Research Article: Telomere Lengthening and Other Functions of Telomerase

Date Published: , 2012

Publisher: A.I. Gordeyev

Author(s): M.P. Rubtsova, D.P. Vasilkova, A.N. Malyavko, Yu.V. Naraikina, M.I. Zvereva, O.A. Dontsova.



Telomerase is an enzyme that maintains the length of the telomere. The telomere
length specifies the number of divisions a cell can undergo before it finally
dies (i.e. the proliferative potential of cells). For example, telomerase is
activated in embryonic cell lines and the telomere length is maintained at a
constant level; therefore, these cells have an unlimited fission potential. Stem
cells are characterized by a lower telomerase activity, which enables only
partial compensation for the shortening of telomeres. Somatic cells are usually
characterized by the absence of telomerase activity. Telomere shortening leads
to the attainment of the Hayflick limit, the transition of cells to a state of
senescence. The cells subsequently enter a state of crisis, accompanied by
massive cell death. The surviving cells become cancer cells, which are capable
both of dividing indefinitely and maintaining telomere length (usually with the
aid of telomerase). Telomerase is a reverse transcriptase. It consists of two
major components: telomerase RNA (TER) and reverse transcriptase (TERT). TER is
a non-coding RNA, and it contains the region which serves as a template for
telomere synthesis. An increasing number of articles focussing on the
alternative functions of telomerase components have recently started appearing.
The present review summarizes data on the structure, biogenesis, and functions
of telomerase.

Partial Text

The genetic information in eukaryotic cells is stored in linear DNA molecules known
as chromosomes [1]. It was revealed as early
as in the 1930s that the behavior of the whole chromosome and its fragments in cells
varies. Torn chromosomes can fuse with each another, rearrange, and they are
characterized by instability [2, 3]. An assumption was made back in the 1930s
that these differences are caused by the presence of specific nucleotide sequences
at the chromosome ends; these sequences were referred to as telomeres [3–5]. The telomeres consist of repeating sequences
and a set of special proteins, which interact with these repeats and spatially
organize them in a specific manner, resulting in the formation of the nucleoprotein
complex known as telomeric heterochromatin [6,
7]. Shortening of the 5’-terminus of
the daughter strand, caused by the removal of the terminal RNA-primer and the
subsequent incomplete replication of linear DNA molecules, is observed during the
genome replication occurring upon cell fission. The independent formulation of the
so-called “end-replication problem” was proposed in the 1970s by A.M.
Olovnikov and J. Watson [8, 9]. Olovnikov hypothesised that there is a
special enzyme, i.e. telomerase, which is capable of compensating for the
“end-replication problem.” This enzyme was discovered in 1987 by C.
Greider and E. Blackburn [10].

Telomeres are the repeating nucleotide sequences bound to the specific proteins
protecting chromosome ends against degradation and the double-strand break repair
systems [12, 13]. As data accumulated, a hypothesis was postulated that telomeres
consist of three distinct regions. Firstly, they contain the so-called cap, a
terminal structure protecting the chromosome ends against degradation and the
double-strand break repair systems (DDR – DNA damage response); they also
regulate telomere elongation. The major component of a telomere is a double-stranded
DNA (dsDNA) consisting of repeating and transcribed sequences. The third component
of a telomere is represented by repeating telomere-associated sequences, the
so-called subtelomeric regions [14, 15]. The telomere nucleotide sequence is
enriched in thymidine and guanosine residues and is appreciably conserved. Mammalian
telomeres are a double-stranded region consisting of TTAGGG repeats and the
150–200 nucleotide long 3’ G-strand overhang. According to one of the
hypotheses, the G-strand overhang is intertwined with the double-stranded telomeric
region, thereby forming a t-loop. The so-called D-loop is formed at the site of the
interaction between the protruding 3’-terminus with the double-stranded region
( Fig. 1 ). t-Loops were
detected via electron microscopy after DNA was extracted and treated in a special
manner. However, the existence of these structures in cells has as yet not been
unequivocally proven; therefore, the D-loops are considered as tentative structures.
Telomere functions depend on the minimal length of telomeric repeats and the
activity of the protein complex associated with them. This complex is known as
shelterin and consists of six proteins: TRF1, TRF2, POT1, TIN2, TPP1, and RAP1. The
proteins TRF1, TRF2 (telomeric repeat binding factor 1 and 2) and POT1 (protection
of telomeres protein 1) are bound to telomeric DNA. TRF1 and TRF2 are bound to the
double-strand telomeric regions; РОТ1 can be bound to the
3’-protruding single-stranded region of the G-strand [16]. TRF1 and TRF2 bind telomeres independently; they do not
interact with each other. Both proteins, which have the form of a homodimer and an
oligomer, are capable of specifically binding the DNA duplex to the telomeric
sequence 5’-YTAGGGTTR-3’ [16–20]. POT1 binds highly specifically to the telomeric single-stranded DNA
(ssDNA) 5′-TAGGGTTAG-3′, attesting to a possible interaction both with the G-strand
overhang and with the sequence of the D-loop displaced by it [13, 21–23]. POT1 interacts with TRF1. It is believed
that TRF1 facilitates the binding of РОТ1 to the single-stranded
telomeric region in this manner. Via its independent domains, TIN2 (TRF1-interacting
protein 2) simultaneously interacts with TRF1 and TRF2, as well as with the
ТРР1–РОТ1 complex, forming a bridge
between the shelterin components [24, 25]. The C-terminal domain of TPP1 is bound to
TIN2, the central domain is bound to POT1 [26–29]; thus, POT1 is attracted to the telomeres [30, 31]. Moreover, TPP1
contains a domain interacting with telomerase on its end. This fact supports the
assumption that TPP1 attracts telomerase to the chromosome end [32]. Protein RAP1 forms a complex with TRF2 and
the telomere [33, 34]. It has been demonstrated in studies undertaken by several
research teams that RAP1 is not essential for telomere capping; however, it impedes
recombination at telomeric regions and enhances their stability [35, 36].
Thus, RAP1 (unlike TRF1, TRF2, POT1, and TPP1) does not protect telomeres [32, 35,

The assembly of telomerase, its existence in a cell, and its entry to the telomeres
are processes that are similar in some aspects, yet differ in other aspects with
regards to evolutionary distant organisms [57–59]. Properties common to all telomerase components have been revealed:
reverse transcriptase (TERT), telomerase RNA (TER), and TER-binding proteins, which
stabilize RNA and facilitate the assembly of the active enzyme. It should be noted
that only TERT is a highly conserved telomerase component. The data obtained through
the study of the components within telomerase are rather inconsistent
[60–64]. Telomerase apparently
interacts with various components throughout its vital activity and thus can be
found in various complexes.

As previously mentioned, telomerase consists of the two major components; however,
the synthesis and processing of each component, as well as the formation of an
active enzyme, require the contribution of a large number of additional factors. The
regulation of TERT expression at the transcriptional phase was thoroughly discussed
in the review by Skvortsov et al . [121]. The alternative splicing of the primary transcript of
the hTERT gene yields 13 different mRNA variants [122–125]. Out of these variants, the so-called
α- and β-forms are both the most common and well-studied. When the
α-form is produced, 36 nucleotides are deleted from the sixth exon, resulting
in the change of the reverse transcriptase motif A. The open reading frame is not
disrupted [126, 127]. The deletion of 182 nucleotides from the exons 7 and 8
and the insertion of 38 nucleotides triggers a premature translation termination,
resulting in the formation of the β-hTERT, which does not contain the three
essential reverse transcriptase motifs [128,
129]. Splicing can independently occur
at different sites; therefore, different forms of hTERT mRNA often
co-exist in cells. The combination of different mRNA forms and their number depend
on the particular cell type. Thus, one of the mRNA variants (α-/β+ form)
has regulatory functions by acting as a dominant negative inhibitor of telomerase
activity both in normal and tumor cells.

The major activity of telomerase ensures the RNA-dependent telomere elongation [168]. The telomerase catalytic cycle consists
of several sequential stages. One telomeric repeat is added after the substrate
binding. The resulting product can either dissociate from the enzyme’s active
site or undergo translocation, followed by elongation. The ability of telomerase to
move the DNA synthesized to the template beginning site allows one to use two
processivity types to describe its function. Nucleotide addition (type I
processivity) is intrinsic to all polymerases, since repeat addition (type II
processivity) is unique to telomerase and determines the ability of an enzyme to
repeatedly copy an RNA template region via elongation of the one substrate molecule
only [169, 170].

The first batch of data on the alternative functions of telomerase were reported at
the early stages of the study of this enzyme. The products of other enzyme
activities were detected when studying the activity, substrate specificity, and
other properties of telome­rase. It turned out that telomerase is also capable of
acting as a catalyst for the other reactions ( Fig. 4 ).

Data attesting to the diversity of the functions carried out by the major components
of cell telomerase have recently been reported. Some of these functions (such as the
nuclease and transferase activities) are associated with the major role of
telomerase and its polymerase activity. The other functions (e.g., regulation of
gene expression, protection against apoptosis, and contribution to the DNA response
to damage) are not directly associated with polymerase activity. It should be noted
that the telomerase content in higher eukaryotic cells is very low; hence, almost
all the data have been obtained under conditions of artificial expression of its
components. Under such conditions, conclusions can be drawn that are divorced from
reality. Researchers from different laboratories obtain inconsistent data, which are
difficult to interpret. The inconsistency is most likely a result of the use of
different systems and models. Nevertheless, the new data reported allows one to
assume that telomerase has a more versatile function, and that its impact on the
cell is not confined to the regulation of the length of telomere. 





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