Telomeres 1 & 2 Flashcards
Why are telomeres needed?
Telomeres are buffer sequences at the ends of chromosomes. They are required because a small fragment of DNA is removed every time the chromosome is replicated.
- When the lagging strand is replicated, Okazaki fragments are produced and subsequently ligated to one another. The process of generating okazaki fragments requires a primer binding to the 5’ of the sequence which is to be amplified.
- The telomeres act as caps so that the DNA repair machinery doesn’t recognize the chromosome ends as double-stranded breaks. W/o telomeres, the chromosomes would fuse.
What are normal histone-DNA complexes called and what are telomeric DNA-histone complexes called?
Nucleosomes and telosomes respectively.
What’s a t-loop?
A t-loop is when the 3’ strand of the telomere invades itself, making it inaccessible for telomerase or any other protein. A sub-structure of the t-loop is the d-loop, which is the interaction between the 3’ invading strand and the 5’ acceptor strand. Structurally it reminds me of a Holliday junction.
The t-loop is generated by “the sheltering complex” consisting of TRF1/2.
What’s a possible explanation for how telomerase is such a processive enzyme?
Telomerase is found as dimers. It’s possible that the dimers coordinate their binding and releasing of telomeres in order to increase processivity.
In vivo, what proteins are present within the telomerase holoenzyme?
EST1, EST2, EST3
Describe the context in which telomerase operates.
- Telomerase reverse transcriptase domain (TERT), the domain that does the elongating of telomeres.
- Telomerase-associated RNA, it is used as a scaffolding for recruitment of the other essential proteins (EST1, EST2, EST3).
Why is RAP1 important in the context of telomeres?
RAP1 (repressor/activator protein 1) binds the telomeres. When several RAP1s are bound adjacent to each other, telomerase will extend the telomere.
RAP1 also protects the ssDNA/dsDNA junctions from adverse exonucleases.
Make sense of this: For some reason there’s an inverse relationship between RAP1 binding sites and telomeric length. Perhaps it’s the ratio between bound / unbound RAP1 sequences that matter.
Is it true that telomeres have 5’ overhangs?
No, they have 3’ overhangs. The length isn’t conserved throughout organisms, humans have 100-300 bp 3’ overhang.
How does telomere extension work?
The telomerase-associated RNA has an overhang which will bind the 3’ overhang of the relevant telomere. The telomerase-associated RNA will then provide an overhang onto which the telomere can be lengthened.
Since both telomerase and the telomerase-associated RNA are expressed in most cell lines, why are we telomerase deficient in all but the germ cell lines?
The telomerase RNA is alternatively spliced into being defective. Cancers can point mutate the splice sites which makes hTERT defect, allowing for its functional transcription and translation.
The telomerase-associated RNA doesn’t partake in any telomeric processes in the absence of telomerase.
What happens if telomeres are transcribed?
The telomere transcripts are referred to as TERRA transcripts. These can either repress or enhance telomerase activity, the mechanism is unknown.
Humans and other vertebrates host a telomeric t-loop which makes the telomeres non-reactive. What analogous mechanism does yeast have in its place?
Instead of a t-loop, yeast has a foldback loop in which SIR2, SIR3, and SIR5 interact with RAP1.
Both t-loops and the foldback structure have the same effect. Interestingly, they can fold over genes that are located in the chromosomal “normal” region, leading to gene silencing. Since this length of the telomeres allows for further reach, it makes sense that shorter telomeres are less proficient at silencing genes.
What’s the cellular reaction to a chromosome’s telomere becoming too short?
Senescence.
How does telomere elongation work in yeast?
- Telomerase is recruited intra-telomerically.
- Telomere binds Ku-proteins (complexed w/ SIR4).
- Telomerase is translocated to the telomeric end during the cell cycle.
- TEL1 phosphorylates CDC13 which frees it from its complex w/ STN1 and TEN1.
- Telomere lengthening may proceed at intersititial telomeric sequences (ITSs)
How does alternative lengthening of telomeres work?
Homologous recombination.
This needs to be complemented.
“In drosophila, there are hetA and TART regions - these are retrotransposons which are important for telomere lengthening. ‘gag’ deriving from HeT-A binds the polyA (of the telomerase-associated RNA??) and transports it to the chromosomal end where telomere can be lengthened. In drosophila there are five proteins which complex (hiphop, HOAP, moi, ver and tea). They bind the telomeric ends sequence-independently. HipHop is important for de novo telomere formation and the reparation of dsDNA breaks. The current model for drosophila telomere lengthening is that hiphop binds broken non-telomeric ends and recruits the remaining 4 proteins.”
