**2. Telomere construction**

The basic structure of telomere is conserved among eukaryotes and consists of short tandem DNA repeats, with G-rich sequence at the three-end referred to as the G-strand and the complementary strand is called the C-strand at the five-end (**Figure 1**) [3]. The length of telomeric DNA varies from 2 to 20 kilobase pairs, depending on factors such as tissue type and human age [4]. The telomere is conspicuous owing to the presence of G-overhang which extends beyond its complementary C-rich strand to form a single-stranded overhang, termed the G-tail. It has been proposed that the 3′ G-overhang can be sequestered into a lasso-like structure known as the T-loop (**Figure 2**) [5, 6]. The single-stranded G-overhang invades the double-stranded telomeric DNA, which displaces the bound G-strand base-pairing with the C-strand. As this displaced binding takes place at a distance from the physical end of the telomere, it generates a large duplex structure called the T-loop (**Figure 2**) [3, 5].

**Figure 1.** Telomeric DNA.

Telomeres can also fold into G-quadruplex DNA, an unusual DNA conformation that is based on a guanine quartet [7]. The repetitive and GC-rich nature of telomeric DNA endows it with the capability to form the higher-order DNA conformation, G-quadruplexes [8, 9]. To maintain such unusual structure of telomeres, a set of telomeric protein complex has evolved, termed shelterin. The shelterin complex consists of six individual proteins, telomeric repeat binding factor 1 (TRF1), TRF2, repressor/activator protein 1 (RAP1), TIN2 (TRF1 interacting protein 2), TPP1 (TINT1/PIP1/PTOP 1), and protection of telomeres 1 (POT1) [6, 10]. The proteins TRF1 and TRF2 attach to double-stranded telomeric repeats facilitating the anchoring of the complex along the length of telomeres [11–14], whereas POT1 binds to the single-stranded overhang. TPP1 and TIN2 act as bridging proteins between the above DNA-binding modules and are crucial for chromosome end protection and telomere length regulation. TRF1 and TRF2 with the help of TIN2 [15] bind with POT1 via TPP1 [16–19]. TIN2 also connects TRF1 to TRF2, and this interaction contributes to the stabilization of TRF2 on telomeres [17–19]. Besides this shelterin complex, other proteins like TEN1 and Pinx1, which are not telomere specific, are also present at telomeres and carry out important functions in recruitment of telomerase [20].

**Figure 2.** T-loop formation.

regarding the telomere length are unanswered till now. Till date it is unclear about the mechanistic correlation between telomere size and life span of various species. To prevent senescence of cell, the preservation of telomere length is essential, which is maintained by telomerase enzyme. Telomerase is a reverse transcriptase enzyme which has recently emerged as an attractive target for cancer as it is a crucial factor required for the tumor immortalization of a subset of cells, including cancer stem cells. Studies have proved that 80–85% of the tumor cells express telomerase whereas somatic cells lack the expression of telomerase [1]. The important paradox is that telomerase-negative normal cells have lengthier telomeres than telomerase-positive cancer cells [2]. Thus difference in telomere length and cell kinetics between normal and cancerous cells shows that targeting telomerase is a more effective way to target cancer cells. Owing to the significant role of telomere and telomerase in tumor immortalization understanding their amassed influence on cancer is crucial for cancer therapy. In this context, this book chapter is focused on disseminating the integrated impact of telomere and telomerase on cancer progres-

The basic structure of telomere is conserved among eukaryotes and consists of short tandem DNA repeats, with G-rich sequence at the three-end referred to as the G-strand and the complementary strand is called the C-strand at the five-end (**Figure 1**) [3]. The length of telomeric DNA varies from 2 to 20 kilobase pairs, depending on factors such as tissue type and human age [4]. The telomere is conspicuous owing to the presence of G-overhang which extends beyond its complementary C-rich strand to form a single-stranded overhang, termed the G-tail. It has been proposed that the 3′ G-overhang can be sequestered into a lasso-like structure known as the T-loop (**Figure 2**) [5, 6]. The single-stranded G-overhang invades the double-stranded telomeric DNA, which displaces the bound G-strand base-pairing with the C-strand. As this displaced binding takes place at a distance from the physical end of the

Telomeres can also fold into G-quadruplex DNA, an unusual DNA conformation that is based on a guanine quartet [7]. The repetitive and GC-rich nature of telomeric DNA endows it with the capability to form the higher-order DNA conformation, G-quadruplexes [8, 9]. To maintain such unusual structure of telomeres, a set of telomeric protein complex has evolved, termed shelterin. The shelterin complex consists of six individual proteins, telomeric repeat binding factor 1 (TRF1), TRF2, repressor/activator protein 1 (RAP1), TIN2 (TRF1 interacting protein 2),

telomere, it generates a large duplex structure called the T-loop (**Figure 2**) [3, 5].

sion.

**2. Telomere construction**

18 Telomere - A Complex End of a Chromosome

**Figure 1.** Telomeric DNA.

**Figure 3.** Telomere-telomerase assembly.

#### **3. Telomere: a knight cap**

During the evolvement of linear genome, the natural ends of linear chromosomes resemble DNA breaks and tend to induce DNA damage response (DDR). These natural linear ends are protected by the sequestration of the ribonucleoprotein (RNP) sequence, telomeres which mask the ends from continuous exposure to the DNA damage response (DDR). Telomeres serve as protective caps, preventing the chromosomal ends from being recognized as doublestrand breaks by the DNA damage repair system and the activation of the p53 or p16INK4a pathway and the start of senescence or apoptosis. If the telomere cap is removed, genome instability is induced. Telomeres with its tightly regulated complexes consisting of repetitive G-rich DNA and specialized proteins accomplish the task of not only concealing the linear chromosome ends from detection and undesired repair, but also protect from checkpoints, homologous recombination, end-to-end fusions, or other events that normally promote repair of intra-chromosomal DNA breaks acts [21]. Telomeric proteins and their interacting factors create an environment at chromosome ends that inhibits DNA repair at that point; however, the repair machinery is also essential for proper telomere function.

The closed configuration of the T-loop of telomeric region provides a protective cap that defines the natural end of the chromosome and masks the telomere from the DDR machinery (**Figure 2**) [6]. In particular, T-loops could provide an architectural solution to the repression of the ataxia telangiectasia mutated (ATM) kinase pathway, which relies on a sensor (the MRN (Mre11/Rad50/Nbs1) complex) with DNA end-binding activity. In addition, T-loops could prevent the Ku70/80 heterodimer, a DNA repair factor that binds to DNA ends, from loading onto the telomere terminus, thereby blocking the initiation of the non-homologous end-joining (NHEJ) pathway (**Figure 4**).

**Figure 4.** DNA damage response pathway.

Among the DNA-binding proteins, TRF1 has DNA remodeling activity [5, 22] and also shown to promote efficient replication of telomeres [23, 24]. On the contrary, TRF2 engages in chromosome end protection by inducing topological changes in telomeric DNA [25], T-loop assembly [26, 27] and by suppression of ATM dependent DDR and NHEJ (**Figure 4**) [28, 29]. Besides that, TRF2 plays a critical role in chromatin assembly, which was demonstrated by the observation that overexpression of TRF2 resulted in aberrant nucleosome spacing and decreased abundance of the core histones H3 and H4 at chromosome ends [30]. TRF2 lacking cells are reported to be growth arrested because of up-regulation of p53 and also show other hallmarks of ATM signaling, including the phosphorylation of ATM and Chk2 [31, 32]. Among the two DDR, the ATM kinase pathway at telomeres is repressed by TRF2 subunit, whereas POT1 is responsible for protection of telomere by suppression of ATR (ATM Rad3-related protein)-dependant DDR pathways [28, 33].

The high affinity of POT1 for single-stranded telomeric DNA makes it a G-strand binding component displaced from the T-loop and forms a closed configuration locking in the structure (**Figure 2**). Earlier report models suggest that POT1 and TPP1 compete with telomerase for access to the overhang [33]. Contrarily, direct interaction between TPP1 and telomerase bolsters telomerase processivity [34, 35] whereas increased loading of POT1 along the overhang block telomerase accessibility to the 3′-OH substrate. The role of RAP1 is obscure and its function has recently been elucidated. The presence of RAP1 at telomeres appears as a backup mechanism to prevent NHEJ when topology-mediated telomere protection is impaired [36]. In case of mutation in TRF2 which wraps the DNA, RAP1 has been implicated in the inhibition of NHEJ [37, 38].

Thus physically taken together, the shelterin complex and G-overhang of telomere are the protective cap that have an immensely complex role in convergence of end protection and telomere length maintenance mechanisms.
