High-throughput techniques have identified numerous antisense (AS) transcripts and long non-coding RNAs (ncRNAs). However, their significance in cancer biology remains largely unknown. Here, we report an androgen-responsive long ncRNA, CTBP1-AS, located in the AS region of C-terminal binding protein 1 (CTBP1), which is a corepressor for androgen receptor. CTBP1-AS is predominantly localized in the nucleus and its expression is generally upregulated in prostate cancer. CTBP1-AS promotes both hormonedependent and castration-resistant tumour growth. Mechanistically, CTBP1-AS directly represses CTBP1 expression by recruiting the RNA-binding transcriptional repressor PSF together with histone deacetylases. CTBP1-AS also exhibits global androgen-dependent functions by inhibiting tumour-suppressor genes via the PSF-dependent mechanism thus promoting cell cycle progression. Our findings provide new insights into the functions of ncRNAs that directly contribute to prostate cancer progression.

Vitamin K 2 is a critical nutrient required for blood coagulation. It also plays a key role in bone homeostasis and is a clinically effective therapeutic agent for osteoporosis. We previously demonstrated that vitamin K 2 is a transcriptional regulator of bone marker genes in osteoblastic cells and that it may potentiate bone formation by activating the steroid and xenobiotic receptor, SXR. To explore the SXR-mediated vitamin K 2 signaling network in bone homeostasis, we identified genes up-regulated by both vitamin K 2 and the prototypical SXR ligand, rifampicin, in osteoblastic cells using oligonucleotide microarray analysis and quantitative reverse transcription-PCR. Fourteen genes were up-regulated by both ligands. Among these, tsukushi, matrilin-2, and CD14 antigen were shown to be primary SXR target genes. Moreover, collagen accumulation in osteoblastic MG63 cells was enhanced by vitamin K 2 treatment. Gain-and loss-of-function analyses showed that the small leucine-rich proteoglycan, tsukushi, contributes to vitamin K 2 -mediated enhancement of collagen accumulation. Our results suggest a new function for vitamin K 2 in bone formation as a transcriptional regulator of extracellular matrix-related genes, that are involved in the collagen assembly.

Next-generation sequencing experiments have shown that microRNAs (miRNAs) are expressed in many different isoforms (isomiRs), whose biological relevance is often unclear. We found that mature miR-21, the most widely researched miRNA because of its importance in human disease, is produced in two prevalent isomiR forms that differ by 1 nt at their 3′ end, and moreover that the 3′ end of miR-21 is posttranscriptionally adenylated by the noncanonical poly(A) polymerase PAPD5. PAPD5 knockdown caused an increase in the miR-21 expression level, suggesting that PAPD5-mediated adenylation of miR-21 leads to its degradation. Exoribonuclease knockdown experiments followed by small-RNA sequencing suggested that PARN degrades miR-21 in the 3′-to-5′ direction. In accordance with this model, microarray expression profiling demonstrated that PAPD5 knockdown results in a down-regulation of miR-21 target mRNAs. We found that disruption of the miR-21 adenylation and degradation pathway is a general feature in tumors across a wide range of tissues, as evidenced by data from The Cancer Genome Atlas, as well as in the noncancerous proliferative disease psoriasis. We conclude that PAPD5 and PARN mediate degradation of oncogenic miRNA miR-21 through a tailing and trimming process, and that this pathway is disrupted in cancer and other proliferative diseases.nucleotidyl transferase | microRNA processing

The mitochondrial respiratory chain is essential for oxidative phosphorylation and comprises multiple complexes, including cytochrome c oxidase, assembled in macromolecular supercomplexes. Little is known about factors that contribute to supercomplex organization. Here we identify COX7RP as a factor that promotes supercomplex assembly. Cox7rp-knockout mice exhibit decreased muscular activity and heat production failure in the cold due to reduced COX activity. In contrast, COX7RP-transgenic mice exhibit increased exercise performance with increased cytochrome c oxidase activity. Two-dimensional blue native electrophoresis reveals that COX7RP is a key molecule that promotes assembly of the III 2 /IV n supercomplex with complex I. Our study identified COX7RP as a protein that functions in I/III 2 /IV n supercomplex assembly and is required for full activity of mitochondrial respiration.

Androgen receptor (AR) is a critical transcription factor that regulates various target genes and contributes to the pathophysiology of prostate cancer hormone dependently. Here, we identify amyloid precursor protein (APP) as a primary androgen target through chromatin immunoprecipitation (ChIP) combined with genome tiling array analysis (ChIPchip). ChIP-treated DNA were obtained from prostate cancer LNCaP cells with R1881 or vehicle treatment using AR or acetylated histone H3 antibodies. Ligand-dependent AR binding was further enriched by PCR subtraction. Using chromosome 21/22 arrays, we identified APP as one of the androgen-regulated genes with adjacent functional AR binding sites. APP expression is androgen-inducible in LNCaP cells and APP immunoreactivity was correlated with poor prognosis in patients with prostate cancer. Gain-of-function and lossof-function studies revealed that APP promotes the tumor growth of prostate cancer. The present study reveals a novel APP-mediated pathway responsible for the androgendependent growth of prostate cancer. Our findings will indicate that APP could be a potential molecular target for the diagnosis and treatment of prostate cancer. [Cancer Res 2009;69(1):137-42]

Recent advances in cancer biology reveal that microRNAs (miRNAs) are involved in the regulation of cancer-related genes, or they function as tumor suppressors or oncogenes. In prostate cancer, evidence has accumulated for the contribution of the androgen-dependent gene network to tumor growth, although the precise functions of miRNAs in prostate cancer remain to be investigated. Here, we identified androgen-responsive miRNAs by the short RNA sequencing analysis in LNCaP prostate cancer cells. Among 10 miRNAs with known sequences, we have determined that miR148a reduces the expression of cullin-associated and neddylation-dissociated 1 (CAND1), a negative regulator of SKP1-Cullin1-F-box (SCF) ubiquitin ligases, by binding to the 3 0 -untranslated region of CAND1 mRNA. CAND1 knockdown by small interfering RNA promoted the proliferation of LNCaP cells. Our study indicates the potential contribution of miR-148a to the growth of human prostate cancer.

The androgen receptor (AR) is a critical transcriptional factor that contributes to the development and the progression of prostate cancer (PCa) by regulating the transcription of various target genes. Genome-wide screening of androgen target genes provides useful information to understand a global view of AR-mediated gene network in PCa. In this study, we performed 5'-cap analysis of gene expression (CAGE) to determine androgen-regulated transcription start sites (TSSs) and chromatin immunoprecipitation (ChIP) on array (ChIP-chip) analysis to identify AR binding sites (ARBSs) and histone H3 acetylated (AcH3) sites in the human genome. CAGE determined 13 110 distinct, androgen-regulated TSSs (P<0.01), and ChIP-chip analysis identified 2872 androgen-dependent ARBSs (P<1e-5) and 25 945 AcH3 sites (P<1e-4). Both androgen-regulated coding genes and noncoding RNAs, including microRNAs (miRNAs) were determined as androgen target genes. Besides prototypic androgen-regulated TSSs in annotated gene promoter regions, there are many androgen-dependent TSSs that are widely distributed throughout the genome, including those in antisense (AS) direction of RefSeq genes. Several pairs of sense/antisense promoters were newly identified within single RefSeq gene regions. The integration of CAGE and ChIP-chip analyses successfully identified a cluster of androgen-inducible miRNAs, as exemplified by the miR-125b-2 cluster on chromosome 21. Notably, the number of androgen-upregulated genes was larger in LNCaP cells treated with R1881 for 24 h than for 6 h, and the percentage of androgen-upregulated genes accompanied with adjacent ARBSs was also much higher in cells treated with R1881 for 24 h than 6 h. On the basis of the Oncomine database, the majority of androgen-upregulated genes containing adjacent ARBSs and CAGE tag clusters in our study were previously confirmed as androgen target genes in PCa. The integrated high-throughput genome analyses of CAGE and ChIP-chip provide useful information for elucidating the AR-mediated transcriptional network that contributes to the development and progression of PCa.

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