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Byline: Prashant. Sharma, Sridhar. Panaiyadiyan, Santosh. Kurra, Raman. Kumar, Brusabhanu. Nayak, Seema. Kaushal, Alpana. Sharma, Rajeev. Kumar, Amlesh. Seth, Prabhjot. Singh We appreciate the author's interest in our recently published [...]

Background: Colorectal cancer is one of the most common cancers and the third leading cause of cancer death in both sexes. The disease progresses as a multistep process and is associated with genetic alterations. One of the characteristic features of cancer is telomerase activation. We sought to evaluate the differences in telomerase activity between colon cancer and adjacent normal tissue and to correlate the differences in telomerase activity between different locations with clinicopathological factors and survival. Methods: Matched colon tumour samples and adjacent normal mucosa samples 10 cm away from the tumour were collected during colectomy. We assessed telomerase activity using real time polymerase chain reaction. Several pathological characteristics of tumours, including p53, Ki-67, p21, bcl2 and MLH1 expression were also studied. Results: We collected samples from 49 patients. There was a significantly higher telomerase activity in colon cancer tissue than normal tissue. Adenocarcinomas of the right colon express significantly higher telomerase than left-side cancers. Colon cancers and their adjacent normal tissue had significantly more telomerase and were more positive to MLH1 than rectal cancers. The expression of p53 negatively correlated to telomerase activity and was linked to better patient survival. Conclusion: Colon and rectal cancers seem to have different telomerase and MLH1 profiles, and this could be another factor for their different biologic and clinical behaviour and progression. These results support the idea that the large bowel cannot be considered a uniform organ, at least in the biology of cancer. Contexte: Le cancer colorectal est l'un des cancers les plus repandus; il arrive au troisieme rang des deces attribuables au cancer chez les hommes et les femmes. La maladie progresse en plusieurs etapes et est associee a des anomalies genetiques. L'une des principales caracteristiques du cancer est l'activation de la telomerase. Nous avons voulu evaluer les differences d'activite de la telomerase entre les tissus touches par le cancer du colon et les tissus adjacents normaux afin d'etablir une correlation entre les differences d'activite de la telomerase selon la localisation d'une part et les facteurs clinicopathologiques et la survie d'autre part. Methodes: Lors de colectomies, nous avons recueilli des echantillons tissulaires jumeles de tumeur du colon et de muqueuse adjacente normale a 10 cm du foyer tumoral. Nous avons mesure l'activite de la telomerase a l'aide de la reaction en chaine de la polymerase en temps reel. Plusieurs caracteristiques pathologiques des tumeurs ont aussi ete analysees, y compris l'expression des genes p53, Ki-67, p21, bcl2 et MLH1. Resultats: Nous avons recueilli des echantillons aupres de 49 patients. Nous avons note une activite nettement plus intense de la telomerase dans les tissus touches par le cancer du colon que dans les tissus normaux. Les adenocarcinomes du colon ascendant expriment une activite de la telomerase significativement plus intense que les cancers du colon descendant. Les cancers du colon et les tissus adjacents normaux presentaient une activite significativement plus intense de la telomerase et etaient plus souvent positifs a l'egard du MLH1 comparativement aux cancers rectaux. L'expression du p53 a ete en correlation negative avec l'activite de la telomerase et a ete associee a une meilleure survie chez les patients. Conclusion: Les cancers du colon et du rectum semblent avoir des profils differents au plan de la telomerase et du gene MLH1. Ce facteur pourrait entre autre expliquer leur comportement et leur progression biologiques et cliniques distincts. Ces resultats appuient l'hypothese selon laquelle le colon ne peut etre considere comme un organe uniforme, du moins en ce qui concerne la biologie du cancer., Cancers of the colon and rectum have been classically viewed as the same disease entity, namely colorectal cancer (CRC), one of the most common cancers in humans and the third [...]

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.cell.2009.06.021 Byline: Agnel Sfeir (1), Settapong T. Kosiyatrakul (2), Dirk Hockemeyer (1), Sheila L. MacRae (1), Jan Karlseder (1), Carl L. Schildkraut (2), Titia de Lange (1) Keywords: DNA Abstract: Telomeres protect chromosome ends through the interaction of telomeric repeats with shelterin, a protein complex that represses DNA damage signaling and DNA repair reactions. The telomeric repeats are maintained by telomerase, which solves the end replication problem. We report that the TTAGGG repeat arrays of mammalian telomeres pose a challenge to the DNA replication machinery, giving rise to replication-dependent defects that resemble those of aphidicolin-induced common fragile sites. Gene deletion experiments showed that efficient duplication of telomeres requires the shelterin component TRF1. Without TRF1, telomeres activate the ATR kinase in S phase and show a fragile-site phenotype in metaphase. Single-molecule analysis of replicating telomeres showed that TRF1 promotes efficient replication of TTAGGG repeats and prevents fork stalling. Two helicases implicated in the removal of G4 DNA structures, BLM and RTEL1, were required to repress the fragile-telomere phenotype. These results identify a second telomere replication problem that is solved by the shelterin component TRF1. Author Affiliation: (1) Laboratory for Cell Biology and Genetics, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA (2) Department of Cell Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA Article History: Received 25 February 2009; Revised 19 May 2009; Accepted 11 June 2009 Article Note: (miscellaneous) Published: July 9, 2009

In Saccharomyces cerevisiae, Cdc13 binds telomeric DNA to recruit telomerase and to 'cap' chromosome ends. In temperature-sensitive cdc13-1 mutants telomeric DNA is degraded and cell-cycle progression is inhibited. To identify novel proteins and pathways that cap telomeres, or that respond to uncapped telomeres, we combined cdc13-1 with the yeast gene deletion collection and used high-throughput spot-test assays to measure growth. We identified 369 gene deletions, in eight different phenotypic classes, that reproducibly demonstrated subtle genetic interactions with the cdc13-1 mutation. As expected, we identified DNA damage checkpoint, nonsense-mediated decay and telomerase components in our screen. However, we also identified genes affecting casein kinase II activity, cell polarity, mRNA degradation, mitochondrial function, phosphate transport, iron transport, protein degradation, and other functions. We also identified a number of genes of previously unknown function that we term RTC, for restriction of telomere capping, or MTC, for maintenance of telomere capping. It seems likely that many of the newly identified pathways/ processes that affect growth of budding yeast cdc13-1 mutants will play evolutionarily conserved roles at telomeres. The high-throughput spot-testing approach that we describe is generally applicable and could aid in understanding other aspects of eukaryotic cell biology.

Two-color single molecule fluorescence coincidence method was extended to directly measure the enzymatic turnover of substrate and the efficiency of the translocation step of telomerase activity. The experimental results were able to identify the essential residues in the alignment domain of human telomerase RNA that contribute to the activity and processivity of telomerase.

Human diploid fibroblasts (HDF) immortalized by hTERT and simian virus 40 (SV40) early region (ER) exhibit a limited degree of transformation upon the expression of activated H-RAS (H-RAS V12) compared with rat embryonic fibroblasts (REF) immortalized by SV40 ER. Here, we identified FRA1 as a determinant for this difference in RAS-induced transformation. FRA1 was not induced by H-RAS V12 in the immortalized HDF, in contrast to its marked accumulation in the immortalized REF. Ectopic expression of FRA1 significantly enhanced anchorage-independent growth of various HDF expressing hTERT, SV40 ER, and H-RAS V12. More importantly, FRA1 could induce anchorage-independent growth as well as nude mice tumor formation of the immortalized HDF in the absence of H-RAS V12. The results of an in vitro kinase assay clearly showed that the RAS-induced extracellular signal-regulated kinase (ERK) activation, which is responsible for FRA1 induction, was markedly attenuated in the HDF compared with that in the REF, despite no obvious differences in the phosphorylation status of ERK between the species. Our results strongly suggest that HDF negatively regulate the mitogen-activated protein kinase kinase (MEK)/ERK pathway more efficiently than REF, and consequently express less malignant phenotypes in response to H-RAS V12. human diploid fibroblast | mitogen-activated protein kinase kinase/extracellular signal-regulated kinase | simian virus 40 early region | telomerase | rat embryonic fibroblasts

Human telomerase is a reverse-transcriptase enzyme that synthesizes the multikilobase repeating hexamer telomere sequence [(TTAGGG).sub.n] at the ends of chromosomes. Here we describe a designed approach to mimicry of telomerase, in which synthetic DNA nanocircles act as essentially infinite catalytic templates for efficient synthesis of long telomeres by DNA polymerase enzymes. Results show that the combination of a nanocircle and a DNA polymerase gives a positive telomere-repeat amplification protocol assay result for telomerase activity, and similar to the natural enzyme, it is inhibited by a known telomerase inhibitor. We show that artificial telomeres can be engineered on human chromosomes by this approach. This strategy allows for the preparation of synthetic telomeres for biological and structural study of telomeres and proteins that interact with them, and it raises the possibility of telomere engineering in cells without expression of telomerase itself. Finally, the results provide direct physical support for a recently proposed rolling-circle mechanism for telomerase-independent telomere elongation.

Cell-tracing analysis reveals that a disperse group of cells in the mouse liver express the enzyme telomerase, which preserves chromosome ends. These cells contribute to liver maintenance and regeneration.Cell-tracing analysis reveals that a disperse group of cells in the mouse liver express the enzyme telomerase, which preserves chromosome ends. These cells contribute to liver maintenance and regeneration., Author(s): Kenneth S. ZaretAuthor Affiliations:The telomerase enzyme and liver renewal First, the authors identified telomerase-expressing cells in the mouse liver and tracked descendent cells. The group genetically engineered mice to [...]

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.cell.2010.12.005 Byline: Jeffrey S. Chamberlain (1) Abstract: The lack of appropriate animal models has hampered efforts to develop therapies for Duchenne muscular dystrophy (DMD). A new mouse model lacking both dystrophin and telomerase () closely mimics the pathological progression of human DMD and shows that muscle stem cell activity is a key determinant of disease severity. Author Affiliation: (1) Departments of Neurology, Medicine, and Biochemistry, University of Washington, Seattle, WA 98195-7720, USA