Elizabeth H. Blackburn

Identification of a specific telomere terminal transferase activity in Tetrahymena extracts.

We have found a novel activity in Tetrahymena cell free extracts that adds tandem TTGGGG repeats onto synthetic telomere primers. The single-stranded DNA oligonucleotides (TTGGGG)4 and TGTGTGGGTGTGTGGGTGTGTGGG, consisting of the Tetrahymena and yeast telomeric sequences respectively, each functioned as primers for elongation, while (CCCCAA)4 and two nontelomeric sequence DNA oligomers did not. Efficient synthesis of the TTGGGG repeats depended only on addition of micromolar concentrations of oligomer primer, dGTP, and dTTP to the extract. The activity was sensitive to heat and proteinase K treatment. The repeat addition was independent of both endogenous Tetrahymena DNA and the endogenous alpha-type DNA polymerase; and a greater elongation activity was present during macronuclear development, when a large number of telomeres are formed and replicated, than during vegetative cell growth. We propose that the novel telomere terminal transferase is involved in the addition of telomeric repeats necessary for the replication of chromosome ends in eukaryotes.

Switching and Signaling at the Telomere

This review describes the structure of telomeres, the protective DNA-protein complexes at eukaryotic chromosomal ends, and several molecular mechanisms involved in telomere functions. Also discussed are cellular responses to compromising the functions of telomeres and of telomerase, which synthesizes telomeric DNA.

Telomere states and cell fates

Telomere length has frequently been used as a means to predict the future life of cells. But by itself it can be a poor indicator of ageing or cell viability. What, then, is the important property of a telomere? Here recent findings are integrated into a new, probabilistic view of the telomere to explain how and when it can signal not only its own fate but also that of a cell.

An alternative pathway for yeast telomere maintenance rescues est1− senescence

Yeast cells lacking a functional EST1 gene show progressive shortening of the terminal G1-3T telomeric repeats and a parallel increase in the frequency of cell death. Although the majority of the cells in an est1- culture die, a minor subpopulation survives the potentially lethal consequences of the est1 mutation. We show that these est1- survivors arise as a result of the amplification and acquisition of subtelomeric elements (and their deletion derivatives) by a large number of telomeres. Hence, even when the primary pathway for telomere replication is defective, an alternative backup pathway can restore telomere function and keep the cell alive.

A tandemly repeated sequence at the termini of the extrachromosomal ribosomal RNA genes in Tetrahymena

Abstract The extrachromosomal genes coding for ribosomal RNA (rDNA) in the ciliated protozoan Tetrahymena thermophila were studied with respect to sequences occurring at their termini. The linear rDNA molecules had previously been shown to be palindromic (Karrer and Gall, 1976; Engberg et al. , 1976). Within the terminal rDNA fragment produced by restriction endonuclease digestion, a tandomly repeated hexanucleotide sequence 5′ (C-C-C-C-A-A) n 3′ was found, where n is between 20 and 70. This fragment was heterogeneous in length as judged by gel electrophoresis. The repeating sequence was preferentially synthesized when rDNA was used as the template by Escherichia coli DNA polymerase I. Initiation occurred at specific single-strand discontinuities, probably one-nucleotide gaps, found every few repeats on the C-C-C-C-A-A strand. At least one discontinuity is present on the G-G-G-G-T-T strand. Experiments with T4 DNA polymerase suggested that there are no free cohesive ends on the rDNA of the kind found in bacteriophage λ DNA. The orientation of the strands carrying the repeating hexanucleotide sequence was determined, and a model for the termini of the rDNA based on these findings is presented.

Telomeres and telomerase: the path from maize, Tetrahymena and yeast to human cancer and aging

Telomeres and telomerase: the path from maize, Tetrahymena and yeast to human cancer and aging

Telomeres and telomerase: their mechanisms of action and the effects of altering their functions

The molecular features of telomeres and telomerase are conserved among most eukaryotes. How telomerase and telomeres function and how they interact to promote the chromosome‐stabilizing properties of telomeres are discussed here.

The telomere syndromes.

There has been mounting evidence of a causal role for telomere dysfunction in a number of degenerative disorders. Their manifestations encompass common disease states such as idiopathic pulmonary fibrosis and bone marrow failure. Although these disorders seem to be clinically diverse, collectively they comprise a single syndrome spectrum defined by the short telomere defect. Here we review the manifestations and unique genetics of telomere syndromes. We also discuss their underlying molecular mechanisms and significance for understanding common age-related disease processes.

Telomeric DNA oligonucleotides form novel intramolecular structures containing guanine·guanine base pairs

Structural properties of DNA oligonucleotides corresponding to the single-stranded molecular terminus of telomeres from several organisms were analyzed. Based on physical studies including nondenaturing polyacrylamide gel electrophoresis, absorbance thermal denaturation analysis, and 1H and 31P nuclear magnetic resonance spectroscopy, we conclude that these molecules can self-associate by forming non-Watson-Crick, guanine.guanine based-paired, intramolecular structures. These structures form below 40 degrees C at moderate ionic strength and neutral pH and behave like hairpin duplexes in nondenaturing polyacrylamide gels. Detailed analysis of the hairpin structure formed by the telomeric sequence from Tetrahymena, (T2G4)4, shows that it is a unique structure stabilized by hydrogen bonds and contains G residues in the syn conformation. We propose that this novel form of DNA is important for telomere function and sets a precedent for the biological relevance of non-Watson-Crick base-paired DNA structures.

Cloning yeast telomeres on linear plasmid vectors

We have constructed a linear yeast plasmid by joining fragments from the termini of Tetrahymena ribosomal DNA to a yeast vector. Structural features of the terminus region of the Tetrahymena rDNA plasmid maintained in the yeast linear plasmid include a set of specifically placed single-strand interruptions within the cluster of hexanucleotide (C4A2) repeat units. An artificially constructed hairpin terminus was unable to stabilize a linear plasmid in yeast. The fact that yeast can recognize and use DNA ends from the distantly related organism Tetrahymena suggests that the structural features required for telomere replication and resolution have been highly conserved in evolution. The linear plasmid was used as a vector to clone chromosomal telomeres from yeast. One Tetrahymena end was removed by restriction digestion, and yeast fragments that could function as an end on a linear plasmid were selected. Restriction mapping and hybridization analysis demonstrated that these fragments were yeast telomeres, and suggested that all yeast chromosomes might have a common telomere sequence. Yeast telomeres appear to be similar in structure to the rDNA of Tetrahymena, in which specific nicks or gaps are present within a simple repeated sequence near the terminus of the DNA.