To better understand the role of telomeres and telomerase in cancer, it is important to analyze telomere behavior, the consequences of telomere malfunctioning, and telomerase activation and inactivation in cancer cells. We recently began to probe telomere dynamics in living cells using a newly available, high time-resolution light microscopy system, and found novel classes of telomere movements in live human cells.
Aim 1 will analyze such real-time telomere dynamics over short times (seconds), quantifying time- resolved 3D movements of telomeres in live cells, to analyze responses of telomere dynamics to telomere perturbations, in cancerous and normal human cells. We have previously designed and tested mutant- template hTers (MT-hTer) that force the highly active telomerase (characteristic of cancer cells) to add mutant sequence repeats to telomeres, which induced a rapid uncapping of telomeres, and elicited cellular responses, including apoptosis. These effects were rapid, telomere length-independent and do not require functional p53 or Rb.
Aim 2 will determine the mechanisms and players at the telomeres that, upon telomere uncapping, initiate the signaling response that ultimately ends in apoptosis. Knocking down the nigh telomerase in cancer cells also quickly inhibited their growth, eliciting distinct cellular and transcriptional changes. These and other recent results have indicated that telomerase likely plays roles in other aspects of cancer known to be central to cancer progression. The distinctive alterations in the gene-expression profiles upon telomerase RNA knockdown were predicted to be associated with diminished cancer progression.
Aim 3 will test which aspect(s) of telomerase/telomeres when lost/altered cause the cellular response to telomerase RNA knockdown, and also analyze the cellular and metabolic responses of human melanoma cells to telomerase depletion. By using the single telomere length analysis (STELA) method, we have discovered a novel class of ultra-short telomeres (""""""""t-stumps"""""""") in cancer cells.
In Aim 4 we will pursue further structural analysis of t-stumps, determine the dependence of t-stumps on telomerase and checkpoint pathways, and test the hypothesis that t-stumps, by signaling cells in a telomerase-'specific fashion, may underlie the rapid cellular effects of telomerase knockdown. Significance: Much previous evidence has pointed to telomerase promoting tumor maintenance and growth, and telomerase has been proposed as a target for anti-cancer therapies. Our work will advance the basic understanding of cancer telomere biology, which will be important to develop novel therapeutic strategies to exploit the unique telomerase status of cancer cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA096840-10
Application #
8114109
Study Section
Cancer Molecular Pathobiology Study Section (CAMP)
Program Officer
Pelroy, Richard
Project Start
2002-09-01
Project End
2012-08-07
Budget Start
2011-08-01
Budget End
2012-08-07
Support Year
10
Fiscal Year
2011
Total Cost
$298,913
Indirect Cost
Name
University of California San Francisco
Department
Biochemistry
Type
Schools of Medicine
DUNS #
094878337
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Blackburn, Elizabeth H; Epel, Elissa S; Lin, Jue (2015) Human telomere biology: A contributory and interactive factor in aging, disease risks, and protection. Science 350:1193-8
Gazzaniga, Francesca S; Blackburn, Elizabeth H (2014) An antiapoptotic role for telomerase RNA in human immune cells independent of telomere integrity or telomerase enzymatic activity. Blood 124:3675-84
Listerman, Imke; Gazzaniga, Francesca S; Blackburn, Elizabeth H (2014) An investigation of the effects of the core protein telomerase reverse transcriptase on Wnt signaling in breast cancer cells. Mol Cell Biol 34:280-9
Belin, Brittany J; Cimini, Beth A; Blackburn, Elizabeth H et al. (2013) Visualization of actin filaments and monomers in somatic cell nuclei. Mol Biol Cell 24:982-94
Chen, Baohui; Gilbert, Luke A; Cimini, Beth A et al. (2013) Dynamic imaging of genomic loci in living human cells by an optimized CRISPR/Cas system. Cell 155:1479-91
Listerman, Imke; Sun, Jie; Gazzaniga, Francesca S et al. (2013) The major reverse transcriptase-incompetent splice variant of the human telomerase protein inhibits telomerase activity but protects from apoptosis. Cancer Res 73:2817-28
Blackburn, Elizabeth H (2011) Cancer interception. Cancer Prev Res (Phila) 4:787-92
Blackburn, Elizabeth H (2011) Walking the walk from genes through telomere maintenance to cancer risk. Cancer Prev Res (Phila) 4:473-5
Stohr, Bradley A; Xu, Lifeng; Blackburn, Elizabeth H (2010) The terminal telomeric DNA sequence determines the mechanism of dysfunctional telomere fusion. Mol Cell 39:307-14
Blackburn, Elizabeth H (2010) Telomeres and telomerase: the means to the end (Nobel lecture). Angew Chem Int Ed Engl 49:7405-21

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