Steven Artandi

All Publications

• Distributed hepatocytes expressing telomerase repopulate the liver in homeostasis and injury. Nature Lin, S., Nascimento, E. M., Gajera, C. R., Chen, L., Neuhöfer, P., Garbuzov, A., Wang, S., Artandi, S. E. 2018

  Abstract

  Hepatocytes are replenished gradually during homeostasis and robustly after liver injury1, 2. In adults, new hepatocytes originate from the existing hepatocyte pool3-8, but the cellular source of renewing hepatocytes remains unclear. Telomerase is expressed in many stem cell populations, and mutations in telomerase pathway genes have been linked to liver diseases9-11. Here we identify a subset of hepatocytes that expresses high levels of telomerase and show that this hepatocyte subset repopulates the liver during homeostasis and injury. Using lineage tracing from the telomerase reverse transcriptase (Tert) locus in mice, we demonstrate that rare hepatocytes with high telomerase expression (TERTHighhepatocytes) are distributed throughout the liver lobule. During homeostasis, these cells regenerate hepatocytes in all lobular zones, and both self-renew and differentiate to yield expanding hepatocyte clones that eventually dominate the liver. In response to injury, the repopulating activity of TERTHighhepatocytes is accelerated and their progeny cross zonal boundaries. RNA sequencing shows that metabolic genes are downregulated in TERTHighhepatocytes, indicating that metabolic activity and repopulating activity may be segregated within the hepatocyte lineage. Genetic ablation of TERTHighhepatocytes combined with chemical injury causes a marked increase in stellate cell activation and fibrosis. These results provide support for a 'distributed model' of hepatocyte renewal in which a subset of hepatocytes dispersed throughout the lobule clonally expands to maintain liver mass.

  View details for DOI 10.1038/s41586-018-0004-7

  View details for PubMedID 29618815

• An Activity Switch in Human Telomerase Based on RNA Conformation and Shaped by TCAB1. Cell Chen, L., Roake, C. M., Freund, A., Batista, P. J., Tian, S., Yin, Y. A., Gajera, C. R., Lin, S., Lee, B., Pech, M. F., Venteicher, A. S., Das, R., Chang, H. Y., Artandi, S. E. 2018

  Abstract

  Ribonucleoprotein enzymes require dynamic conformations of their RNA constituents for regulated catalysis. Human telomerase employs a non-coding RNA (hTR) with a bipartite arrangement of domains-a template-containing core and a distal three-way junction (CR4/5) that stimulates catalysis through unknown means. Here, we show that telomerase activity unexpectedly depends upon the holoenzyme protein TCAB1, which in turn controls conformation of CR4/5. Cells lacking TCAB1 exhibit a marked reduction in telomerase catalysis without affecting enzyme assembly. Instead, TCAB1 inactivation causes unfolding of CR4/5 helices that are required for catalysis and for association with the telomerase reverse-transcriptase (TERT). CR4/5 mutations derived from patients with telomere biology disorders provoke defects in catalysis and TERT binding similar to TCAB1 inactivation. These findings reveal a conformational "activity switch" in human telomerase RNA controlling catalysis and TERT engagement. The identification of two discrete catalytic states for telomerase suggests an intramolecular means for controlling telomerase in cancers and progenitor cells.

  View details for DOI 10.1016/j.cell.2018.04.039

  View details for PubMedID 29804836

• Control of Cellular Aging, Tissue Function, and Cancer by p53 Downstream of Telomeres. Cold Spring Harbor perspectives in medicine Roake, C. M., Artandi, S. E. 2017; 7 (5)

  Abstract

  Telomeres, the nucleoprotein complex at the ends of eukaryotic chromosomes, perform an essential cellular role in part by preventing the chromosomal end from initiating a DNA-damage response. This function of telomeres can be compromised as telomeres erode either as a consequence of cell division in culture or as a normal part of cellular ageing in proliferative tissues. Telomere dysfunction in this context leads to DNA-damage signaling and activation of the tumor-suppressor protein p53, which then can prompt either cellular senescence or apoptosis. By culling cells with dysfunctional telomeres, p53 plays a critical role in protecting tissues against the effects of critically short telomeres. However, as telomere dysfunction worsens, p53 likely exacerbates short telomere-driven tissue failure diseases, including pulmonary fibrosis, aplastic anemia, and liver cirrhosis. In cells lacking p53, unchecked telomere shortening drives chromosomal end-to-end fusions and cycles of chromosome fusion-bridge-breakage. Incipient cancer cells confronting these telomere barriers must disable p53 signaling to avoid senescence and eventually up-regulate telomerase to achieve cellular immortality. The recent findings of highly recurrent activating mutations in the promoter for the telomerase reverse transcriptase (TERT) gene in diverse human cancers, together with the widespread mutations in p53 in cancer, provide support for the idea that circumvention of a telomere-p53 checkpoint is essential for malignant progression in human cancer.

  View details for DOI 10.1101/cshperspect.a026088

  View details for PubMedID 28289249

• Pulmonary arteriovenous malformations: an uncharacterised phenotype of dyskeratosis congenita and related telomere biology disorders EUROPEAN RESPIRATORY JOURNAL Khincha, P. P., Bertuch, A. A., Agarwal, S., Townsley, D. M., Young, N. S., Keel, S., Shimamura, A., Boulad, F., Simoneau, T., Justino, H., Kuo, C., Artandi, S., McCaslin, C., Cox, D. W., Chaffee, S., Collins, B. F., Giri, N., Alter, B. P., Raghu, G., Savage, S. A. 2017; 49 (1)

  View details for DOI 10.1183/13993003.01640-2016

  View details for Web of Science ID 000397423500036

  View details for PubMedID 27824607

  View details for PubMedCentralID PMC5841586

• DNA repair: Telomere-lengthening mechanism revealed. Nature Roake, C. M., Artandi, S. E. 2016; 539 (7627): 35-36

  View details for DOI 10.1038/nature19483

  View details for PubMedID 27760112

• Dual SMAD Signaling Inhibition Enables Long-Term Expansion of Diverse Epithelial Basal Cells. Cell stem cell Mou, H., Vinarsky, V., Tata, P. R., Brazauskas, K., Choi, S. H., Crooke, A. K., Zhang, B., Solomon, G. M., Turner, B., Bihler, H., Harrington, J., Lapey, A., Channick, C., Keyes, C., Freund, A., Artandi, S., Mense, M., Rowe, S., Engelhardt, J. F., Hsu, Y., Rajagopal, J. 2016; 19 (2): 217-231

  Abstract

  Functional modeling of many adult epithelia is limited by the difficulty in maintaining relevant stem cell populations in culture. Here, we show that dual inhibition of SMAD signaling pathways enables robust expansion of primary epithelial basal cell populations. We find that TGFβ/BMP/SMAD pathway signaling is strongly activated in luminal and suprabasal cells of several epithelia, but suppressed in p63+ basal cells. In airway epithelium, SMAD signaling promotes differentiation, and its inhibition leads to stem cell hyperplasia. Using dual SMAD signaling inhibition in a feeder-free culture system, we have been able to expand airway basal stem cells from multiple species. Expanded cells can produce functional airway epithelium physiologically responsive to clinically relevant drugs, such as CFTR modulators. This approach is effective for the clonal expansion of single human cells and for basal cell populations from epithelial tissues from all three germ layers and therefore may be broadly applicable for modeling of epithelia.

  View details for DOI 10.1016/j.stem.2016.05.012

  View details for PubMedID 27320041

• High telomerase is a hallmark of undifferentiated spermatogonia and is required for maintenance of male germline stem cells. Genes & development Pech, M. F., Garbuzov, A., Hasegawa, K., Sukhwani, M., Zhang, R. J., Benayoun, B. A., Brockman, S. A., Lin, S., Brunet, A., Orwig, K. E., Artandi, S. E. 2015; 29 (23): 2420-2434

  Abstract

  Telomerase inactivation causes loss of the male germline in worms, fish, and mice, indicating a conserved dependence on telomere maintenance in this cell lineage. Here, using telomerase reverse transcriptase (Tert) reporter mice, we found that very high telomerase expression is a hallmark of undifferentiated spermatogonia, the mitotic population where germline stem cells reside. We exploited these high telomerase levels as a basis for purifying undifferentiated spermatogonia using fluorescence-activated cell sorting. Telomerase levels in undifferentiated spermatogonia and embryonic stem cells are comparable and much greater than in somatic progenitor compartments. Within the germline, we uncovered an unanticipated gradient of telomerase activity that also enables isolation of more mature populations. Transcriptomic comparisons of Tert(High) undifferentiated spermatogonia and Tert(Low) differentiated spermatogonia by RNA sequencing reveals marked differences in cell cycle and key molecular features of each compartment. Transplantation studies show that germline stem cell activity is confined to the Tert(High) cKit(-) population. Telomere shortening in telomerase knockout strains causes depletion of undifferentiated spermatogonia and eventual loss of all germ cells after undifferentiated spermatogonia drop below a critical threshold. These data reveal that high telomerase expression is a fundamental characteristic of germline stem cells, thus explaining the broad dependence on telomerase for germline immortality in metazoans.

  View details for DOI 10.1101/gad.271783.115

  View details for PubMedID 26584619

  View details for PubMedCentralID PMC4691947

• Keeping It in the Family: ATRX Loss Promotes Persistent Sister Telomere Cohesion in ALT Cancer Cells. Cancer cell Roake, C. M., Artandi, S. E. 2015; 28 (3): 277-279

  Abstract

  In this issue of Cancer Cell, Ramamoorthy and Smith report that cancer cells that maintain their chromosome ends through alternative lengthening of telomeres (ALT) display persistent sister telomere cohesion. This delayed resolution of sister telomere cohesion depends upon the loss of ATRX and its histone-sequestering function and is associated with increased recombination between sister telomeres.

  View details for DOI 10.1016/j.ccell.2015.08.005

  View details for PubMedID 26373274

• Reversibility of Defective Hematopoiesis Caused by Telomere Shortening in Telomerase Knockout Mice PLOS ONE Raval, A., Behbehani, G. K., Le Xuan Truong Nguyen, L. X., Thomas, D., Kusler, B., Garbuzov, A., Ramunas, J., Holbrook, C., Park, C. Y., Blau, H., Nolan, G. P., Artandi, S. E., Mitchell, B. S. 2015; 10 (7)

  View details for DOI 10.1371/journal.pone.0131722

  View details for Web of Science ID 000358154400065

• A platform for rapid exploration of aging and diseases in a naturally short-lived vertebrate. Cell Harel, I., Benayoun, B. A., Machado, B., Singh, P. P., Hu, C., Pech, M. F., Valenzano, D. R., Zhang, E., Sharp, S. C., Artandi, S. E., Brunet, A. 2015; 160 (5): 1013-1026

  Abstract

  Aging is a complex process that affects multiple organs. Modeling aging and age-related diseases in the lab is challenging because classical vertebrate models have relatively long lifespans. Here, we develop the first platform for rapid exploration of age-dependent traits and diseases in vertebrates, using the naturally short-lived African turquoise killifish. We provide an integrative genomic and genome-editing toolkit in this organism using our de-novo-assembled genome and the CRISPR/Cas9 technology. We mutate many genes encompassing the hallmarks of aging, and for a subset, we produce stable lines within 2-3 months. As a proof of principle, we show that fish deficient for the protein subunit of telomerase exhibit the fastest onset of telomere-related pathologies among vertebrates. We further demonstrate the feasibility of creating specific genetic variants. This genome-to-phenotype platform represents a unique resource for studying vertebrate aging and disease in a high-throughput manner and for investigating candidates arising from human genome-wide studies.

  View details for DOI 10.1016/j.cell.2015.01.038

  View details for PubMedID 25684364

• A platform for rapid exploration of aging and diseases in a naturally short-lived vertebrate. Cell Harel, I., Benayoun, B. A., Machado, B., Singh, P. P., Hu, C., Pech, M. F., Valenzano, D. R., Zhang, E., Sharp, S. C., Artandi, S. E., Brunet, A. 2015; 160 (5): 1013-1026

  View details for DOI 10.1016/j.cell.2015.01.038

  View details for PubMedID 25684364

• Inhibition of pluripotency networks by the rb tumor suppressor restricts reprogramming and tumorigenesis. Cell stem cell Kareta, M. S., Gorges, L. L., Hafeez, S., Benayoun, B. A., Marro, S., Zmoos, A., Cecchini, M. J., Spacek, D., Batista, L. F., O'Brien, M., Ng, Y., Ang, C. E., Vaka, D., Artandi, S. E., Dick, F. A., Brunet, A., Sage, J., Wernig, M. 2015; 16 (1): 39-50

  Abstract

  Mutations in the retinoblastoma tumor suppressor gene Rb are involved in many forms of human cancer. In this study, we investigated the early consequences of inactivating Rb in the context of cellular reprogramming. We found that Rb inactivation promotes the reprogramming of differentiated cells to a pluripotent state. Unexpectedly, this effect is cell cycle independent, and instead reflects direct binding of Rb to pluripotency genes, including Sox2 and Oct4, which leads to a repressed chromatin state. More broadly, this regulation of pluripotency networks and Sox2 in particular is critical for the initiation of tumors upon loss of Rb in mice. These studies therefore identify Rb as a global transcriptional repressor of pluripotency networks, providing a molecular basis for previous reports about its involvement in cell fate pliability, and implicate misregulation of pluripotency factors such as Sox2 in tumorigenesis related to loss of Rb function.

  View details for DOI 10.1016/j.stem.2014.10.019

  View details for PubMedID 25467916

  View details for PubMedCentralID PMC4389904

• Reversibility of Defective Hematopoiesis Caused by Telomere Shortening in Telomerase Knockout Mice. PloS one Raval, A., Behbehani, G. K., Nguyen, L. X., Thomas, D., Kusler, B., Garbuzov, A., Ramunas, J., Holbrook, C., Park, C. Y., Blau, H., Nolan, G. P., Artandi, S. E., Mitchell, B. S. 2015; 10 (7)

  Abstract

  Telomere shortening is common in bone marrow failure syndromes such as dyskeratosis congenita (DC), aplastic anemia (AA) and myelodysplastic syndromes (MDS). However, improved knowledge of the lineage-specific consequences of telomere erosion and restoration of telomere length in hematopoietic progenitors is required to advance therapeutic approaches. We have employed a reversible murine model of telomerase deficiency to compare the dependence of erythroid and myeloid lineage differentiation on telomerase activity. Fifth generation Tert-/- (G5 Tert-/-) mice with shortened telomeres have significant anemia, decreased erythroblasts and reduced hematopoietic stem cell (HSC) populations associated with neutrophilia and increased myelopoiesis. Intracellular multiparameter analysis by mass cytometry showed significantly reduced cell proliferation and increased sensitivity to activation of DNA damage checkpoints in erythroid progenitors and in erythroid-biased CD150hi HSC, but not in myeloid progenitors. Strikingly, Cre-inducible reactivation of telomerase activity restored hematopoietic stem and progenitor cell (HSPC) proliferation, normalized the DNA damage response, and improved red cell production and hemoglobin levels. These data establish a direct link between the loss of TERT activity, telomere shortening and defective erythropoiesis and suggest that novel strategies to restore telomerase function may have an important role in the treatment of the resulting anemia.

  View details for DOI 10.1371/journal.pone.0131722

  View details for PubMedID 26133370

  View details for PubMedCentralID PMC4489842

• Proteostatic Control of Telomerase Function through TRiC-Mediated Folding of TCAB1 CELL Freund, A., Zhong, F. L., Venteicher, A. S., Meng, Z., Veenstra, T. D., Frydman, J., Artandi, S. E. 2014; 159 (6): 1389-1403

  Abstract

  Telomere maintenance by telomerase is impaired in the stem cell disease dyskeratosis congenita and during human aging. Telomerase depends upon a complex pathway for enzyme assembly, localization in Cajal bodies, and association with telomeres. Here, we identify the chaperonin CCT/TRiC as a critical regulator of telomerase trafficking using a high-content genome-wide siRNA screen in human cells for factors required for Cajal body localization. We find that TRiC is required for folding the telomerase cofactor TCAB1, which controls trafficking of telomerase and small Cajal body RNAs (scaRNAs). Depletion of TRiC causes loss of TCAB1 protein, mislocalization of telomerase and scaRNAs to nucleoli, and failure of telomere elongation. DC patient-derived mutations in TCAB1 impair folding by TRiC, disrupting telomerase function and leading to severe disease. Our findings establish a critical role for TRiC-mediated protein folding in the telomerase pathway and link proteostasis, telomere maintenance, and human disease.

  View details for DOI 10.1016/j.cell.2014.10.059

  View details for Web of Science ID 000346652900017

  View details for PubMedID 25467444

• Understanding telomere diseases through analysis of patient-derived iPS cells. Current opinion in genetics & development Batista, L. F., Artandi, S. E. 2013; 23 (5): 526-533

  Abstract

  A unique characteristic of tissue stem cells is the ability to self-renew, a process that enables the life-long maintenance of many organs. Stem cell self-renewal is dependent in part on the synthesis of telomere repeats by the enzyme telomerase. Defects in telomerase and in genes in the telomere maintenance pathway result in diverse disease states, including dyskeratosis congenita, pulmonary fibrosis, aplastic anemia, liver cirrhosis and cancer. Many of these disease states share a tissue failure phenotype, such as loss of bone marrow cells or failure of pulmonary epithelium, suggesting that stem cell dysfunction is a common pathophysiological mechanism underlying these telomere diseases. Studies of telomere diseases in undifferentiated iPS cells have provided a quantitative relationship between the magnitude of biochemical defects in the telomerase pathway and disease severity in patients, thereby establishing a clear correlation between genotype and phenotype in telomere disease states. Modeling telomere diseases in iPS cells has also revealed diverse underlying disease mechanisms, including reduced telomerase catalytic activity, diminished assembly of the telomerase holoenzyme and impaired trafficking of the enzyme within the nucleus. These studies highlight the need for therapies tailored to the underlying biochemical defect in each class of patients.

  View details for DOI 10.1016/j.gde.2013.07.006

  View details for PubMedID 23993228

• TPP1 OB-Fold Domain Controls Telomere Maintenance by Recruiting Telomerase to Chromosome Ends CELL Zhong, F. L., Batista, L. F., Freund, A., Pech, M. F., Venteicher, A. S., Artandi, S. E. 2012; 150 (3): 481-494

  Abstract

  Telomere synthesis in cancer cells and stem cells involves trafficking of telomerase to Cajal bodies, and telomerase is thought to be recruited to telomeres through interactions with telomere-binding proteins. Here, we show that the OB-fold domain of the telomere-binding protein TPP1 recruits telomerase to telomeres through an association with the telomerase reverse transcriptase TERT. When tethered away from telomeres and other telomere-binding proteins, the TPP1 OB-fold domain is sufficient to recruit telomerase to a heterologous chromatin locus. Expression of a minimal TPP1 OB-fold inhibits telomere maintenance by blocking access of telomerase to its cognate binding site at telomeres. We identify amino acids required for the TPP1-telomerase interaction, including specific loop residues within the TPP1 OB-fold domain and individual residues within TERT, some of which are mutated in a subset of pulmonary fibrosis patients. These data define a potential interface for telomerase-TPP1 interaction required for telomere maintenance and implicate defective telomerase recruitment in telomerase-related disease.

  View details for DOI 10.1016/j.cell.2012.07.012

  View details for Web of Science ID 000307301400009

  View details for PubMedID 22863003

• Reversible cell-cycle entry in adult kidney podocytes through regulated control of telomerase and Wnt signaling. Nature medicine Shkreli, M., Sarin, K. Y., Pech, M. F., Papeta, N., Chang, W., Brockman, S. A., Cheung, P., Lee, E., Kuhnert, F., Olson, J. L., Kuo, C. J., Gharavi, A. G., D'Agati, V. D., Artandi, S. E. 2012; 18 (1): 111-119

  Abstract

  Mechanisms of epithelial cell renewal remain poorly understood in the mammalian kidney, particularly in the glomerulus, a site of cellular damage in chronic kidney disease. Within the glomerulus, podocytes--differentiated epithelial cells crucial for filtration--are thought to lack substantial capacity for regeneration. Here we show that podocytes rapidly lose differentiation markers and enter the cell cycle in adult mice in which the telomerase protein component TERT is conditionally expressed. Transgenic TERT expression in mice induces marked upregulation of Wnt signaling and disrupts glomerular structure, resulting in a collapsing glomerulopathy resembling those in human disease, including HIV-associated nephropathy (HIVAN). Human and mouse HIVAN kidneys show increased expression of TERT and activation of Wnt signaling, indicating that these are general features of collapsing glomerulopathies. Silencing transgenic TERT expression or inhibiting Wnt signaling through systemic expression of the Wnt inhibitor Dkk1 in either TERT transgenic mice or in a mouse model of HIVAN results in marked normalization of podocytes, including rapid cell-cycle exit, re-expression of differentiation markers and improved filtration barrier function. These data reveal an unexpected capacity of podocytes to reversibly enter the cell cycle, suggest that podocyte renewal may contribute to glomerular homeostasis and implicate the telomerase and Wnt-β-catenin pathways in podocyte proliferation and disease.

  View details for DOI 10.1038/nm.2550

  View details for PubMedID 22138751

• Reversible cell-cycle entry in adult kidney podocytes through regulated control of telomerase and Wnt signaling NATURE MEDICINE Shkreli, M., Sarin, K. Y., Pech, M. F., Papeta, N., Chang, W., Brockman, S. A., Cheung, P., Lee, E., Kuhnert, F., Olson, J. L., Kuo, C. J., Gharavi, A. G., D'Agati, V. D., Artandi, S. E. 2012; 18 (1): 111-119

  View details for DOI 10.1038/nm.2550

  View details for Web of Science ID 000299018600036

• Genomic Maps of Long Noncoding RNA Occupancy Reveal Principles of RNA-Chromatin Interactions MOLECULAR CELL Chu, C., Qu, K., Zhong, F. L., Artandi, S. E., Chang, H. Y. 2011; 44 (4): 667-678

  Abstract

  Long noncoding RNAs (lncRNAs) are key regulators of chromatin state, yet the nature and sites of RNA-chromatin interaction are mostly unknown. Here we introduce Chromatin Isolation by RNA Purification (ChIRP), where tiling oligonucleotides retrieve specific lncRNAs with bound protein and DNA sequences, which are enumerated by deep sequencing. ChIRP-seq of three lncRNAs reveal that RNA occupancy sites in the genome are focal, sequence-specific, and numerous. Drosophila roX2 RNA occupies male X-linked gene bodies with increasing tendency toward the 3' end, peaking at CES sites. Human telomerase RNA TERC occupies telomeres and Wnt pathway genes. HOTAIR lncRNA preferentially occupies a GA-rich DNA motif to nucleate broad domains of Polycomb occupancy and histone H3 lysine 27 trimethylation. HOTAIR occupancy occurs independently of EZH2, suggesting the order of RNA guidance of Polycomb occupancy. ChIRP-seq is generally applicable to illuminate the intersection of RNA and chromatin with newfound precision genome wide.

  View details for DOI 10.1016/j.molcel.2011.08.027

  View details for Web of Science ID 000297387800017

  View details for PubMedID 21963238

  View details for PubMedCentralID PMC3249421

• In Situ Genetic Correction of the Sickle Cell Anemia Mutation in Human Induced Pluripotent Stem Cells Using Engineered Zinc Finger Nucleases STEM CELLS Sebastiano, V., Maeder, M. L., Angstman, J. F., Haddad, B., Khayter, C., Yeo, D. T., Goodwin, M. J., Hawkins, J. S., Ramirez, C. L., Batista, L. F., Artandi, S. E., Wernig, M., Joung, J. K. 2011; 29 (11): 1717-1726

  Abstract

  The combination of induced pluripotent stem cell (iPSC) technology and targeted gene modification by homologous recombination (HR) represents a promising new approach to generate genetically corrected, patient-derived cells that could be used for autologous transplantation therapies. This strategy has several potential advantages over conventional gene therapy including eliminating the need for immunosuppression, avoiding the risk of insertional mutagenesis by therapeutic vectors, and maintaining expression of the corrected gene by endogenous control elements rather than a constitutive promoter. However, gene targeting in human pluripotent cells has remained challenging and inefficient. Recently, engineered zinc finger nucleases (ZFNs) have been shown to substantially increase HR frequencies in human iPSCs, raising the prospect of using this technology to correct disease causing mutations. Here, we describe the generation of iPSC lines from sickle cell anemia patients and in situ correction of the disease causing mutation using three ZFN pairs made by the publicly available oligomerized pool engineering method (OPEN). Gene-corrected cells retained full pluripotency and a normal karyotype following removal of reprogramming factor and drug-resistance genes. By testing various conditions, we also demonstrated that HR events in human iPSCs can occur as far as 82 bps from a ZFN-induced break. Our approach delineates a roadmap for using ZFNs made by an open-source method to achieve efficient, transgene-free correction of monogenic disease mutations in patient-derived iPSCs. Our results provide an important proof of principle that ZFNs can be used to produce gene-corrected human iPSCs that could be used for therapeutic applications.

  View details for DOI 10.1002/stem.718

  View details for Web of Science ID 000296565500009

  View details for PubMedID 21898685

  View details for PubMedCentralID PMC3285772

• Telomere shortening and loss of self-renewal in dyskeratosis congenita induced pluripotent stem cells NATURE Batista, L. F., Pech, M., Zhong, F. L., Nguyen, H. N., Xie, K. T., Zaug, A. J., Crary, S. M., Choi, J., Sebastiano, V., Cherry, A., Giri, N., Wernig, M., Alter, B. P., Cech, T. R., Savage, S. A., Pera, R. A., Artandi, S. E. 2011; 474 (7351): 399-?

  Abstract

  The differentiation of patient-derived induced pluripotent stem cells (iPSCs) to committed fates such as neurons, muscle and liver is a powerful approach for understanding key parameters of human development and disease. Whether undifferentiated iPSCs themselves can be used to probe disease mechanisms is uncertain. Dyskeratosis congenita is characterized by defective maintenance of blood, pulmonary tissue and epidermal tissues and is caused by mutations in genes controlling telomere homeostasis. Short telomeres, a hallmark of dyskeratosis congenita, impair tissue stem cell function in mouse models, indicating that a tissue stem cell defect may underlie the pathophysiology of dyskeratosis congenita. Here we show that even in the undifferentiated state, iPSCs from dyskeratosis congenita patients harbour the precise biochemical defects characteristic of each form of the disease and that the magnitude of the telomere maintenance defect in iPSCs correlates with clinical severity. In iPSCs from patients with heterozygous mutations in TERT, the telomerase reverse transcriptase, a 50% reduction in telomerase levels blunts the natural telomere elongation that accompanies reprogramming. In contrast, mutation of dyskerin (DKC1) in X-linked dyskeratosis congenita severely impairs telomerase activity by blocking telomerase assembly and disrupts telomere elongation during reprogramming. In iPSCs from a form of dyskeratosis congenita caused by mutations in TCAB1 (also known as WRAP53), telomerase catalytic activity is unperturbed, yet the ability of telomerase to lengthen telomeres is abrogated, because telomerase mislocalizes from Cajal bodies to nucleoli within the iPSCs. Extended culture of DKC1-mutant iPSCs leads to progressive telomere shortening and eventual loss of self-renewal, indicating that a similar process occurs in tissue stem cells in dyskeratosis congenita patients. These findings in iPSCs from dyskeratosis congenita patients reveal that undifferentiated iPSCs accurately recapitulate features of a human stem cell disease and may serve as a cell-culture-based system for the development of targeted therapeutics.

  View details for DOI 10.1038/nature10084

  View details for Web of Science ID 000291647100050

  View details for PubMedID 21602826

  View details for PubMedCentralID PMC3155806

• TRAPping telomerase within the intestinal stem cell niche EMBO JOURNAL Pech, M. F., Artandi, S. E. 2011; 30 (6): 986-987

  View details for DOI 10.1038/emboj.2011.51

  View details for Web of Science ID 000289672900002

  View details for PubMedID 21407250

• Disruption of telomerase trafficking by TCAB1 mutation causes dyskeratosis congenita GENES & DEVELOPMENT Zhong, F., Savage, S. A., Shkreli, M., Giri, N., Jessop, L., Myers, T., Chen, R., Alter, B. P., Artandi, S. E. 2011; 25 (1): 11-16

  Abstract

  Dyskeratosis congenita (DC) is a genetic disorder of defective tissue maintenance and cancer predisposition caused by short telomeres and impaired stem cell function. Telomerase mutations are thought to precipitate DC by reducing either the catalytic activity or the overall levels of the telomerase complex. However, the underlying genetic mutations and the mechanisms of telomere shortening remain unknown for as many as 50% of DC patients, who lack mutations in genes controlling telomere homeostasis. Here, we show that disruption of telomerase trafficking accounts for unknown cases of DC. We identify DC patients with missense mutations in TCAB1, a telomerase holoenzyme protein that facilitates trafficking of telomerase to Cajal bodies. Compound heterozygous mutations in TCAB1 disrupt telomerase localization to Cajal bodies, resulting in misdirection of telomerase RNA to nucleoli, which prevents telomerase from elongating telomeres. Our findings establish telomerase mislocalization as a novel cause of DC, and suggest that telomerase trafficking defects may contribute more broadly to the pathogenesis of telomere-related disease.

  View details for DOI 10.1101/gad.2006411

  View details for Web of Science ID 000285870300002

  View details for PubMedID 21205863

• Short Telomeres and Stem Cell Exhaustion Model Duchenne Muscular Dystrophy in mdx/mTR Mice CELL Sacco, A., Mourkioti, F., Tran, R., Choi, J., Llewellyn, M., Kraft, P., Shkreli, M., Delp, S., Pomerantz, J. H., Artandi, S. E., Blau, H. M. 2010; 143 (7): 1059-1071

  Abstract

  In Duchenne muscular dystrophy (DMD), dystrophin mutation leads to progressive lethal skeletal muscle degeneration. For unknown reasons, dystrophin deficiency does not recapitulate DMD in mice (mdx), which have mild skeletal muscle defects and potent regenerative capacity. We postulated that human DMD progression is a consequence of loss of functional muscle stem cells (MuSC), and the mild mouse mdx phenotype results from greater MuSC reserve fueled by longer telomeres. We report that mdx mice lacking the RNA component of telomerase (mdx/mTR) have shortened telomeres in muscle cells and severe muscular dystrophy that progressively worsens with age. Muscle wasting severity parallels a decline in MuSC regenerative capacity and is ameliorated histologically by transplantation of wild-type MuSC. These data show that DMD progression results, in part, from a cell-autonomous failure of MuSC to maintain the damage-repair cycle initiated by dystrophin deficiency. The essential role of MuSC function has therapeutic implications for DMD.

  View details for DOI 10.1016/j.cell.2010.11.039

  View details for Web of Science ID 000285625400005

  View details for PubMedID 21145579

  View details for PubMedCentralID PMC3025608

• Shared molecular mechanisms regulate multiple catenin proteins: canonical Wnt signals and components modulate p120-catenin isoform-1 and additional p120 subfamily members JOURNAL OF CELL SCIENCE Hong, J. Y., Park, J., Cho, K., Gu, D., Ji, H., Artandi, S. E., McCrea, P. D. 2010; 123 (24): 4351-4365

  Abstract

  Wnt signaling pathways have fundamental roles in animal development and tumor progression. Here, employing Xenopus embryos and mammalian cell lines, we report that the degradation machinery of the canonical Wnt pathway modulates p120-catenin protein stability through mechanisms shared with those regulating β-catenin. For example, in common with β-catenin, exogenous expression of destruction complex components, such as GSK3β and axin, promotes degradation of p120-catenin. Again in parallel with β-catenin, reduction of canonical Wnt signals upon depletion of LRP5 and LRP6 results in p120-catenin degradation. At the primary sequence level, we resolved conserved GSK3β phosphorylation sites in the amino-terminal region of p120-catenin present exclusively in isoform-1. Point-mutagenesis of these residues inhibited the association of destruction complex components, such as those involved in ubiquitylation, resulting in stabilization of p120-catenin. Functionally, in line with predictions, p120 stabilization increased its signaling activity in the context of the p120-Kaiso pathway. Importantly, we found that two additional p120-catenin family members, ARVCF-catenin and δ-catenin, associate with axin and are degraded in its presence. Thus, as supported using gain- and loss-of-function approaches in embryo and cell line systems, canonical Wnt signals appear poised to have an impact upon a breadth of catenin biology in vertebrate development and, possibly, human cancers.

  View details for DOI 10.1242/jcs.067199

  View details for Web of Science ID 000284837100017

  View details for PubMedID 21098636

• Telomeres and telomerase in cancer CARCINOGENESIS Artandi, S. E., DePinho, R. A. 2010; 31 (1): 9-18

  Abstract

  Myriad genetic and epigenetic alterations are required to drive normal cells toward malignant transformation. These somatic events commandeer many signaling pathways that cooperate to endow aspiring cancer cells with a full range of biological capabilities needed to grow, disseminate and ultimately kill its host. Cancer genomes are highly rearranged and are characterized by complex translocations and regional copy number alterations that target loci harboring cancer-relevant genes. Efforts to uncover the underlying mechanisms driving genome instability in cancer have revealed a prominent role for telomeres. Telomeres are nucleoprotein structures that protect the ends of eukaryotic chromosomes and are particularly vulnerable due to progressive shortening during each round of DNA replication and, thus, a lifetime of tissue renewal places the organism at risk for increasing chromosomal instability. Indeed, telomere erosion has been documented in aging tissues and hyperproliferative disease states-conditions strongly associated with increased cancer risk. Telomere dysfunction can produce the opposing pathophysiological states of degenerative aging or cancer with the specific outcome dictated by the integrity of DNA damage checkpoint responses. In most advanced cancers, telomerase is reactivated and serves to maintain telomere length and emerging data have also documented the capacity of telomerase to directly regulate cancer-promoting pathways. This review covers the role of telomeres and telomerase in the biology of normal tissue stem/progenitor cells and in the development of cancer.

  View details for DOI 10.1093/carcin/bgp268

  View details for Web of Science ID 000273493100002

  View details for PubMedID 19887512

• Reverse Transcribing the Code for Chromosome Stability MOLECULAR CELL Artandi, S. E., Cooper, J. P. 2009; 36 (5): 715-719

  Abstract

  The linearity of eukaryotic chromosomes presents challenges to cells, as the presence of DNA "ends" poses problems for the DNA replication machinery and the cell's damage response systems. This year's Nobel Prize in Physiology or Medicine recognized groundbreaking studies establishing the telomere field as a crucial area of biomedical research.

  View details for DOI 10.1016/j.molcel.2009.11.030

  View details for Web of Science ID 000272965400001

  View details for PubMedID 20005831

• Telomerase modulates Wnt signalling by association with target gene chromatin NATURE Park, J., Venteicher, A. S., Hong, J. Y., Choi, J., Jun, S., Shkreli, M., Chang, W., Meng, Z., Cheung, P., Ji, H., McLaughlin, M., Veenstra, T. D., Nusse, R., McCrea, P. D., Artandi, S. E. 2009; 460 (7251): 66-U77

  Abstract

  Stem cells are controlled, in part, by genetic pathways frequently dysregulated during human tumorigenesis. Either stimulation of Wnt/beta-catenin signalling or overexpression of telomerase is sufficient to activate quiescent epidermal stem cells in vivo, although the mechanisms by which telomerase exerts these effects are not understood. Here we show that telomerase directly modulates Wnt/beta-catenin signalling by serving as a cofactor in a beta-catenin transcriptional complex. The telomerase protein component TERT (telomerase reverse transcriptase) interacts with BRG1 (also called SMARCA4), a SWI/SNF-related chromatin remodelling protein, and activates Wnt-dependent reporters in cultured cells and in vivo. TERT serves an essential role in formation of the anterior-posterior axis in Xenopus laevis embryos, and this defect in Wnt signalling manifests as homeotic transformations in the vertebrae of Tert(-/-) mice. Chromatin immunoprecipitation of the endogenous TERT protein from mouse gastrointestinal tract shows that TERT physically occupies gene promoters of Wnt-dependent genes. These data reveal an unanticipated role for telomerase as a transcriptional modulator of the Wnt/beta-catenin signalling pathway.

  View details for DOI 10.1038/nature08137

  View details for Web of Science ID 000267545200030

  View details for PubMedID 19571879

  View details for PubMedCentralID PMC4349391

• Stem Cell Aging and Aberrant Differentiation within the Niche CELL STEM CELL Choi, J., Artandi, S. 2009; 5 (1): 6-8

  Abstract

  Stem cells age, but the underlying mechanisms remain unclear. In a recent issue of Cell, Inomata and colleagues (2009) show that DNA damage, a prime suspect in stem cell aging, causes graying and loss of melanocyte stem cells by inducing premature differentiation, without inducing apoptosis or senescence.

  View details for DOI 10.1016/j.stem.2009.06.006

  View details for Web of Science ID 000267879200004

  View details for PubMedID 19570507

• Telomere Uncapping, Chromosomes, and Carcinomas CANCER CELL Batista, L. F., Artandi, S. E. 2009; 15 (6): 455-457

  Abstract

  Data from mouse models and from human cancers have supported the idea that telomere shortening leads to chromosomal instability and epithelial carcinogenesis. In this issue of Cancer Cell, Else et al. demonstrate that telomere uncapping-altering a protein that protects chromosome ends without shortening telomeres-also results in epithelial cancers.

  View details for DOI 10.1016/j.ccr.2009.05.006

  View details for Web of Science ID 000266686500001

  View details for PubMedID 19477422

• TCAB1 Driving telomerase to Cajal bodies CELL CYCLE Venteicher, A. S., Artandi, S. E. 2009; 8 (9): 1329-1331

  Abstract

  Telomerase supports the proliferation of progenitor cells and tumor cells by adding telomere repeats to chromosome ends. The low abundance and restricted expression pattern of telomerase have limited our knowledge of this important enzyme. A new telomerase protein, TCAB1, sheds light on the pathway that governs telomerase holoenzyme assembly and function in vivo. TCAB1 is a component of active telomerase and is required for the telomerase holoenzyme to accumulate in Cajal bodies and to elongate telomeres. These findings provide important new insights into how telomerase functions in cancer and in stem cell biology.

  View details for Web of Science ID 000266114600016

  View details for PubMedID 19342896

• A Human Telomerase Holoenzyme Protein Required for Cajal Body Localization and Telomere Synthesis SCIENCE Venteicher, A. S., Abreu, E. B., Meng, Z., McCann, K. E., Terns, R. M., Veenstra, T. D., Terns, M. P., Artandi, S. E. 2009; 323 (5914): 644-648

  Abstract

  Telomerase is a ribonucleoprotein (RNP) complex that synthesizes telomere repeats in tissue progenitor cells and cancer cells. Active human telomerase consists of at least three principal subunits, including the telomerase reverse transcriptase, the telomerase RNA (TERC), and dyskerin. Here, we identify a holoenzyme subunit, TCAB1 (telomerase Cajal body protein 1), that is notably enriched in Cajal bodies, nuclear sites of RNP processing that are important for telomerase function. TCAB1 associates with active telomerase enzyme, established telomerase components, and small Cajal body RNAs that are involved in modifying splicing RNAs. Depletion of TCAB1 by using RNA interference prevents TERC from associating with Cajal bodies, disrupts telomerase-telomere association, and abrogates telomere synthesis by telomerase. Thus, TCAB1 controls telomerase trafficking and is required for telomere synthesis in human cancer cells.

  View details for DOI 10.1126/science.1165357

  View details for Web of Science ID 000262862800046

  View details for PubMedID 19179534

• Telomerase-dependent and -independent chromosome healing in mouse embryonic stem cells DNA REPAIR Gao, Q., Reynolds, G. E., Wilcox, A., Miller, D., Cheung, P., Artandi, S. E., Murnane, J. P. 2008; 7 (8): 1233-1249

  Abstract

  Telomeres play an important role in protecting the ends of chromosomes and preventing chromosome fusion. We have previously demonstrated that double-strand breaks near telomeres in mammalian cells result in either the addition of a new telomere at the site of the break, termed chromosome healing, or sister chromatid fusion that initiates chromosome instability. In the present study, we have investigated the role of telomerase in chromosome healing and the importance of chromosome healing in preventing chromosome instability. In embryonic stem cell lines that are wild type for the catalytic subunit of telomerase (TERT), chromosome healing at I-SceI-induced double-strand breaks near telomeres accounted for 22 of 35 rearrangements, with the new telomeres added directly at the site of the break in all but one instance. In contrast, in two TERT-knockout embryonic stem cell lines, chromosome healing accounted for only 1 of 62 rearrangements, with a 23 bp insertion at the site of the sole chromosome-healing event. However, in a third TERT-knockout embryonic stem cell line, 10PTKO-A, chromosome healing was a common event that accounted for 20 of 34 rearrangements. Although this chromosome healing also occurred at the I-SceI site, differences in the microhomology at the site of telomere addition demonstrated that the mechanism was distinct from that in wild-type embryonic stem cell lines. In addition, the newly added telomeres in 10PTKO-A shortened with time in culture, eventually resulting in either telomere elongation through a telomerase-independent mechanism or loss of the subtelomeric plasmid sequences entirely. The combined results demonstrate that chromosome healing can occur through both telomerase-dependent and -independent mechanisms, and that although both mechanisms can prevent degradation and sister chromatid fusion, neither mechanism is efficient enough to prevent sister chromatid fusion from occurring in many cells experiencing double-strand breaks near telomeres.

  View details for DOI 10.1016/j.dnarep.2008.04.004

  View details for Web of Science ID 000258259000006

  View details for PubMedID 18502190

  View details for PubMedCentralID PMC2597172

• Identification of ATPases pontin and reptin as telomerase components essential for holoenzyme assembly CELL Venteicher, A. S., Meng, Z., Mason, P. J., Veenstra, T. D., Artandi, S. E. 2008; 132 (6): 945-957

  Abstract

  Telomerase is a multisubunit ribonucleoprotein (RNP) complex that adds telomere repeats to the ends of chromosomes. Three essential telomerase components have been identified thus far: the telomerase reverse transcriptase (TERT), the telomerase RNA component (TERC), and the TERC-binding protein dyskerin. Few other proteins are known to be required for human telomerase function, limiting our understanding of both telomerase regulation and mechanisms of telomerase action. Here, we identify the ATPases pontin and reptin as telomerase components through affinity purification of TERT from human cells. Pontin interacts directly with both TERT and dyskerin, and the amount of TERT bound to pontin and reptin peaks in S phase, evidence for cell-cycle-dependent regulation of TERT. Depletion of pontin and reptin markedly impairs telomerase RNP accumulation, indicating an essential role in telomerase assembly. These findings reveal an unanticipated requirement for additional enzymes in telomerase biogenesis and suggest alternative approaches for inhibiting telomerase in cancer.

  View details for DOI 10.1016/j.cell.2008.01.019

  View details for Web of Science ID 000254273600014

  View details for PubMedID 18358808

• Telomere uncapping in progenitor cells with critical telomere shortening is coupled to S-phase progression in vivo PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rajaraman, S., Choi, J., Cheung, P., Beaudry, V., Moore, H., Artandi, S. E. 2007; 104 (45): 17747-17752

  Abstract

  Telomeres protect chromosome ends and serve as a substrate for telomerase, a reverse transcriptase that adds DNA repeats to the telomere terminus. In the absence of telomerase, telomeres progressively shorten, ultimately leading to telomere uncapping, a structural change at the telomere that activates DNA damage responses and leads to ligation of chromosome ends. Telomere uncapping has been implicated in aging and cancer, yet the precise mechanism of uncapping and its relationship to cell cycle remain to be defined. Here, we show that telomeres uncap in an S-phase-dependent manner in gastrointestinal progenitors of TERT(-/-) mice. We develop an in vivo assay that allows a quantitative kinetic assessment of telomere dysfunction-induced apoptosis and its relationship to cell cycle. By exploiting the mathematical relationship between rates of generation and clearance of apoptotic cells, we show that 86.2 +/- 8.8% of apoptotic gastrointestinal cells undergo programmed cell death either late in S-phase or in G2. Apoptosis is primarily triggered via a signaling cascade from newly uncapped telomeres to the tumor suppressor p53, rather than by chromosome fusion-bridge breakage, because mitotic blockade did not alter the rate of newly generated apoptotic bodies. These data support a model in which rapidly dividing progenitor cells within a tissue with short telomeres are vulnerable to telomere uncapping during or shortly after telomere replication.

  View details for DOI 10.1073/pnas.0706485104

  View details for Web of Science ID 000250897600038

  View details for PubMedID 17965232

  View details for PubMedCentralID PMC2077026

• DNA-dependent protein kinase catalytic subunit is not required for dysfunctional telomere fusion and checkpoint response in the telomerase-deficient mouse MOLECULAR AND CELLULAR BIOLOGY Maser, R. S., Wong, K., Sahin, E., Xia, H., Naylor, M., Hedberg, H. M., Artandi, S. E., DePinho, R. A. 2007; 27 (6): 2253-2265

  Abstract

  Telomeres are key structural elements for the protection and maintenance of linear chromosomes, and they function to prevent recognition of chromosomal ends as DNA double-stranded breaks. Loss of telomere capping function brought about by telomerase deficiency and gradual erosion of telomere ends or by experimental disruption of higher-order telomere structure culminates in the fusion of defective telomeres and/or the activation of DNA damage checkpoints. Previous work has implicated the nonhomologous end-joining (NHEJ) DNA repair pathway as a critical mediator of these biological processes. Here, employing the telomerase-deficient mouse model, we tested whether the NHEJ component DNA-dependent protein kinase catalytic subunit (DNA-PKcs) was required for fusion of eroded/dysfunctional telomere ends and the telomere checkpoint responses. In late-generation mTerc(-/-) DNA-PKcs(-/-) cells and tissues, chromosomal end-to-end fusions and anaphase bridges were readily evident. Notably, nullizygosity for DNA Ligase4 (Lig4)--an additional crucial NHEJ component--was also permissive for chromosome fusions in mTerc(-/-) cells, indicating that, in contrast to results seen with experimental disruption of telomere structure, telomere dysfunction in the context of gradual telomere erosion can engage additional DNA repair pathways. Furthermore, we found that DNA-PKcs deficiency does not reduce apoptosis, tissue atrophy, or p53 activation in late-generation mTerc(-/-) tissues but rather moderately exacerbates germ cell apoptosis and testicular degeneration. Thus, our studies indicate that the NHEJ components, DNA-PKcs and Lig4, are not required for fusion of critically shortened telomeric ends and that DNA-PKcs is not required for sensing and executing the telomere checkpoint response, findings consistent with the consensus view of the limited role of DNA-PKcs in DNA damage signaling in general.

  View details for DOI 10.1128/MCB.01354-06

  View details for Web of Science ID 000245003600023

  View details for PubMedID 17145779

• Aging, graying and loss of melanocyte stem cells STEM CELL REVIEWS Sarin, K. Y., Artandi, S. E. 2007; 3 (3): 212-217

  Abstract

  Hair graying is one of the prototypical signs of human aging. Maintenance of hair pigmentation is dependent on the presence and functionality of melanocytes, neural crest derived cells which synthesize pigment for growing hair. The melanocytes, themselves, are maintained by a small number of stem cells which reside in the bulge region of the hair follicle. The recent characterization of the melanocyte lineage during aging has significantly accelerated our understanding of how age-related changes in the melanocyte stem cell compartment contribute to hair graying. This review will discuss our current understanding of hair graying, drawing on evidence from human and mouse studies, and consider the contribution of melanocyte stem cells to this process. Furthermore, using the melanocyte lineage as an example, it will discuss common theories of tissue and stem cell aging.

  View details for DOI 10.1007/s12015-007-0028-0

  View details for Web of Science ID 000249929800004

  View details for PubMedID 17917134

• Telomeres, telomerase, and human disease. New England journal of medicine Artandi, S. E. 2006; 355 (12): 1195-1197

  View details for PubMedID 16990382

• Telomerase flies the coop: the telomerase RNA component as a viral-encoded oncogene JOURNAL OF EXPERIMENTAL MEDICINE Artandi, S. E. 2006; 203 (5): 1143-1145

  Abstract

  Telomerase, the enzyme that elongates our telomeres, is crucial for cancer development based on extensive analyses of human cells, human cancers, and mouse models. New data now suggest that a viral telomerase RNA gene encoded by Marek's disease virus (MDV), an oncogenic herpesvirus of chickens, promotes tumor formation. These findings highlight the importance of telomerase in cancer and raise new questions regarding the mechanisms by which the telomerase RNA component supports tumorigenesis.

  View details for DOI 10.1084/jem.20060849

  View details for Web of Science ID 000237803700003

  View details for PubMedID 16682501

• Regulation of cellular immortalization and steady-state levels of the telomerase reverse transcriptase through its carboxy-terminal domain MOLECULAR AND CELLULAR BIOLOGY Middleman, E. J., Choi, J. K., Venteicher, A. S., Cheung, P., Artandi, S. E. 2006; 26 (6): 2146-2159

  Abstract

  Telomerase maintains cell viability and chromosomal stability through the addition of telomere repeats to chromosome ends. The reactivation of telomerase through the upregulation of TERT, the telomerase protein subunit, is an important step during cancer development, yet TERT protein function and regulation remain incompletely understood. Despite its close sequence similarity to human TERT (hTERT), we find that mouse TERT (mTERT) does not immortalize primary human fibroblasts. Here we exploit these differences in activity to understand TERT protein function by creating chimeric mouse-human TERT proteins. Through the analysis of these chimeric TERT proteins, we find that sequences in the human carboxy-terminal domain are critical for telomere maintenance in human fibroblasts. The substitution of the human carboxy-terminal sequences into the mouse TERT protein is sufficient to confer immortalization and maintenance of telomere length and function. Strikingly, we find that hTERT protein accumulates to markedly higher levels than does mTERT protein and that the sequences governing this difference in protein regulation also reside in the carboxy-terminal domain. These elevated protein levels, which are characteristic of hTERT, are necessary but not sufficient for telomere maintenance because stabilized mTERT mutants cannot immortalize human cells. Thus, the TERT carboxy terminus contains sequences that regulate TERT protein levels and determinants that are required for productive action on telomere ends.

  View details for DOI 10.1128/MCB.26.6.2146-2159.2006

  View details for Web of Science ID 000235915400012

  View details for PubMedID 16507993

  View details for PubMedCentralID PMC1430280

• Conditional telomerase induction causes proliferation of hair follicle stem cells NATURE Sarin, K. Y., Cheung, P., Gilison, D., Lee, E., Tennen, R. I., Wang, E., Artandi, M. K., Oro, A. E., Artandi, S. E. 2005; 436 (7053): 1048-1052

  Abstract

  TERT, the protein component of telomerase, serves to maintain telomere function through the de novo addition of telomere repeats to chromosome ends, and is reactivated in 90% of human cancers. In normal tissues, TERT is expressed in stem cells and in progenitor cells, but its role in these compartments is not fully understood. Here we show that conditional transgenic induction of TERT in mouse skin epithelium causes a rapid transition from telogen (the resting phase of the hair follicle cycle) to anagen (the active phase), thereby facilitating robust hair growth. TERT overexpression promotes this developmental transition by causing proliferation of quiescent, multipotent stem cells in the hair follicle bulge region. This new function for TERT does not require the telomerase RNA component, which encodes the template for telomere addition, and therefore operates through a mechanism independent of its activity in synthesizing telomere repeats. These data indicate that, in addition to its established role in extending telomeres, TERT can promote proliferation of resting stem cells through a non-canonical pathway.

  View details for DOI 10.1038/nature03836

  View details for Web of Science ID 000231263900057

  View details for PubMedID 16107853

  View details for PubMedCentralID PMC1361120

• Pathways connecting telomeres and p53 in senescence, apoptosis, and cancer BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Artandi, S. E., Attardi, L. D. 2005; 331 (3): 881-890

  Abstract

  The ends of eukaryotic chromosomes are protected by specialized structures termed telomeres that serve in part to prevent the chromosome end from activating a DNA damage response. However, this important function for telomeres in chromosome end protection can be lost as telomeres shorten with cell division in culture or in self-renewing tissues with advancing age. Impaired telomere function leads to induction of a DNA damage response and activation of the tumor suppressor protein p53. p53 serves a critical role in enforcing both senescence and apoptotic responses to dysfunctional telomeres. Loss of p53 creates a permissive environment in which critically short telomeres are inappropriately joined to generate chromosomal end-to-end fusions. These fused chromosomes result in cycles of chromosome fusion-bridge-breakage, which can fuel cancer initiation, especially in epithelial tissues, by facilitating changes in gene copy number.

  View details for Web of Science ID 000229135900023

  View details for PubMedID 15865944

• Complex roles for telomeres and telomerase in breast carcinogenesis BREAST CANCER RESEARCH Artandi, S. E. 2003; 5 (1): 37-41

  Abstract

  Telomerase - an enzyme that endows cells with unlimited proliferative potential - is differentially expressed in cancer cells and in normal cells. Although most primary human cells lack telomerase, the enzyme is upregulated in more than 90% of invasive breast cancers. As a result, much of breast cancer development occurs before telomerase is reactivated during a critical transition from a telomerase-negative to a telomerase-positive state. During this transition, the telomere shortening that accompanies cell division may either prevent or facilitate tumorigenesis by activating checkpoints and impairing chromosomal stability. In mature cancers, telomerase probably serves a crucial role in tumor progression and maintenance by stabilizing telomeres and supporting the immortal growth of breast cancer cells.

  View details for DOI 10.1186/bcr553

  View details for Web of Science ID 000180046700010

  View details for PubMedID 12559044

• Constitutive telomerase expression promotes mammary carcinomas in aging mice PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Artandi, S. E., Alson, S., Tietze, M. K., Sharpless, N. E., Ye, S., Greenberg, R. A., Castrillon, D. H., Horner, J. W., Weiler, S. R., Carrasco, R. D., DePinho, R. A. 2002; 99 (12): 8191-8196

  Abstract

  Telomerase is up-regulated in the vast majority of human cancers and serves to halt the progressive telomere shortening that ultimately blocks would-be cancer cells from achieving a full malignant phenotype. In contrast to humans, the laboratory mouse possesses long telomeres and, even in early generation telomerase-deficient mice, the level of telomere reserve is sufficient to avert telomere-based checkpoint responses and to permit full malignant progression. These features in the mouse provide an opportunity to determine whether enforced high-level telomerase activity can serve functions that extend beyond its ability to sustain telomere length and function. Here, we report the generation and characterization of transgenic mice that express the catalytic subunit of telomerase (mTERT) at high levels in a broad variety of tissues. Expression of mTERT conferred increased telomerase enzymatic activity in several tissues, including mammary gland, splenocytes, and cultured mouse embryonic fibroblasts. In mouse embryonic fibroblasts, mTERT overexpression extended telomere lengths but did not prevent culture-induced replicative arrest, thus reinforcing the view that this phenomenon is not related to occult telomere shortening. Robust telomerase activity, however, was associated with the spontaneous development of mammary intraepithelial neoplasia and invasive mammary carcinomas in a significant proportion of aged females. These data indicate that enforced mTERT expression can promote the development of spontaneous cancers even in the setting of ample telomere reserve.

  View details for DOI 10.1073/pnas.112515399

  View details for Web of Science ID 000176217700071

  View details for PubMedID 12034875

• Telomere shortening and cell fates in mouse models of neoplasia TRENDS IN MOLECULAR MEDICINE Artandi, S. E. 2002; 8 (1): 44-47

  Abstract

  Cell division in the absence of telomerase leads to telomere shortening that can activate checkpoint responses and impair chromosomal stability. The absence of telomerase in primary human cells and its near universal reactivation in human cancers has highlighted the importance of telomere shortening and telomerase reactivation during tumor development. Data from telomerase-deficient mouse models of cancer have indicated that telomere shortening can exert profoundly different influences on cell fates in developing cancers, limiting tumorigenesis by enhancing cell death or facilitating carcinogenesis by compromising chromosomal stability. These alternate fates depend on the integrity of the p53 pathway and on cell type.

  View details for Web of Science ID 000173187000008

  View details for PubMedID 11796266

• Telomere dysfunction promotes non-reciprocal translocations and epithelial cancers in mice NATURE Artandi, S. E., Chang, S., Lee, S. L., Alson, S., Gottlieb, G. J., Chin, L., DePinho, R. A. 2000; 406 (6796): 641-645

  Abstract

  Aged humans sustain a high rate of epithelial cancers such as carcinomas of the breast and colon, whereas mice carrying common tumour suppressor gene mutations typically develop soft tissue sarcomas and lymphomas. Among the many factors that may contribute to this species variance are differences in telomere length and regulation. Telomeres comprise the nucleoprotein complexes that cap the ends of eukaryotic chromosomes and are maintained by the reverse transcriptase, telomerase. In human cells, insufficient levels of telomerase lead to telomere attrition with cell division in culture and possibly with ageing and tumorigenesis in vivo. In contrast, critical reduction in telomere length is not observed in the mouse owing to promiscuous telomerase expression and long telomeres. Here we provide evidence that telomere attrition in ageing telomerase-deficient p53 mutant mice promotes the development of epithelial cancers by a process of fusion-bridge breakage that leads to the formation of complex non-reciprocal translocations--a classical cytogenetic feature of human carcinomas. Our data suggest a model in which telomere dysfunction brought about by continual epithelial renewal during life generates the massive ploidy changes associated with the development of epithelial cancers.

  View details for Web of Science ID 000088653800051

  View details for PubMedID 10949306

• Mice without telomerase: what can they teach us about human cancer? NATURE MEDICINE Artandi, S. E., DePinho, R. A. 2000; 6 (8): 852-855

  Abstract

  Unicellular organisms, human cells and mice have provided insights into the processes of senescence, crisis, genomic instability and cancer in humans. Here, Artandi and DePinho discuss how studies in mice have uncovered a complex interplay between the ARF-p53 pathway, genomic instability due to telomere dysfunction, and the suppression or promotion of cancer.

  View details for Web of Science ID 000165473800015

  View details for PubMedID 10932211

• A critical role for telomeres in suppressing and facilitating carcinogenesis CURRENT OPINION IN GENETICS & DEVELOPMENT Artandi, S. E., DePinho, R. A. 2000; 10 (1): 39-46

  Abstract

  Progressive telomere shortening occurs with the division of primary human cells and activates tumor suppressor pathways, triggering senescence and inhibiting tumorigenesis. Loss of p53 function, however, allows continued cell division despite increasing telomere dysfunction and entry into telomere crisis. Recent data suggest that the severe chromosomal instability of telomere crisis promotes secondary genetic changes that facilitate carcinogenesis. Reactivation of telomerase stabilizes telomere ends and allows continued tumor growth.

  View details for Web of Science ID 000085533800005

  View details for PubMedID 10679392

• p53 deficiency rescues the adverse effects of telomere loss and cooperates with telomere dysfunction to accelerate carcinogenesis CELL Chin, L., Artandi, S. E., Shen, Q., Tam, A., Lee, S. L., Gottlieb, G. J., Greider, C. W., DePinho, R. A. 1999; 97 (4): 527-538

  Abstract

  Maintenance of telomere length and function is critical for the efficient proliferation of eukaryotic cells. Here, we examine the interactions between telomere dysfunction and p53 in cells and organs of telomerase-deficient mice. Coincident with severe telomere shortening and associated genomic instability, p53 is activated, leading to growth arrest and/or apoptosis. Deletion of p53 significantly attenuated the adverse cellular and organismal effects of telomere dysfunction, but only during the earliest stages of genetic crisis. Correspondingly, the loss of telomere function and p53 deficiency cooperated to initiate the transformation process. Together, these studies establish a key role for p53 in the cellular response to telomere dysfunction in both normal and neoplastic cells, question the significance of crisis as a tumor suppressor mechanism, and identify a biologically relevant stage of advanced crisis, termed genetic catastrophe.

  View details for Web of Science ID 000080299100013

  View details for PubMedID 10338216