Expression and functional role of a transcribed noncoding RNA with an ultraconserved element in hepatocellular carcinoma

Abstract

Although expression of non-protein-coding RNA (ncRNA) can be altered in human cancers, their functional relevance is unknown. Ultraconserved regions are noncoding genomic segments that are 100% conserved across humans, mice, and rats. Conservation of gene sequences across species may indicate an essential functional role, and therefore we evaluated the expression of ultraconserved RNAs (ucRNA) in hepatocellular cancer (HCC). The global expression of ucRNAs was analyzed with a custom microarray. Expression was verified in cell lines by real-time PCR or in tissues by in situ hybridization using tissue microarrays. Cellular ucRNA expression was modulated with siRNAs, and the effects on global gene expression and growth of human and murine HCC cells were evaluated. Fifty-six ucRNAs were aberrantly expressed in HepG2 cells compared with nonmalignant hepatocytes. Among these ucRNAs, the greatest change was noted for ultraconserved element 338 (uc.338), which was dramatically increased in human HCC compared with noncancerous adjacent tissues. Although uc.338 is partially located within the poly(rC) binding protein 2 (PCBP2) gene, the transcribed ncRNA encoding uc.338 is expressed independently of PCBP2 and was cloned as a 590-bp RNA gene, termed TUC338. Functional gene annotation analysis indicated predominant effects on genes involved in cell growth. These effects were experimentally demonstrated in both human and murine cells. siRNA to TUC338 decreased both anchorage-dependent and anchorage-independent growth of HCC cells. These studies identify a critical role for TUC338 in regulation of transformed cell growth and of transcribed ultraconserved ncRNA as a unique class of genes involved in the pathobiology of HCC.liver cancer | hepatocarcinoma | exaptation | transposon

CAGE profiling of ncRNAs in hepatocellular carcinoma reveals widespread activation of retroviral LTR promoters in virus-induced tumors

“…A growing number of ncRNAs are being considered as regulators of various cellular processes (Mercer and Mattick 2013), whereas relatively few ncRNAs are reported to be associated with hepatocarcinogenesis (Braconi et al 2011;Spizzo et al 2012). We have established a comprehensive map of promoters for ncRNAs actively transcribed in a large set of human HCCs as well as in the Mdr2 KO mouse model of HCC using CAGE.…”

Long Non-coding RNAs are Differentially Expressed in Hepatocellular Carcinoma Cell Lines with Differing Metastatic Potential

“…According to another study, levels of the lncRNA uc.338 are elevated in HCC cells and may be a promising marker for this type of cancer. Aberrant expression of its transcript, TUC338, is found in transformed hepatocytes, and its functional role in modulating growth may make it a desirable therapeutic target for selected HCC cases (Braconi et al, 2011). Collectively, these studies lead us to propose that lncRNAs may serve as key regulatory hubs in HCC progression.…”

TUC338 promotes cell migration and invasion by targeting TIMP1 in cervical cancer

“…The siRNA sequences (5'-3') were as follows: siRNA-1, UGA CAG CCC UGG AGA CUG A; siRNA-2, CCA CAG GAC AGG UAC AGC A (8). HeLa and C33A cells were transfected with 60 nM TUC338 siRNA (8) or 0 nM siRNA as the negative control (NC) by Lipofectamine ® 2000 (Invitrogen Life Technologies; Thermo Fisher Scientific, Inc.) for 48 h prior to further experiments, according to the manufacturer's instructions. The transfected cells were incubated at 37˚C with 5% CO 2 .…”

“…Braconi et al (8) demonstrated that TUC338 is markedly overexpressed in hepatocellular cancer (HCC) cell lines and tissue, and it promotes tumor cellular proliferation and modulates progression through regulation of the cell cycle. However, the expression level and biological role of TUC338 in CC remains unclear.…”