Telomeric repeats

The most striking and recent example of structural complexity is the intramolecular human telomeric motif. In sodium, the AG3(T2AG3)3 sequence adopts an intramolecular fold with a central diagonal loop. As a result, each strand has one parallel and one antiparallel neighbour. In potassium, the situation is different. Crystallographic studies indicate that the very same sequence adopts a completely different folding scheme all four DNA strands are parallel, with the three linking trinucleotide loops positioned on the exterior 

Telomeric repeats from other organisms are also prone to quadruplex polymorphism. The Tetrahymena TG4T2G4T telomeric sequence, for example, may adopt several conformations. This sequence forms two novel G-quadruplex structures in Na -containing solution. In the first structure (head-to-head), the two loops are at one end of the G-tetrad core in the second structure (head-to-tail), the two loops are located on opposite ends of the G-tetrad core. In contrast to the human telomere sequence, the proportions of the two forms are similar for a wide range of temperatures their unfolding rates are also similar, with an activation enthalpy of 37 kcal mol . The (G4T4) sequence may also interconvert between parallel and antiparallel structures. 

Low concentrations of either Na or K facilitate formation of the antiparallel quadruplex, while high cation concentration promotes formation of the parallel 

---

Gavathiotis E et al. (2003) Drug recognition and stabilisation of the parallel-stranded DNA quadruplex d(TTAGGGT)4 containing the human telomeric repeat. J Mol Biol 334(l) 25-36... 

At last, nucleolin might play a specific role in telomeric replication and maintenance, as suggested by two types of data. First, it binds telomeric repeat (TTAGGG)n in vitro (Ishikawa et al, 1993 Pollice et al, 2000), with a marked preference for the single-stranded form. Secondly, it interacts in vitro and in vivo with hTERT (Khurts et al, 2004), the protein catalytic component of human telom-erase. This interaction takes place both in the cytoplasm and in the nucleolus, where it could promote the assembly of hTERT with the RNA subunit hTERC. As a conclusion, many data regarding the involvement of nucleolin in DNA replication are indirect and an experimental demonstration through knockdown or knockout studies is still awaited. 

Although the existence of this enzyme may not be surprising, the mechanism by which it acts is remarkable and unprecedented. Telomerase, like some other enzymes described in this chapter, contains both RNA and protein components. The RNA component is about 150 nucleotides long and contains about 1.5 copies of the appropriate CyKx telomere repeat. This region of the RNA acts as a template for synthesis of the T -G strand of the telomere. Telomerase thereby acts as a cellular reverse transcriptase that provides the active site for RNA-dependent DNA synthesis. Unlike retroviral reverse transcriptases, telomerase copies only a small segment of RNA that it carries within itself. Telomere synthesis requires the 3 end of a chromosome as primer and proceeds in the usual 5 —>3 direction. Having syn-... 

Telomerase is a reverse transcriptase that copies the DNA sequence of the telomeric repeats from a small guide RNA that is part of the enzyme. Tire first telomerase studied was the relatively abundant enzyme from Tetrahymena. It contains a 159-nucleotide RNA with the sequence 5 -CAACCCCAA-3 at positions 43-51. This sequence is complementary to the 5 -TTGGGG-3 repeat sequence of the Tetrahymena telomeres.450 451 A 127-kDa human protein contains a similar guide RNA with the sequence 5 -CUAACCCUACC-3, which is complementary to the human telomere repeat sequence as is illustrated in Fig. 27-21.452-454 Telomerases455 456 evidently allow the cell to elongate the telomere 5 -ends using the... 

Joffe, B.I., Solovei, I.V. and Macgregor, H.C. (1 996) Ends of chromosomes in Polycelis tenuis (Platyhelminthes) have telomere repeat TTAGGG. Chromosome Research 4, 323-324. 

Greider, C. W., and Blackburn, E. H. (1989). A telomeric sequence in the RNA of Tetrahymena telomerase required for telomere repeat synthesis. Nature337, 331-337. 

The results from Zhou et al. revealed that the introduction of electron-donating groups such as substituted amino groups at the C-ll position of the quindoline significantly enhanced the ability of the molecule to inhibit telomerase activity (IC50 > 138 iM for quindoline, 0.44-12.3 iM for quindoline derivatives 1-10). The quindoline derivatives not only stabilized the G-quadruplex structure but also induced the G-rich telomeric repeated DNA sequence to fold into a quadruplex [31]. 

Figure 12.9 (A) Schematic illustration of overall assay design for ZnO NR-based telomeric repeat elongation assays. TS is an oligonucleotide whose sequence is recognized by telomerase. (B) Fluorescence panel obtained from positive (1) versus negative (2) samples after performing telomeric repeat elongation assays on ZnO NR stripe platforms.

Dorfman, A., Parajuli, O., Kumar, N., and Hahm, J. (2008). Novel telomeric repeat elongation assay performed on ZnO nanorod array supports. J. Nanosci. Nanotech. 8 410-415. 

The length of the telomere repeats at individual chromosome ends influences biological functions that range from aging to carcinogenesis (9). Measurements of the telomere length can be... 

Wang Y, Patel DJ. 1993. Solution structure of the human telomeric repeat d[AG3(T2AG3)3] G-tetraplex. Structure 1 263-82... 

The TRAP (telomeric repeat amplification protocol) assay is a widely used method for detection of telomerase activity. This technique measures the telomerase activity present in cell extracts. Briefly, cellular extract containing telomerase activity is incubated with a telomeric substrate (a short strand of DNA onto which the telomerase wiU. attach the telomeric repeats) followed by polymerase chain reaction (PCR) amplification of the elongated telomere. Detection of the PCR product is by a number of methods, including gel electrophoresis, radiometric detection, ELISA, and real-time PCR detection. ... 

Sequencing of telomeres from a dozen or so organisms, including humans, has shown that most are repetitive oligomers with a high C content in the strand with its 3 end at the end of the chromosome. The telomere repeat sequence In humans and other vertebrates is TTACCC. These simple sequences are repeated at the very termini of chromosomes for a total of a few hundred base pairs in yeasts and protozoans and a few thousand base pairs in vertebrates. The 3 end of the C-rich strand extends 12-16 nucleotides beyond the 5 end of the complementary C-rich strand. This region is bound by specific proteins that both protect the ends of... 

---