IGEM:Harvard/2006/Brainstorming Papers - David Ramos

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Telomeres

  1. LeBel C and Wellinger RJ. . pmid:15976439. PubMed HubMed [t1]
  2. Murray AW, Claus TE, and Szostak JW. . pmid:3062364. PubMed HubMed [t2]
  3. Feld H, Navarro V, Kleeman H, and Shani J. . pmid:1458522. PubMed HubMed [t3]
All Medline abstracts: PubMed HubMed

Telomeres: What's new at your end?

  • Gives some general information on telomeres.
  • Telomeres exist at the end of DNA strands in yeast and human cells.
  • They protect DNA from being modified or degraded
    • Also prevent chromosomal ends from being recognized as DNA damage
    • Relies on proteins associated with the G-tails and proteins associated with the double-strand portion of the repeats.
  • Telomere structure is dynamic and udergoes dramatic changes during the cell cycle particularly during the S phase.
  • G-tails are the G-rich single-strand extensions at the very end of telomeres that, in species with particularly long G-tails (humans), invade and displace G-strands in the double-stranded portion of the telomere, creating a t-loop.
  • Budding yeast telomeres are irregular in length (TG)^1-6 TG^2-3, are about 300 bp long, and have a relatively short G-tail, however this can change, especially during S phase.
  • Telomeric repeat tract lengths are not precise, but rather centered around a genetically determined mean.
  • The telomorase holoenzyme seems to play a role in growing telomorase repeats
  • An active end elongation restriction process is thought to be involved in regulating telomere lengths. This involved the telomeric proteins involved in assembling the complexes on the double-strand repeats.
  • Entire process is still somewhat unknown, and "an emphasis of recent research has been on how this telomere-specific end replication becomes actively engaged and how it is regulated to maintain telomeres at lengths that are within the species-specific window."
  • Evidence has been found suggesting that telomeres form form the interior of the chromosome towards the ends.
    • The C-strand will always be synthesized by lagging-strand synthesis and the G-strand by leading-strand synthesis.
  • For telomere lengths to stay within certain bounds, an active end elongation complex must be halted.
    • "The precise molecular mechanisms involved in this regulatory step are nebulous."

Characterization of Two Telomeric DNA Processing Reactions in Saccharomyces cerevisiae

  • Telomeric DNA needs to overcome two problems:
    • A continuing decrease in chromosome length caused by incomplete replication.
    • The reactivity of DNA ends, which are subject to degradation by nucleases and to fusion by ligation.
  • (not too important) Telomere sequences are usually described with respect to the complementary C-rich strand.
  • In many species, long arrays of short repeats are found. However:
    • Oxytricha is unusual in having a shorter stretch of C^4A^4 repeats. The telomere structure is precisely defined: there are a fixed number of repeats, and in the end of the molecule is a double-stranded break with a 16-base-pair 3' overhang.
    • In Dictyostelium and in S. cerevisiae, the repeat unit is irregular.
    • In yeast and Tetrahymena, single-strand interruptions are found within the telomeric repeats on both the CA- and GT-rich strands.
  • "Telomeric DNA is subject to a novel DNA-processing activity that results in the addition of new telomeric sequence to the end of the DNA duplex." All telomeres appear to be subject to this activity.
  • In trypanosomes, the elongation reaction is particularly synchronous and regular telomere growth is observable.
  • "The efficiency of the elongation reaction suggests that it plays a central role in telomere replication." The experiments conducted in this paper support non-template-directed DNA synthesis in the elongation reaction and argue against a recombinatorial mechanism.
  • Experimental highlights:
    • The C^4A^2 telomeric repeat of Tetrahymena is an excellent substrate for the elongation reaction.
    • The unique DNA adjacent to the C^4A^2 repeats is not necessary for telomere function in yeast cells.
    • The amont of C^1-3A DNA which is added to a plasmid during the elongation reaction is not influenced by either the unique Tetrahymena DNA or the amount of C^4A^2 DNA at the end.
    • Alternating CA is also a substrate for the elongation reaction.
    • The alternating CA cluster is a substrate only when the CA-rich strand runs in the same direction as it does in a natural telomere.
  • Since the elongation reaction is so efficient, it needn't occur every cell cycle, unlike the resolution reaction. Therefore, "the structure of telomeres is not static but in dynamic equilibrium as the resul of the opposing actions of non-template-directed DNA synthesis on one hand and incomplete replication and exonucleolytic degradation on the other."
  • The constant average length of yeast telomeres implies a feedback mechanism which sense the length of telomeric DNA and reduces the extent of non-templat-directed DNA synthesis when the telomeric DNA exceeds a certain length.
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