ‫؍‬ 4.0 ؋ 10 ؊4 ), suggesting that it may be involved in the pathogenesis of type I diabetes. SUMO-11 is involved in the post-translational modification of cellular proteins and regulates various cellular processes such as nuclear transport, oncogenesis, stress response, inflammation, and the response to viral infection (1). SUMO-1 is related to ubiquitin, and although both share only an 18% sequence homology, they have a similar three-dimensional structure (1). In contrast to ubiquitin, which tags proteins for degradation, SUMO-1 seems to enhance protein stability and modulate the subcellular localization. The list of mammalian proteins, which are known to be sumoylated, is growing continually and includes RAN GTPase-activating protein 1 (nuclear import) (2), IB␣ (NF-B inhibition) (3), c-Jun (transcription factor), p53 (tumor suppressor) (4), GLUT1 and GLUT4 (glucose transport) (5), heat shock transcription factors 1 and 2 (HSF1 and 2) (6, 7), and FAS/APO-1 (cell death domain/ apoptosis) (8).SUMO-1 is expressed initially as an inactive precursor of 101 amino acids. A C-terminal proteolytic event exposes two glycine residues at the C terminus that are involved in the formation of a peptide bond with the ⑀-amino group of a lysine residue of a target protein. The activating enzymes E1 (AOS1 and UBA2) and the conjugating enzyme E2 (UBC9) are required for sumoylation in vitro (1). In vivo, an additional enzyme E3 is required for efficient sumoylation (9). The E2 enzyme, UBC9, is specific for sumoylation, and recently, two additional SUMO family members have been identified based on the interaction with UBC9, SUMO-2, and SUMO-3 (10, 11). SUMO-1 shares a 48% identity with SUMO-2 and a 46% with SUMO-3. Very little is known regarding the functions of SUMO-2 and SUMO-3. However, SUMO-2 and SUMO-3 contain a consensus SUMO modification site (Fig. 1), which enables self-sumoylation and the formation of polymeric chains (11), in contrast to SUMO-1. Thus, SUMO-2 and SUMO-3 may have different roles in cellular functions.Here, we identify and characterize a novel fou...

Cancers with dysfunctional mutations in BRCA1 or BRCA2, most commonly associated with some breast cancers, are deficient in the DNA damage repair pathway called homologous recombination (HR), which makes them exquisitely vulnerable to poly(ADP-ribose) polymerase (PARP) inhibitors, such as olaparib. This functional state and therapeutic sensitivity is referred to as “BRCAness”. Pharmaceutical induction of BRCAness could expand the use of PARP inhibitors to other tumor types. For example, BRCA mutations are present in only a small proportion of prostate cancer (PCa) patients. We found that castration-resistant PCa (CRPC) cells increased expression of a set of HR-associated genes, including BRCA1, RAD54L and RMI2. Androgen-targeted therapy is typically not effective in CRPC patients. However, the androgen receptor (AR) inhibitor enzalutamide suppressed the expression of those HR genes, thus creating HR deficiency and BRCAness in CRPC cells. In a manner dependent on these gene expression effects, a “lead-in” treatment strategy, in which enzalutamide was followed by the combination of enzalutamide and olaparib, promoted DNA damage-induced cell death and inhibited clonal proliferation of PCa cells in culture and suppressed the growth of PCa xenografts in mice. Thus, our study suggests that anti-androgen and PARP inhibitor combination therapy may be effective for patients with CRPC, and that pharmaceutically-induced BRCAness may expand the clinical use of PARP inhibitors.

Bone metastases remain as a serious health concern because of limited therapeutic options. Here, we report that crosstalk between ROR1-HER3 and the Hippo-YAP pathway promotes breast cancer bone metastasis in a long noncoding RNA-dependent fashion. Mechanistically, the orphan receptor tyrosine kinase ROR1 phosphorylates HER3 at a previously unidentified site Tyr1307, upon neuregulin stimulation, independently of other ErbB family members. p-HER3 Tyr1307 recruits the LLGL2-MAYA-NSUN6 RNA-protein complex to methylate Hippo/MST1 at Lys59. This methylation leads to MST1 inactivation and activation of YAP target genes in tumor cells, which elicits osteoclast differentiation and bone metastasis. Furthermore, increased ROR1, p-HER3 Tyr1307 and MAYA levels correlate with tumor metastasis and unfavorable outcomes. Our data provide insights into the mechanistic regulation and linkage of the ROR1-HER3 and Hippo-YAP pathway in cancer-specific context, and also imply valuable therapeutic targets for bone metastasis and possible therapy-resistant tumors.

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