OCT4 controls mitotic stability and inactivates the RB tumor suppressor pathway to enhance ovarian cancer aggressiveness

OCT4 (Octamer-binding transcription factor 4) is essential for embryonic stem cell self-renewal. Here we show that OCT4 increases the aggressiveness of high-grade serous ovarian cancer (HG-SOC) by inactivating the Retinoblastoma tumor suppressor pathway and enhancing mitotic stability in cancer cells. OCT4 drives the expression of Nuclear Inhibitor of Protein Phosphatase type 1 (NIPP1) and Cyclin F (CCNF) that together inhibit Protein Phosphatase 1 (PP1). This results in pRB hyper-phosphorylation, accelerated cell proliferation and increased in vitro tumorigenicity of ovarian cancer cells. In parallel, OCT4 and NIPP1/CCNF drive the expression of the central Chromosomal Passenger Complex (CPC) components, Borealin, Survivin and the mitotic kinase Aurora B, promoting the clustering of supernumerary centrosomes to increase mitotic stability. Loss of OCT4 or NIPP1/CCNF results in severe mitotic defects, multipolar spindles and supernumerary centrosomes, finally leading to the induction of apoptosis. These phenotypes were recapitulated in different cancer models indicating general relevance for human cancer. Importantly, activation of these parallel pathways leads to dramatically reduced overall survival of HG-SOC patients. Altogether, our data highlights an unprecedented role for OCT4 as central regulator of mitotic fidelity and RB tumor suppressor pathway activity. Disrupting this pathway represents a promising strategy to target an aggressive subpopulation of HG-SOC cells.Oncogene advance online publication, 20 March 2017; doi:10.1038/onc.2017.20

TERRA regulate the transcriptional landscape of pluripotent cells through TRF1-dependent recruitment of PRC2

The mechanisms that regulate pluripotency are still largely unknown. Here, we show that Telomere Repeat Binding Factor 1 (TRF1), a component of the shelterin complex, regulates the genome-wide binding of polycomb and polycomb H3K27me3 repressive marks to pluripotency genes, thereby exerting vast epigenetic changes that contribute to the maintenance of mouse ES cells in a na\uefve state. We further show that TRF1 mediates these effects by regulating TERRA, the lncRNAs transcribed from telomeres. We find that TERRAs are enriched at polycomb and stem cell genes in pluripotent cells and that TRF1 abrogation results in increased TERRA levels and in higher TERRA binding to those genes, coincidental with the induction of cell-fate programs and the loss of the na\uefve state. These results are consistent with a model in which TRF1-dependent changes in TERRA levels modulate polycomb recruitment to pluripotency and differentiation genes. These unprecedented findings explain why TRF1 is essential for the induction and maintenance of pluripotency

TGS1 mediates 2,2,7-trimethyl guanosine capping of the human telomerase RNA to direct telomerase dependent telomere maintenance

Pathways that direct the selection of the telomerase-dependent or recombination-based, alternative lengthening of telomere (ALT) maintenance pathway in cancer cells are poorly understood. Using human lung cancer cells and tumor organoids we show that formation of the 2,2,7-trimethylguanosine (TMG) cap structure at the human telomerase RNA 5′ end by the Trimethylguanosine Synthase 1 (TGS1) is central for recruiting telomerase to telomeres and engaging Cajal bodies in telomere maintenance. TGS1 depletion or inhibition by the natural nucleoside sinefungin impairs telomerase recruitment to telomeres leading to Exonuclease 1 mediated generation of telomere 3′ end protrusions that engage in RAD51-dependent, homology directed recombination and the activation of key features of the ALT pathway. This indicates a critical role for 2,2,7-TMG capping of the RNA component of human telomerase (hTR) in enforcing telomerase-dependent telomere maintenance to restrict the formation of telomeric substrates conductive to ALT. Our work introduces a targetable pathway of telomere maintenance that holds relevance for telomere-related diseases such as cancer and aging

A ‘higher order' of telomere regulation: telomere heterochromatin and telomeric RNAs

Protection of chromosome ends from DNA repair and degradation activities is mediated by specialized protein complexes bound to telomere repeats. Recently, it has become apparent that epigenetic regulation of the telomric chromatin template critically impacts on telomere function and telomere-length homeostasis from yeast to man. Across all species, telomeric repeats as well as the adjacent subtelomeric regions carry features of repressive chromatin. Disruption of this silent chromatin environment results in loss of telomere-length control and increased telomere recombination. In turn, progressive telomere loss reduces chromatin compaction at telomeric and subtelomeric domains. The recent discoveries of telomere chromatin regulation during early mammalian development, as well as during nuclear reprogramming, further highlights a central role of telomere chromatin changes in ontogenesis. In addition, telomeres were recently shown to generate long, non-coding RNAs that remain associated to telomeric chromatin and will provide new insights into the regulation of telomere length and telomere chromatin. In this review, we will discuss the epigenetic regulation of telomeres across species, with special emphasis on mammalian telomeres. We will also discuss the links between epigenetic alterations at mammalian telomeres and telomere-associated diseases

Electromechanical analysis of an adaptive piezoelectric energy harvester controlled by two segmented electrodes with shunt circuit networks

This paper presents an adaptive power harvester using a shunted piezoelectric control system with segmented electrodes. This technique has spurred new capability for widening the three simultaneous resonance frequency peaks using only a single piezoelectric laminated beam where normally previous works only provide a single peak for the resonance at the first mode. The benefit of the proposed techniques is that it provides effective and robust broadband power generation for application in self-powered wireless sensor devices. The smart structure beam with proof mass offset is considered to have simultaneous combination between vibration-based power harvesting and shunt circuit control-based electrode segments. As a result, the system spurs new development of the two mathematical methods using electromechanical closed-boundary value techniques and Ritz method-based weak-form analytical approach. The two methods have been used for comparison giving accurate results. For different electrode lengths using certain parametric tuning and harvesting circuit systems, the technique enables the prediction of the power harvesting that can be further proved to identify the performance of the system using the effect of varying circuit parameters so as to visualize the frequency and time waveform responses

TERRA Promotes Telomere Shortening through Exonuclease 1–Mediated Resection of Chromosome Ends

The long noncoding telomeric repeat containing RNA (TERRA) is expressed at chromosome ends. TERRA upregulation upon experimental manipulation or in ICF (immunodeficiency, centromeric instability, facial anomalies) patients correlates with short telomeres. To study the mechanism of telomere length control by TERRA in Saccharomyces cerevisiae, we mapped the transcriptional start site of TERRA at telomere 1L and inserted a doxycycline regulatable promoter upstream. Induction of TERRA transcription led to telomere shortening of 1L but not of other chromosome ends. TERRA interacts with the Exo1-inhibiting Ku70/80 complex, and deletion of EXO1 but not MRE11 fully suppressed the TERRA–mediated short telomere phenotype in presence and absence of telomerase. Thus TERRA transcription facilitates the 5′-3′ nuclease activity of Exo1 at chromosome ends, providing a means to regulate the telomere shortening rate. Thereby, telomere transcription can regulate cellular lifespan through modulation of chromosome end processing activities

Active Chromatin Marks Are Retained on X Chromosomes Lacking Gene or Repeat Silencing Despite XIST/Xist Expression in Somatic Cell Hybrids

X-chromosome inactivation occurs early in mammalian development and results in the inactive X chromosome acquiring numerous hallmarks of heterochromatin. While XIST is a key player in the inactivation process, the method of action of this ncRNA is yet to be determined.To assess which features of heterochromatin may be directly recruited by the expression and localization of the XIST RNA we have analyzed a mouse/human somatic cell hybrid in which expression of human and mouse XIST/Xist has been induced from the active X by demethylation. Such hybrids had previously been demonstrated to disconnect XIST/Xist expression from gene silencing and we confirm maintenance of X-linked gene expression, even close to the Xist locus, despite the localized expression of mouse Xist.Loss of the active chromatin marks H3 acetylation and H3 lysine 4 methylation was not observed upon XIST/Xist expression, nor was there a gain of DNA methylation; thus these marks of facultative heterochromatin are not solely dependent upon Xist expression. Cot-1 holes, regions of depleted RNA hybridization with a Cot-1 probe, were observed upon Xist expression; however, these were at reduced frequency and intensity in these somatic cells. Domains of human Cot-1 transcription were observed corresponding to the human chromosomes in the somatic cell hybrids. The Cot-1 domain of the X was not reduced with the expression of XIST, which fails to localize to the human X chromosome in a mouse somatic cell background. The human inactive X in a mouse/human hybrid cell also shows delocalized XIST expression and an ongoing Cot-1 domain, despite X-linked gene silencing. These results are consistent with recent reports separating Cot-1 silencing from genic silencing, but also demonstrate repetitive element expression from an otherwise silent X chromosome in these hybrid cells

Distinct Differences in Chromatin Structure at Subtelomeric X and Y' Elements in Budding Yeast

In Saccharomyces cerevisiae, all ends of telomeric DNA contain telomeric repeats of (TG1–3), but the number and position of subtelomeric X and Y' repeat elements vary. Using chromatin immunoprecipitation and genome-wide analyses, we here demonstrate that the subtelomeric X and Y' elements have distinct structural and functional properties. Y' elements are transcriptionally active and highly enriched in nucleosomes, whereas X elements are repressed and devoid of nucleosomes. In contrast to X elements, the Y' elements also lack the classical hallmarks of heterochromatin, such as high Sir3 and Rap1 occupancy as well as low levels of histone H4 lysine 16 acetylation. Our analyses suggest that the presence of X and Y' elements govern chromatin structure and transcription activity at individual chromosome ends

5′-UTR RNA G-quadruplexes: translation regulation and targeting

RNA structures in the untranslated regions (UTRs) of mRNAs influence post-transcriptional regulation of gene expression. Much of the knowledge in this area depends on canonical double-stranded RNA elements. There has been considerable recent advancement of our understanding of guanine(G)-rich nucleic acids sequences that form four-stranded structures, called G-quadruplexes. While much of the research has been focused on DNA G-quadruplexes, there has recently been a rapid emergence of interest in RNA G-quadruplexes, particularly in the 5′-UTRs of mRNAs. Collectively, these studies suggest that RNA G-quadruplexes exist in the 5′-UTRs of many genes, including genes of clinical interest, and that such structural elements can influence translation. This review features the progresses in the study of 5′-UTR RNA G-quadruplex-mediated translational control. It covers computational analysis, cell-free, cell-based and chemical biology studies that have sought to elucidate the roles of RNA G-quadruplexes in both cap-dependent and -independent regulation of mRNA translation. We also discuss protein trans-acting factors that have been implicated and the evidence that such RNA motifs have potential as small molecule target. Finally, we close the review with a perspective on the future challenges in the field of 5′-UTR RNA G-quadruplex-mediated translation regulation

Telomerase Efficiently Elongates Highly Transcribing Telomeres in Human Cancer Cells

RNA polymerase II transcribes the physical ends of linear eukaryotic chromosomes into a variety of long non-coding RNA molecules including telomeric repeat-containing RNA (TERRA). Since TERRA discovery, advances have been made in the characterization of TERRA biogenesis and regulation; on the contrary its associated functions remain elusive. Most of the biological roles so far proposed for TERRA are indeed based on in vitro experiments carried out using short TERRA-like RNA oligonucleotides. In particular, it has been suggested that TERRA inhibits telomerase activity. We have exploited two alternative cellular systems to test whether TERRA and/or telomere transcription influence telomerase-mediated telomere elongation in human cancer cells. In cells lacking the two DNA methyltransferases DNMT1 and DNMT3b, TERRA transcription and steady-state levels are greatly increased while telomerase is able to elongate telomeres normally. Similarly, telomerase can efficiently elongate transgenic inducible telomeres whose transcription has been experimentally augmented. Our data challenge the current hypothesis that TERRA functions as a general inhibitor of telomerase and suggest that telomere length homeostasis is maintained independently of TERRA and telomere transcription