telomerase and function.pptx

Telomere is the end part of a chromosome.its length is maintained by na enzyme called telomerase.if telomerase is lacking,many genetic diseases may result( like progeria)

1. TELOMERASE AND IT’S FUNCTION

2. “Telomeres” is just a fancy scientific term for “the ends of a chromosome.’’  -Carol Greider TELOMERE and TELOMERASE Elizabeth Blackburn and Jack Szostak discovered that a unique DNA sequence in the telomeres protects the chromosomes from degradation. Carol Greider and Elizabeth Blackburn identified telomerase, the enzyme that makes telomere DNA

3. The Nobel Prize in Physiology or Medicine 2009 jointly to Elizabeth H. Blackburn, Carol W. Greider and Jack W. Szostak for the discovery of “how chromosomes are protected by telomeres and the enzyme telomerase “

4. Tetrahymena thermophila  5′-CAACCCCAA-3′, Telomerase RNA sequence

5. Telomere DNA protects the chromosomes These end-points are often compared to the plastic protective covers on the tips of shoelaces Barbara McClintock and H J Muller revealed that the ends of chromosome are protected,and telomere is essential for chromosome stability(1930s) They were able to recover chromosomes without deletions when treated with xray and fusion of chromosome occurred when telomere is lost.

6. EFFECTS OF TELOMERE LOSS Essential genes are lost When telomere length reaches zero, difficulty of cells to divide successfully Chromosome breaks and rearrangement Nuclease attacks Apotopsis Premature aging syndromes Replicative cell senescence Chromosome fusion Chromosome instability

7. STRUCTURE OF TELOMERASE- RIBONUCLEOPROTEIN Multiple protein subunits+ RNA component(TERC) Catalytic subunit contain 3 domains it consist of: A reverse transcriptase domain-(TERT) palm and finger subdomains RNA binding domain(TRBD) Thumb domain

8. Telomere terminal transferase( telomerase)

9. END REPLICATION PROBLEM Loss of telomeric region

10. HOW IS END REPLICATION PROBLEM SOLVED? Telomerase extends the 3′ end of template DNA Thereby providing an extended 3′ end,additional template is available for replication machinery.

11. Telomere binding proteins regulate telomerase activity and telomere length When telomere is short,telomere binding proteins accumulate and induce telomere extension.As telomere becomes longer more of the telomere binding proteins accumulate and inhibit telomere extension at 3’ OH end of telomere Cdc13: recruitment of telomerase to telomeres pOT1: inhibitor of telomerase Rap1,Rif1,Rif2: inhibition of telomerase binding TRF1&TRF2 binds to dsDNA, RAP1,TIN2,TPP1,POT1- All these proteins together form shelterin complex that shelter telomere from DNA repair enzymes

12. Telomere length maintenance by telomerase It is a complex multistep process that involves a series of molecular events including hTERT protein transport and trafficking into nucleus. hTR and hTERT assembly with accessory components in nucleus Recruitment to telomere at appropriate time during replication Assembly of functional telomerase holoenzyme

13. SENSITIVITY OF TELOMERIC REGIONS TO DSBs Highly sensitive to DSBs Presence of DNA end is considered as a double stranded break in the DNA ,which is targeted by the DNA repair machinery,leads to fusion events results in random chromosome breaks and an abnormal chromosome with 2 centromeres

14. Irreparable DSBs at telomeres contribute to aging and stress induced senescence. Ectopic localisation of TRF2 caused delay of interstitial DSB repair A single DSB is enough to kill a cell or cause chromosomal aberrations leading to cancer. Chromosome healing act as a mechanism to compensate the deficiency of repair mechanism. Telomere binding proteins distinguish telomeres from other DNA ends in the cell,Thereby preventing fusions Elimination of these proteins leads to recognition of telomeres as normal Dna breaks and are attacked by enzymes TRF2 directed t-loop formation protects the ends of DNA

15. But telomerase cannot recognise this form of telomere because it lacks an obvious single strand 3’ .it has been proposed that as Telomeres shorten and they would have increasingly difficult time forming the t loop ,thereby allowing increased access to the three prime end of the Telomere.

16. What happens if telomerases are non functional 😳? Loss of protective function of Telomeres has been hypothesized to cause DNA damage response (DDR) .telomerase deletion response occurs when telomeres no longer be maintained by telomerase. A discrete set of genes called telomerase deletion signature is upregulated in TDR. Chromosome instability result from Telomere loss B/F/B cycles occur- results in chromosome fusions and chromosome with 2 centromeres formed.broken chromosomes resulted during anaphase, daughter cromatids on the next replication again fuses since telomere is lost and gradually chromosome is lost and cell dies

17. Bfb cycles can generate many types of chromosomal rearrangements associated with the cancer, for example inverted repeats most common in pancreatic Cancer The BFB cycle begins when the end region of a chromosome, called its telomere, breaks off. When that chromosome subsequently replicates it forms two sister chromatids which both lack a telomere.the lack of a telomere on these two sister chromatids causes them to fuse with one another. During anaphase the sister chromatids will form a bridge . Being pulled in opposite directions will cause the two sister chromatids to break apart from each other, but not necessarily at the site that they fused. This results in the two daughter cells receiving an uneven chromatid. Since the two resulting chromatids lack telomeres, when they replicate the BFB cycle will repeat, and will continue every subsequent cell division until those chromatids receive a telomere, usually from a different chromatid through the process of translocation

18. TELOMERASE IN HUMAN Telomerase is on during Fetal development and remains active in proliferative cells. Each zygote begins their life witH telomeres of maximum length - Stem cells , activated lymphocytes,hair follicles Telomerase is down regulated but still detectable in: Epithelial, fibroblasts and endothelial cells high content of telomerase in 80-90%of invasive cancers Absent in most of the somatic cells

19. Functions of telomerase 1.solving end replication problem

20. Like all other DNA polymerases telomerase act to extend 3 prime end of DNA substrate but unlike DNA polymerases telomerase doesn’t need an exogenous DNA template to direct the addition of new dntps instead r n a component of telomerase act as template for adding telomere sequences to 3 prime terminus the newly synthesized DNA is single stranded.this elongated template is then used to extend 5’end of new DNA telomerase is a novel DNA polymerase that doesn’t require an exogenous template

21. 2.CELL SENESCENCE

22. 3 markers of cellular aging Telomerase activity Telomere length Cellular oxidative stress

23. HOW DO WE AGE?👶👴 The Hayflick Limit is a concept that helps to explain the mechanisms behind cellular aging. The concept states that a normal human cell can only replicate and divide  forty to sixty times  before it cannot divide anymore, and will break down by programmed cell death or apoptosis Hayflick hypothesized that the limited replicative capability of the cell related to aging in cells and, consequently, to human aging. Red blood cells live for about four months, while white blood cells live on average more than a year. Skin cells live about two or three weeks. Colon cells have it rough: They die off after about four days In 2009, Blackburn and Szostak received the Nobel Prize in Physiology or Medicine for their work on telomerase, in which the Hayflick Limit played an essential role

24. Telomerase activity is implicated in cell senescence- replication senescence and aging Natural aging Telomerase activity is normal in early embryo but after birth it is active Only in reproductive cells and stem cells . Chromosome shortening after each replication,reaches astage where it can no longer divide- replicative senescence Essential genes are lost ,cell dies When telomere length reaches a critical point,TRF2 forms a protective cap-t loop Telomere shortening act as longevity clock. Leads to termination of a cell lineage.

25. Telomere deficiency Dyskeratosis congenita (DC)  is a rare congenital disorder that is characterized by premature aging and, at the molecular level, the inheritance of short telomeres. The disease is caused by mutations in a number of genes, all of which encode products involved in telomere maintenance.

26. DC is an inherited disease which result from progressive bone marrow failure . Individuals have shorter telomeres.,weak immune system,bone marrow is unable to produce enough blood cells. Genetics: a full human lifespan requires both telomerase RNA alleles to be functional ( maternal and paternal)because telomerase gene product quantity matters.But in DC, one copy of telomerase RNA is deficient either maternal or paternal. In about half of this people disorder is caused due to mutation in TERC,TERT,TINF2 Genes these genes provide instructions for maintaining telomeres

27. A lacy-looking rash on the face, neck and chest White patches in the mouth, called leukoplakia Fingernails and toenails that are not shaped normally Many children with DKC develop  bone marrow failure . Have more infections because of problems with white blood cells Be tired and have low energy because they do not have enough red blood cells (anemia) Have bleeding problems because of low levels of platelets

28. Hutchinson-Gilford Progeria Syndrome (HGPS) Rare premature aging disorder caused by mutations in the gene LMNA, which encodes the nuclear matrix protein lamin A. Previous research has shown that the average telomere length in fibroblasts from HGPS patients is shorter than in age-matched controls.

29. Stress and telomerase activity Chronic stress reduces the ability of adult cell to replenish itself When cells are exposed to years of chronic stress ,effects override normal aging making our telomeres look like they’re from an older person . Groups of people with Psychiatric disorders have Shorter telomers It is due to the fact that epigenetic changes altered the overall chromatin and transcriptional properties, ultimately resulting in senescence

30. TELOMERASE FUNCTION IN MITOCHONDRIA-OXIDATIVE STRESS AND AGING During aging, damaged mitochondria that produce less ATP and more reactive oxygen species (ROS) accumulate. ROS cause oxidative stress, damaging mitochondria and resulting in an energetic crisis that triggers neurodegenerative diseases and accelerates aging. Prolonged exposure to oxidative stress is associated with Shorter telomere and reduced telomerase activity Due to the high content of guanine residue telomeres are highly sensitive to oxidative damage

31. Transport of hTERT into mitochondria mediated by mitochondrial sequence at n-terminal of hTERT accumulating endogenous stress-induced the nuclear export of hTERT to cytosol and decrease nuclear and total telomerase activity. The addition of antioxidants decelerated this process Mitochondrial hTERT increases membrane potential reduce ROS production ,protect mitochondrial DNA indicating an additional protective function of telomerase in mitochondria.

32. Role of telomerase in regulating immune system Lymphocytes express higher level of telomerase activity ,this may sometimes result in autoimmune disorders.during autoimmunity, causes prematurity of aging or whether entering the senescence program alters the function of immune cells in such a way that they lose their ability to discriminate between self and non-self and cause disease. Auto antibody production also increases with age

33. 2 . Hyperactivity of TELOMERASE is implicated in cancer When normal cells have telomerase activity,: Absence of senescence, immortality Telomeres are longer than normal telomere shortening plays a key role in protecting human from cancer by limiting the number of divisions a potential tumor cell can or by preventing the transformation of a normal cell into tumor cell cancer cells achieve proliferative immortality by activating normally silent hTERT genes that encode telomerase complexes with other proteins and functional RNA to make ribonucleoprotein enzyme complex ( via mutation)

34. 2.(a)Mutation in hTERT promoter 70%melanomas 80%gliblastomas 60%bladder cancer 30%thyroid cancer

35. Telomerase as a target for anticancer therapeutics Antitelomerase therapeutics: Induction of apoptosis and PCD Via-development of vaccine Small molecular inhibitors of hTERT Eg: BIBR1532 is a potent telomerase inhibitor that showed potential anti-tumor activities in several types of cancer, by triggering replicative senescence and apoptosis.

36. Imetelstat is a telomerase inhibitor that acts on cells with hyperactive telomerase and short telomere lengths Act as a competitive inhibitor

37. ANTITELOMERASE IMMUNOTHERAPEUTICS Anti-telomerase cancer immunotherapy, using human telomerase reverse transcriptase (hTERT) as a tumor antigen. Vaccination, using hTERT peptides or adaptive transfer of hTERT-specific cytotoxic T lymphocytes, induces augmented tumor regression.eg:GV1001 Degradation of telomerase by proteosome result in the formation of protein fragments that are detected by immune system and targeted by T cells and tumour cells are destroyed.eg

38. Exploiting telomerase activity to selectively kill tumor cells Introducing a modified nucleoside into cells so that telomerase would incorporate it into TELOMERIC DNA .an altered nucleotide incorporated into telomere would not bind to shelter in proteins efficiently and lead to telomere dysfunction and cell death. Eg:nucleoside analogue 6-thio-2’-deoxyguanosine (6-thio-dG) is recognized by telomerase and is incorporated into de novo synthesized telomeres. This results in modified telomeres, leading to telomere dysfunction

39. 2( b).ALTERNATION IN ALTERNATIVE SPLICING OF hTERT PRE mRNA Some pre mRNAs are spliced to full length hTERT ,makes functional telomerase in somatic cells This reprogramming of alternative splicing is the cause of tumorigenesis

40. 2(c).ALTERNATIVE LENGTHENING OF TELOMERE Alternative Lengthening of Telomeres (also known as “ALT”) is a telomerase-independent mechanism by which cancer cells avoid the degradation of telomeres recombination-mediated telomere synthesis

41. Lifestyle Interventions targeting telomerase

42. What telomere has to tell you?🧐 Telomeres listen to you, they listen to your behaviors, they listen to your state of mind,” said Blackburn You don’t have to go to the gym three hours a day or run a marathon a week. People who do moderate aerobic exercise – about three times a week for 45 minutes – have telomeres pretty much as long as marathon runners. Have a good state of mind,have good practices and habits,eat healthy,Take care of Your body,Spend some minutes for exercise,stay healthy ,stay happy this is the mantra of long telomeres which help you surpass the effect of ageing.

43. Cellular senescence is also thought to serve as a safety mechanism against cancer, preventing cells with potentially dangerous levels of DNA damage from replicating and removing them from the system. However, the problems begin when these cells are allowed to linger. A Double-Edged Sword – Cancer and Senescence

44. REFERENCES Brown. Genome.Oxford ; New York : Wiley-Liss, 2002 Jafri, Shakeel A Ansari, Mohammed H Alqahtani, Jerry W Shay.Roles of telomeres and telomerase in cancer, and advances in telomerase-targeted therapies
Janet, Wallace F. Marshall, and Gerald Karp. Karp’s Cell and Molecular Biology: Concepts and Experiments. Principles of molecular biology by Burton E Tropp · Weaver, Robert Franklin, 1942-. Molecular biology / Robert F. Weaver Molecular Biology of the Gene. James D. Watson. Pearson Education

45. Thank you