Structure and Synthesis of Telomere

In this article we will discuss about the structure and synthesis of telomere.

Structure of Telomere:

Molecular genetic studies have shown that the telomere consists of several short sequences which are tandemly repeated (Table 8.8). The general formula for the composition of the repeating units in one strand of the telomeres of all the eukaryotes is as follows:

5′ C_n (A/T)_m3′ …(8.4)

where n is greater than 1 and m varies from 1 to 4. The other strand has its complementary sequence.

There exist single strand breaks in the telomeric region. The terminal part of the telomere is folded in some specific way so that neither ligase can seal it nor nucleases can degrade it. Some specific proteins are also bound to the telomeres and thereby afford additional protection.

Although one strand (G-rich strand) of the telomeric DNA is long and single stranded (unpaired), it forms a duplex structure through folding back upon itself. Two models have been suggested for the folding mechanisms at the telomeric ends.

1. Four G-rich repeating units (5′ TTGGGG3′) are involved in forming the duplex struc­ture. Two of the repeating units fold back on the other two to form a hairpin like structure within which G : G pairing occurs. (Fig. 8.9).

2. According to the alternative model, proposed by Williamson and coworkers in 1989, four repeating units (5′ TTGGGG 3′) fold in such a way that the second G of each unit acts as a member of a “quartet”. Rest of the nucleotides form a loop (Fig. 8.9). This model is known as G quartet model.

Synthesis of Telomeric DNA:

Synthesis of the telomeric repeating sequences has been well understood in Tetrahymena. In this species an enzymes called telomerase was discovered in 1987 by Greider and Blackburn; this enzyme synthesize the telomeric DNA. The enzyme telomerase contains a short chain of single- stranded RNA made of 159 ribonucleotides; thus the enzyme is a ribonucleoprotein.

The enzymatic RNA includes a 15-22 base long sequence which is similar to the C-rich sequence (3′ AACCCC 5′) (Table 8.8) and it is complementary to the G-rich sequence (5′ TTGGGG 3′) of the telomere. This RNA sequence functions as a template for the synthesis of the G-rich sequence (Fig. 8.10).

Thus the enzyme telomerase functions like “reverse transcriptase” During synthesis, the nucleotide G or T is added one-by-one at the 3′-OH end of the G-rich DNA primer. When the whole sequence is synthesized, a new cycle of DNA synthesis starts through the movement of the enzyme at the new 3′-OH end.

The length of the telomeric DNA increases by this process of synthesis. However, the telomeric length is controlled by some regulatory mechanism.

Yeast Artificial Chromosome (YAC):

A chromosome requires three necessary components for its existence:

(i) Telomeres to seal the ends of the chromosome and to ensure survival,

(ii) An origin of initiation of DNA replication and

(iii) A centromere for movement and segregation into daughter cells.

Yeast artificial chromosomes have been constructed by putting the above three elements together; they can propagate in yeast. YACs are maintained in circular form before insertion of foreign DNA into them. The circular YAC has a BamHI cleavage site at the end of telomeric sequences, and Smal cleavage site at some other position.

In addition, it has a marker TRP_X on one side of the centromere, while other marker URA₃ is carried on the other side. The enzyme BamHI cleaves the circular YAC to produce a linear molecule whose both ends act as telomeres.

The other enzyme Smal cleaves this linear molecule into two pieces between which foreign DNA can be inserted (ligated) to produce a linear chromosome carrying different genes. The markers TRP_X (in one arm) and URA₃ (in the other arm) help in selection of the molecules in which the two arms have been joined. The DNA inserted may be 50-300 kbp in length, thus eukaryotic genes can be carried in YACs. The whole genomic library can be maintained in the YACs and can be propagated in yeast.