Telomeres are specialized structures at the ends of linear chromosomes. Their maintenance is essential for the unlimited proliferation of cells due to end-replication problem. Progressive telomere shortening in somatic cells can lead to the induction of senescence or apoptosis, thus acting as a barrier to unlimited proliferation. Therefore, maintaining telomere length is a crucial feature of cancers. To elongate their telomeres, most cancer cells reactivate telomerase, an enzyme which is normally only active in stem cells. However, 5-25 % of cancers employ the alternative lengthening of telomeres (ALT) pathway, which is relying on DNA repair and recombination mechanisms. Accordingly, inhibition of telomere length maintenance is considered an important goal in tumor therapy, however treatment of telomerase-positive tumors with telomerase inhibitors could select for ALT cells instead. Therefore, understanding ALT is crucial for (i) the identification of tumors in which it is active, (ii) the development of prognostic markers, and (iii) the targeting of this pathway with novel anti-cancer therapies. This work is conducted in the framework of the BMBF funded CancerTelSys consortium that is decribed at www.CancerTelSys.org.
Work of the group
ALT-positive cells are characterized by the absence of telomerase activity, the presence of ALT-associated PML (promyelocytic leukemia) bodies (APBs), an extreme heterogeneity of telomere repeat length, an elevated level of telomere sister chromatid exchanges (T-SCEs), extrachromosomal telomere repeats (ECTRs), and increased transcription of telomeric-repeat containing RNA (TERRA) (Fig. 1). Several mutations and deregulations involved in the ALT pathway were discovered, however, the detailed mechanism remains elusive. We are particularly interested in a chromatin remodeler called ATRX, which is frequently mutated in ALT cancers. In regards to ATRX, and related mutations, some cancers seem to rely on epigenetic mechanisms in order to promote ALT (Chudasama et al., 2018).Within the CancerTelSys consortium we dissect various aspects of the ALT mechanism.
|Figure 1. Hallmarks of ALT cancers and related assays.|
Automated 3D imaging analysis
Since APBs contain proteins that are known to function in DNA repair and recombination processes it is thought that the telomere elongation takes place in these nuclear structures. Using automated microscopy images, we identified 29 proteins that had an effect on the formation of PML nuclear bodies and therefore play a role in the complex process of alternative telomere lengthening (Osterwald et al., 2015).An automated 3D imaging-based workflow was also applied to tissue samples, where we quantified individual telomere features on tissue sections for a large number of cells (Gunkel et al., 2017)
To provide a reference for studies that dissect TM features, we constructed the TelNet database (Braun et al., 2017). It offers a comprehensive compilation of human and yeast genes linked to telomere maintenance. These genes were annotated in terms of TM mechanism, associated specific functions and orthologous genes, a TM significance score and information from peer-reviewed literature. TelNet can be integrated into the annotation of genes identified from bioinformatics analysis pipelines to determine possible connections with TM.
Bioinformatical analysis of telomere maintenance and network modeling
We integrate a bioinformatical analysis of (epi)genomic data with our automated high-throughput quantification of 3D fluorescence microscopy images. With this we analyzed a panel of pediatric glioblastoma patient samples in respect to the active telomere maintenance mechanisms (TMM) (Deeg et al., 2017). In addition, we established a classification system called "Predicting ALT IN Tumors" (PAINT) to distinguish between the different TMMs. For this we combine specific cytogenetic TMM features, genetic variants and RNA-Seq data of pediatric glioblastoma patient samples. The TMM classification will be applied in a clinical setting for prognostic, predictive and stratifying analysis of tumor samples. Furthermore, we developed a mathematical model to identify the most important regulators of the telomerase in Saccharomyces cerevisiae (Poos et al. 2016). Currently, we apply these models to human cancer data and focus on the regulation of the telomerase reverse transcriptase (TERT) gene in different cancer types.
We showed that AURKB interacts with both telomerase and cenRNA and activates telomerase in trans (Mallm et al., 2015). Thus, in mouse ESCs, telomere maintenance is regulated via expression of cenRNA in a cell-cycle-dependent manner.
Chudasama P, Mughal S, Sanders M, Hübschmann D, Chung I, Deeg KI, Wong SH, Rabe S, Hlevnjak M, Zapatka M, Ernst A, Kleinheinz K, Schlesner M, Sieverling L, Klink B, Schröck E, Hoogenboezem R, Kasper B, Heilig C, Egerer G, Wolf S, von Kalle C, Eils R, Stenzinger A, Weichert W, Glimm H, Gröschel S, Kopp H-G, Omlor G, Lehner B, Bauer S, Schimmack S, Ulrich A, Mechtersheimer G, Rippe K, Brors B, Hutter B, Renner M, Hohenberger P, Scholl C & Fröhling S (2018). Integrative genomic and transcriptomic analysis of leiomyosarcoma. Nat Commun 9, 144. doi: 10.1038/s41467-017-02602-0 | Abstract | Reprint (3.1 MB) | Article metrics
Gunkel M, Chung I, Wörz S, Deeg KI, Simon R, Sauter G, Jones DT, Korshunov A, Rohr K, Erfle H & Rippe K (2017). Quantification of telomere features in tumor tissue sections by an automated 3D imaging-based workflow. Methods 114, 60-73. doi: 10.1016/j.ymeth.2016.09.014 | Abstract | Reprint (7.3 MB) | Article metrics
Poos AM, Maicher A, Dieckmann A, Oswald M, Eils R, Kupiec M, Luke B & König R (2016). Mixed integer linear programming based machine learning approach identifies regulators of telomerase in yeast. Nucleic Acids Res 44, e93. doi: 10.1093/nar/gkw111 | Article metrics
Deeg KI, Chung I, Bauer C & Rippe K (2016). Cancer cells with alternative lengthening of telomeres do not display a general hypersensitivity to ATR inhibition. Front Oncol 6, 186. doi: 10.3389/fonc.2016.00186 | Abstract | Reprint (1.4 MB) | Article metrics.
Mallm JP & Rippe K (2015). Aurora kinase B regulates telomerase activity via a centromeric RNA in stem cells. Cell Rep 11, 1667-1678. doi: 10.1016/j.celrep.2015.05.015 | Abstract | Reprint (5.9 MB) | Article metrics
Osterwald S, Deeg KI, Chung I, Parisotto D, Wörz S, Rohr K, Erfle H & Rippe K (2015). PML induces compaction, partial TRF2 depletion and DNA damage signaling at telomeres and promotes alternative lengthening of telomeres. J Cell Sci 128, 1887-1900. doi: 10.1242/jcs.148296 | Abstract | Reprint (4.3 MB) | Article metrics
Osterwald, S., Wörz, S., Reymann, J., Sieckmann, F., Rohr, K., Erfle, H. & Rippe, K. (2012). A three-dimensional colocalization RNA interference screening platform to elucidate the alternative lengthening of telomeres pathway. Biotechnol. J. 7, 103-116. doi: 10.1002/biot.201000474 | Abstract | Reprint (1.8 MB)
Braun DM, Chung I, Kepper N & Rippe K (2017). TelNet - a database for human and yeast genes involved in telomere maintenance. biorxiv 130153. doi: 10.1101/130153
Deeg KI, Chung I, Poos AM, Braun DM, Korshunov A, Oswald M, Kepper, N, Bender S, Castel D, Lichter P, Grill J, Pfister SM, König R, Jones DT & Rippe K (2017). Dissecting telomere maintenance mechanisms in pediatric glioblastoma. biorxiv 129106. doi: 10.1101/129106
Sieverling L, Chen Hong, Koser SD, Ginsbach P, Kleinheinz K, Hutter B, Braun DM, Cortes-Ciriano I, Xi R, Kabbe R, Park PJ, Eils R, Schlesner M, Rippe K, Jones DTW, Brors B & Feuerbach L (2017). Genomic footprints of activated telomere maintenance mechanisms in cancer. biorxiv 157560. doi: 10.1101/157560