Browsing by Subject "Telomeric Repeat Binding Protein 2"
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Item Open Access Distinct functions of POT1 at telomeres.(2008) Kendellen, Megan FullerTelomeres are nucleoprotein complexes that constitute the ends of eukaryotic chromosomes. Telomeres differentiate the end of the chromosome from sites of DNA damage and control cellular replicative potential. The loss of function of telomeres results in several biological consequences. First, dysfunctional telomeres elicit DNA damage responses and repair activities, which frequently induce cytogenetic abnormalities and genomic instability that are characteristic of human cancer. Second, cellular immortalization resulting from inappropriate elongation of telomeres is a critical component of tumorigenesis. Alternatively, as telomere shortening limits replicative potential, abnormally short telomeres can result in premature cellular senescence that is associated with human pathology ranging from anemia to atherosclerosis. Telomeric DNA is composed of tandem repeats of G‐rich double‐stranded (ds)DNA that terminates in a G‐rich 3’ single‐stranded (ss)DNA overhang. Telomeres are thought to assume a lariat structure termed the t‐loop, which is decorated by an assortment of telomere‐associated proteins. The most unique and least well characterized of these proteins is POT1. POT1 binds telomeric ssDNA via a pair of Nterminal OB‐folds. Through its C‐terminal protein‐interaction domain, POT1 directly binds the telomeric dsDNA‐binding protein TRF2 and participates in heterodimeric complex with the protein TPP1. Inhibition of POT1 induces a robust DNA damage response at telomeres and deregulation of telomere length homeostasis, indicating that POT1 is important in maintaining telomere stability and in regulating telomere length. The goal of my thesis work was to determine which of the three major functions of POT1– telomeric ssDNA‐, TPP1‐, or TRF2‐binding – were required to properly localize POT1 to telomeres and to prevent the telomere instability and length deregulation that occur in the absence of POT1. Using separation‐of‐function mutants of POT1 deficient in at least one of these activities, I found that POT1 depends on its heterodimeric partner TPP1 in cis with telomeric ssDNA‐binding to preserve telomere stability, while POT1 depends on its protein interaction with TRF2 to localize to telomeres and its TRF2‐ and telomeric ssDNA‐binding activities in cis to regulate telomere length.Item Open Access Nucleolar organization, ribosomal DNA array stability, and acrocentric chromosome integrity are linked to telomere function.(PLoS One, 2014) Stimpson, Kaitlin M; Sullivan, Lori L; Kuo, Molly E; Sullivan, Beth AThe short arms of the ten acrocentric human chromosomes share several repetitive DNAs, including ribosomal RNA genes (rDNA). The rDNA arrays correspond to nucleolar organizing regions that coalesce each cell cycle to form the nucleolus. Telomere disruption by expressing a mutant version of telomere binding protein TRF2 (dnTRF2) causes non-random acrocentric fusions, as well as large-scale nucleolar defects. The mechanisms responsible for acrocentric chromosome sensitivity to dysfunctional telomeres are unclear. In this study, we show that TRF2 normally associates with the nucleolus and rDNA. However, when telomeres are crippled by dnTRF2 or RNAi knockdown of TRF2, gross nucleolar and chromosomal changes occur. We used the controllable dnTRF2 system to precisely dissect the timing and progression of nucleolar and chromosomal instability induced by telomere dysfunction, demonstrating that nucleolar changes precede the DNA damage and morphological changes that occur at acrocentric short arms. The rDNA repeat arrays on the short arms decondense, and are coated by RNA polymerase I transcription binding factor UBF, physically linking acrocentrics to one another as they become fusogenic. These results highlight the importance of telomere function in nucleolar stability and structural integrity of acrocentric chromosomes, particularly the rDNA arrays. Telomeric stress is widely accepted to cause DNA damage at chromosome ends, but our findings suggest that it also disrupts chromosome structure beyond the telomere region, specifically within the rDNA arrays located on acrocentric chromosomes. These results have relevance for Robertsonian translocation formation in humans and mechanisms by which acrocentric-acrocentric fusions are promoted by DNA damage and repair.