Regulation and function of R-loops at repetitive elements

R-loops are atypical, three-stranded nucleic acid structures that contain a stretch of RNA:DNA hybrids and an unpaired, single stranded DNA loop. R-loops are physiological relevant and can act as regulators of gene expression, chromatin structure, DNA damage repair and DNA replication. However, unscheduled and persistent R-loops are mutagenic and can mediate replication-transcription conflicts, leading to DNA damage and genome instability if left unchecked. Detailed transcriptome analysis unveiled that 85% of the human genome, including repetitive regions, hold transcriptional activity. This anticipates that R-loops management plays a central role for the regulation and integrity of genomes. This function is expected to have a particular relevance for repetitive sequences that make up to 75% of the human genome. Here, we review the impact of R-loops on the function and stability of repetitive regions such as centromeres, telomeres, rDNA arrays, transposable elements and triplet repeat expansions and discuss their relevance for associated pathological conditions

Altered telomere homeostasis and resistance to skin carcinogenesis in Suv39h1 transgenic mice

The Suv39h1 and Suv39h2 H3K9 histone methyltransferases (HMTs) have a conserved role in the formation of constitutive heterochromatin and gene silencing. Using a transgenic mouse model system we demonstrate that elevated expression of Suv39h1 increases global H3K9me3 levels in vivo. More specifically, Suv39h1 overexpression enhances the imposition of H3K9me3 levels at constitutive heterochromatin at telomeric and major satellite repeats in primary mouse embryonic fibroblasts. Chromatin compaction is paralleled by telomere shortening, indicating that telomere length is controlled by H3K9me3 density at telomeres. We further show that increased Suv39h1 levels result in an impaired clonogenic potential of transgenic epidermal stem cells and Ras/E1A transduced transgenic primary mouse embryonic fibroblasts. Importantly, Suv39h1 overexpression in mice confers resistance to a DMBA/TPA induced skin carcinogenesis protocol that is characterized by the accumulation of activating H-ras mutations. Our results provide genetic evidence that Suv39h1 controls telomere homeostasis and mediates resistance to oncogenic stress in vivo. This identifies Suv39h1 as an interesting target to improve oncogene induced senescence in premalignant lesions

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

Nuove strategie di terapie antitumorali di precisione: il progetto Interreg – PreCanMed

PreCanMed è un progetto strategico finanziato dall’U.E., Fondo Europeo Regionale e Interreg V-A Italia-Austria 2014-2020. Lo scopo è quello di implementare la tecnologia degli organoidi tumorali da paziente, un nuovo potente strumento d’indagine che permette di studiare i tessuti sani e malati di pazienti con neoplasie, sviluppando trattamenti innovativi e personalizzati.Laureato in scienze della vita all'Università di Vienna, Stefan Schoeftner ha conseguito il dottorato di ricerca presso l'Università di Vienna, nel 2005. Gli è stata quindi assegnata una borsa di studio EMBO a lungo termine per svolgere la sua ricerca post-dottorato nel laboratorio di Maria A. Blasco allo Spanish National Cancer Center (CNIO) di Madrid dal 2006 al 2009. Si è successivamente trasferito a Trieste per svolgere ricerche post-dottorato nel laboratorio di Roberta Benetti presso il Laboratorio Nazionale CIB (LNCIB). Nel marzo 2013 ha trasferito il suo gruppo di ricerca al Laboratorio Nazionale CIB presso l'AREA Science Park di Trieste dove lavora sul ruolo degli RNA non codificanti nella regolazione dei telomeri nel cancro, nell'invecchiamento e nella biologia delle cellule staminali. Dal febbraio 2015 è professore associato di biologia molecolare presso il Dipartimento di scienze della vita dell'Università di Trieste

Novel diagnostic and therapeutic agent

The invention relates to a diagnostic comprising a polynucleotide and to therapeutic agent comprising a polyribonucleotide capable of inhibiting and/or reducing the activity of a telomerase for use in medicine. The invention further provides a method of diagnosing a proliferative disorder. Also provided are therapeutic agent and methods of using the sam

microRNAs control the function of telomeres in cancer

Telomeres are located at the end of chromosomes and consist of DNA tandem repeats that recruit the specialized protein complex “shelterin”. Shelterin has a crucial role in controlling chromosome end protection, telomere recombination and telomere length. Telomeres shorten with every cell division, finally leading to telomere-dysfunction and the induction of senescence or apoptosis. Cancer formation is paralleled by a change in telomere regulation. Re-activation of telomerase ensures the maintenance of telomere function to facilitate unlimited proliferative potential. Importantly, aberrant function of the shelterin complex contributes to tumor formation and genomic instability in human cancer. miRNAs are important regulators of central cancer pathways and are of high clinical relevance. We recently showed that the onco-microRNA miR-155 controls the expression of TRF1 in human breast cancer to promote increased telomere fragility and genomic instability. Importantly, low TRF1 expression correlates with poor prognosis in estrogen receptor positive cancer patients, underlining the clinical relevance of miR-155 dependent regulation of TRF1 in human breast cancer. Our work suggests that multiple miRNAs exist that control telomere function in telomere related pathologies. Identification and functional validation of “telo-miRNAs” is expected to open new avenues in the understanding of telomere related maladies such as cancer and aging

Brevetto Internazionale WO2021/240340A1: Titolo: "Pharmacological composition for the chemical inhibition of TGS1 in the therapeutic treatment of telomeropathies" Inventori: Grazia Daniela Raffa, Stefano Cacchione, Stefan Schoeftner (Università La Sapienza, Roma; Università degli Studi di Trieste)

The Patent describes a function of the RNA methyltransferase TGS1 in modulating the telomere maintenance in human call

miR-335 Directly Targets Rb1 (pRb/p105) in a Proximal Connection to p53-Dependent Stress Response

Loss-of-function mutations of retinoblastoma family (Rb) proteins drive tumorigenesis by overcoming barriers to cellular proliferation. Consequently, factors modulating Rb function are of great clinical import. Here, we show that miR-335 is differentially expressed in human cancer cells and that it tightly regulates the expression of Rb1 (pRb/p105) by specifically targeting a conserved sequence motif in its 3' untranslated region. We found that by altering Rb1 (pRb/p105) levels, miR-335 activates the p53 tumor suppressor pathway to limit cell proliferation and neoplastic cell transformation. DNA damage elicited an increase in miR-335 expression in a p53-dependent manner. miR-335 and p53 cooperated in a positive feedback loop to drive cell cycle arrest. Together, these results indicate that miR-335 helps control proliferation by balancing the activities of the Rb and p53 tumor suppressor pathways. Further, they establish that miR-335 activation plays an important role in the induction of p53-dependent cell cycle arrest after DNA damage

An Oct4-pRB axis, controlled by MiR-335, integrates stem cell self-renewal and cell cycle control

The pluripotency of mouse embryonic stem cells (mESCs) is controlled by a network of transcription factors, mi-RNAs, and signaling pathways. Here, we present a new regulatory circuit that connects miR-335, Oct4, and the Retinoblastoma pathway to control mESC self-renewal and differentiation. Oct4 drives the expression of Nipp1 and Ccnf that inhibit the activity of the protein phosphatase 1 (PP1) complex to establish hyperphosphorylation of the retinoblastoma protein 1 (pRb) as a hallmark feature of self-renewing mESCs. The Oct4-Nipp1/Ccnf-PP1-pRb axis promoting mESC self-renewal is under control of miR-335 that regulates Oct4 and Rb expression. During mESC differentiation, miR-335 upregulation co-operates with the transcriptional repression of Oct4 to facilitate the collapse of the Oct4-Nipp1/Ccnf-PP1-pRb axis, pRb dephosphorylation, the exit from self-renewal, and the establishment of a pRb-regulated cell cycle program. Our results introduce Oct4-dependent control of the Rb pathway as novel regulatory circuit controlling mESC self-renewal and differentiation