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Telomerase, Cancer and Aging

Group no. 6
MED 1 -B2
Jamee Dela Rosa
Haiezel dela Cruz
Lalaine dela Cruz
Maureen dela Cruz
Erika dela Fuente
Mariel dela Cruz

OBJECTIVES
1. Describe the physical structure of a chromosome and describe what
makes up the centromere and telomeres of a chromosome
2. Briefly review the process of replication of a linear chromosome and
explain what accounts for the natural shortening of the lagging strand
3. Explain Cellular Senescence, some mechanisms that accounts for it and
how is it related to cancer
4. Describe the enzyme Telomerase and its structure.
5. Identify specific cell populations that express the enzyme h.

6.
7.
8.

9.

Discuss the mechanism involve in Telometric


lengthening and of sealing the telometric ends.
Explain the role of telomere binding proteins in the
regulation of telomerase function.
Explain the relationship between telomeres in regards
to aging and cancer.
Discuss the potential advantages of using antitelomerase agents in treating cancer cells.

CHROMOSOME

Tightly coiled DNA


Carriers of gene Unit of Heredity
2 pairs of 23 filamentous rod shape body
Present in the nucleus
Chromosomes are not visible in active nucleus,
but are clearly seen during cell division

PHYSICAL STRUCTURE
Parts:
CHROMATIDS
Double Stranded DNA

Long arm (q)


Short arm (p)
CENTROMERE
Joins chromatids together

TELOMERE
Found at each end of
Chromatid

CENTROMERE
Part of a chromosome that links sister chromatids
Rich in adenine-thymine base pairs
Condensed regions within the chromosome that are
responsible for the accurate segregation of the replicated
chromosome during mitosis and meiosis

TELOMERE
Telos - END'
Mers -PART
Short thymine-guanine sequence
5-TTAGGG-3 Sequence
Found at each end of a chromatid,
which protects the end of the
chromosome from deterioration or
from fusion with neighbouring
chromosomes

DNA Replication: Review

KEY POINTS
ORIGIN is the beginning
REPLICATION FORKS are
the sites at which DNA
synthesis is occurring
New chains grow 5-3
Bidirectional
Semi-conservative

ORDER OF ACTION

Unwinding proteins
Preventing the Reannealing
Primase makes RNA primer
DNA polymerase makes DNA
RNAse H removes RNA primer
DNA polymerase fills in gaps
DNA ligase joins gaps

UNWINDING
OF PROTEINS
HELICASE
Made up of 6 proteins
arranged in a ring shape

Motor proteins
Unpackage an organisms
gene

PREVENTING THE
REANNEALING
SINGLE STRAND BINDING
PROTEINS
Tetramers
Attached to the postreplication fork single
strands of DNA,
preventing their
"reannealing

PRIMASE MAKES
RNA
PRIMER
PRIMASE
A type of RNA
Polymerase
Creates a RNA Primer
Key importance in
DNA Replication

DNA POLYMERASE MAKES DNA

DNA POLYMERASE
Creates DNA Molecules
by assembling
nucleotides

RNAse H
REMOVES RNA
PRIMER
RNAse H
Non- Specific
Endonuclease that
catalyzes the cleavage
of RNA
Removing the RNA
primer

DNA POLYMERASE
FILLS IN THE GAPS

DNA LIGASE JOINS


GAPS

DNA LIGASE
A ligase that facilitates
the joining of DNA strand
together by catalyzing the
formation of a
phosphodiester bond.

WHAT ACCOUNTS FOR THE NATURAL


SHORTENING OF LAGGING STRAND?
During chromosome replication, the enzymes
that duplicate the DNA cannot continue
duplicating all the way to the end of the
chromosome.
OKAZAKI FRAGMENTS
RNA Primers attached ahead on the lagging strand

CELLULAR SENESCENCE
Latin: Senescere Grow Old
It is the phase or stage in which normal cells
cease to divide
Associated with a loss of telomerase activity
As human telomeres grow shorter, eventually
cells reach the limit of their replicative
capacity and progress into senescence

PROPERTIES OF CELLS IN CELLULAR


SENESCENCE
Increase synthesis of proteases
Decrease in synthesis of pro-collagen and tissue
inhibitors of metalloproteases
Irreversible growth arrest
Dramatic change in morphology
-lose original shape
-acquire distinct flattened cytoplasm
-change in nuclear structure, gene expression,
protein processing and metabolism

CELLULAR SENESCENCE
Today it is known that somatic cells derived from
human newborns will usually divide 80 to 90
times in culture
Whereas those from a 70-year-old are likely to
divide only 20 to 30 times
When human cells that are normally capable of
dividing stop reproducingor, in Hay- flick's
words, become "senescent"they look different
and function less efficiently than they did in
youth, and after a while they die.

CELLULAR SENESCENCE: TELOMERE


SHORTENING
Telomeres shorten because of the lagging
strand phenomenon.
A section of telomeres is lost during each cycle
of replication.
Progressively lose approximately 50-200
nucleotides during each mitotic replication.
Cellular senescence is triggered when cells
acquire one or a few critically short telomeres

CELLULAR SENESCENCE:
ACCUMULATION OF DAMAGE
Major cause of aging which is due to highly
reactive substances containing oxygen (oxygen
free radicals)
Oxygen free radicals damages the DNA
Damaged DNA accumulate mutations with
fewer proliferation

CELLULAR SENESCENCE: GLYCATION


Glycation is the result of a sugar reducing
molecule, such as fructose or glucose,
bonding to a protein or Lipid molecule
without the controlling action of an enzyme.
This reaction products [AGEs or advanced
glycation end products] are irreversible and
detrimental for extracellular protein function.

CELLULAR SENESCENCE:
MITOCHONDRIAL DNA DAMAGE
The mitochondria have their own genetic material
(mtDNA), which is distinct from the nuclear DNA in
the cell.
Mitochondria produce ATP (energy) and free
radical (byproducts of respiration)
Aerobic condition 4% of oxygen are metabolize by
mitochondria.
In normal condition, the oxygen is reduced to
produce water.

CELLULAR SENESCENCE:
MITOCHONDRIAL DNA DAMAGE
However, the oxygen is instead prematurely
and incompletely reduced to give the
superoxide ion.
The superoxide ion hydrogen peroxide
hydroxyl radical Reactive oxygen species
(ROS)
ROS leads to damage the cell membrane,
structural protein and mitochondrial and
nuclear DNA.

CELLULAR SENESCENCE SUMMARY


TELOMERE
SHORTENING

MITOCHONDRIA

ROS
CELLULAR
SENESCENCE

OXIDATIVE STRESS

OXYGEN FREE RADICALS

AGEs

GLYCATION

CELLULAR SENESCENCE AND CANCER


Senescence involves p53 and pRb pathways
and leads to the arrest of cell proliferation
plays an important role in suppression of
emergence of cancer, although inheriting shorter
telomeres probably does not protect against
cancer. Why?
Because with critically shortened telomeres,
further cell proliferation can be achieved by
inactivation of p53 and pRb pathways.
Cells entering proliferation after inactivation of
p53 and pRb pathways undergo crisis.

CELLULAR SENESCENCE AND CANCER


Usually almost all cells die when it enters crisis
but rare cells emerge from crisis and
immortalized through telomere elongation by
either activated telomerase (becomes cancer
cells)

WHAT IS TELOMERASE?

TELOMERASE
Background
Telomerase is an enzyme, discovered by
Elizabeth Helen Blackburn (professor of
biochemistry and biophysics at the University of
California, San Francisco School of Medicine)
and Carol Greider (professor of molecular biology
and genetics at the Johns Hopkins University
School of Medicine ) in 1985.

TELOMERASE
A ribonucleoprotein reverse transcriptase
(enzyme) that synthezises telomeric DNA.
Tetrahymena in which the enzyme was first
discovered.
Tetrahymena 5 TTGGGG 3
In humans 5 TTAGGG 3

TELOMERASE
is an enzyme that adds telomeric sequences to the
ends of each chromosome. Unlike most enzymes,
which consist entirely of protein, telomerase is a
combination of a protein and an RNA.
The enzyme is a protein and RNA complex called
telomere terminal transferase, or telomerase.
Telomerase is present in most fetal tissues, normal
adult male germ cells, stem cells, in proliferative
cells of renewal tissues, and in most tumor cells.

TELOMERASE STRUCTURE

TELOMERASE STRUCTURE
Human telomerase is composed of at least
two sub-units:
human Telomerase Reverse Transcriptase
(hTERT) - protein component
human Telomerase RNA (hTR or hTERC)
- RNA Component

human Telomerase Reverse


Transcriptase (hTERT)
Protein Component
The coding region of the hTERT gene is
3396bp, and translates to a protein of 1131
amino acids
The hTERT gene maps to chromosome band
5p15.33.

human Telomerase RNA


(hTR or hTERC)
RNA component
referred to as the RNA guide
template region of hTR is
3'-CAAUCCCAAUC-5'
sequence complementary to telomere repeat
[TTAGGG]
serve as template for telomere synthesis and
elongation

MECHANISM OF TELOMERIC
LENGTHENING

hRNA
CAAUCCCAAUC

TELOMERE

hTERT

GGTTAGGGTTAGGG
CCAAUCC

TELOMERASE

STEP 1: BINDING
CAAUCCCAAUC

GGTTAGGGTTAGGG
CCAAUCC

STEP 2: ELONGATION

GGTTAGGGTTAGGGTTAGGG TTAG
CCAAUCC

CAAUCCCAAUC

STEP 3: TRANSLOCATION

GGTTAGGGTTAGGGTTAGGG TTAG
CCAAUCC

CAAUCCCAAUC

STEP 4: ELONGATION

GGTTAGGGTTAGGGTTAGGG TTAGTTAGGG TTAG


CCAAUCC

CAAUCCCAAUC

MECHANISM OF SEALING THE


TELOMERIC ENDS

T-loop

HETERO-DUPLEX OR T-LOOP

It was proposed that the t-loop is formed by strand


invasion of the 3' G-rich overhang into the preceding
telomeric tract to form a lariat with a D-loop at the
looptail junction .

Importance of T-loop
stabilizes the telomere
prevents the telomere ends from being recognized
as break points by the DNA repair machinery thus
it gives protection from exonucleases.
preventing the telomere from eliciting a DNA
damage response manifested as cell-cycle arrest
or apoptosis

Importance of T-loop
prevention of chromosome end fusions or non
homologous end joining (NHEJ)
prevention of homologous recombination
between telomeric regions
regulation of telomere length homeostasis.

What prevents the telomerase


from over-extending the ends of
a linear chromosome?

Shelterin Complex
The T-loop is held together by six known
proteins:
TRF1, TRF2, POT1, TIN2, RAP 1 and TPP1,

Shelterin Complex
1. TRF 1 (Telomeric Repeat
binding Factor 1)

It binds along the length of


the T-loop.
along with TRF2, it normally
prevents telomerase from
adding more telomere units
to telomeres.
But
when
telomere
lengthening is required,
TRF1 recruits helicases to
facilitate the process.

Shelterin Complex
2. TRF2 (Telomeric Repeat
binding Factor 2)

It appears to promote
formation of D-loop
prevents
ataxia
telangiectasia
mutated
(ATM) activation, which is
a DNA damage response
(DDR) to DNA double strand
breaks.
But when DNA repair of
telomeres is required, TRF2
recruits
DNA
repair
proteins.

Shelterin Complex
3. TIN 2 (TRF1 Interacting
Nuclear factor 2)

links TRF1 with TRF2, and


connects both to TPP1 .
TIN2 is believed to facilitate
recruitment oft singlestranded telomere-binding
proteins to telomeres.
TIN2 interacts with TRF1 and
has been suggested to
stabilize the T-loop.

Shelterin Complex
4. POT 1 (Protector Of
Telomeres 1)

only binds to the singlestranded 3-end DNA


overhang.
POT1 prevents ataxia
telangiectasia and Rad3
related (ATR) activation,
which is a DNA damage
response (DDR) to DNA
double strand breaks.
Humans only have a single
POT1, whereas mice have
POT1a and POT1b.

Shelterin Complex
5. RAP1 (Repressor/Activator
Protein 1)

binds to TRF2, and facilitates


TRF2 function.
RAP1 protects telomeres
from non homologous end
joining (NHEJ).
Unlike the other shelterin
proteins, RAP1 has functions
independent of its function
within the shelterin complex:
RAP1 regulates transcription
and affects NF- kb signaling.

Shelterin Complex
6. TPP1 (TINT1, PTOP,
PIP1 POT1-TIN2

organizing protein)

interacts with POT1 and


regulates its function.
When telomeres are to be
lengthened, TPP1 is a
central factor in recruiting
telomerase to telomeres.
Deletion of TPP1 from
shelterin elicits an ATRmediated DDR.

AGING

What is aging?
Aging is a degenerative process that is
associated with progressive accumulation of
deleterious changes with time, reduction of
physiological function and increase in the
chance of disease and death.

Telomeres alone do not reduce lifespan


there are some factors that also plays
important role in aging.

but
an

According to geneticist Richard Cawthon and


the colleagues at the University of Utah,
Shorter telomeres are associated with shorter
lives. Among people older than 60, those with
shorter telomeres were three times more likely
to die from heart disease and eight times more
likely to die from infectious disease.

Telomere shortening
When people are divided
into two groups based on
telomere length, the half
with longer telomeres lives
an average of five years
longer than those with
shorter telomeres.

Chronological Age
After age 60, the risk of death doubles every 8 years. So a
68-year-old has twice the chance of dying within a year
compared with a 60-year-old.

Oxidative Stress
Oxidative stress is the damage to DNA, proteins, and
lipids (fats) caused by oxidants, which are highly reactive
substances containing oxygen

Glycation
Glycation happens when glucose, the main sugar we use
as energy, binds to some of our DNA, proteins, and lipids,
leaving them unable to do their jobs.

Is it possible to revert old cells into


young cells again?

Is it possible to revert old cells into young cells


again?
No. Why?
Using that in humans will be more difficult
because Mice make telomerase throughout their
lives but the enzyme is switched off in adult
humans.

Telomerase and Cancer


Telomerase is expressed in
almost all human cancers but
is inactive in most normal
cells.
Cancer cells are malignant
cells which multiply until
they form a tumor that
grows uncontrollably.
Telomerase is a good
biomarker
for
cancer
detection because most
human cancers cells express
high levels of telomerase.

Telomerase as an Agent to Extend Cellular Life Span

Telomere shortening has been observed in most


dividing somatic cells, eventually leading to cell
senescence when critically short telomeres are
reached.
Telomerase has been identified as a
ribonucleoprotein enzyme that can synthesize
telomeric repeats onto chromosomes.
Telomerase can be an agent to extend cellular
life span especially in cancer cells.

Telomerase as an Agent to Extend Cellular Life Span


ADVANTAGES

DISADVANTAGES

If telomerase are activated in If telomerase are inactivated,


somatic cell, human cells will not
telomere shortening can be
age.
anticipated.

If telomerase are inactivated, Telomerase has been detected in


telomeres in cancer cells would
human cancer cells and is found
shorten, just like they do in normal
to be 10-20 times more active
body cells
than in normal body cells
It will be able to mass produce cells Blocking telomerase could impair
for transplantation
fertility, wound healing and

production of blood cells and


immune cells

Anti-telomerase Agents to Treat Cancer Cells

Harley & Greider


inhibitory agent could
cause the telomeres of
cultured tumor cells to
shrink
Apoptosis

Blackburn
cells sometimes
compensate for the loss
of telomerase, repair
shorter ends for
Recombination

Potential advantages of antitelomerase agents


SPECIFICITY
Anti-telomerase treatment would be very
selective in that only cells with an activated
telomerase would be affected (most normal
adult tissues have NO telomerase activity).

THANK YOU!

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