DNA Recombination, Rearrangement Flashcards
Homologous recombination in meiosis
(6)
- ds break in one of two homologs is converted to a ds gap by exonucleases (5’ ends are shorter)
- exposed 3’OH searches for complement in intact homolog, displacing other strand in duplex
- DNA pol extends invading 3’ end
- Two cross overs result: Holliday intermediate
- DNA pol replaces DNA missinf from site of original ds break
- Specialized nucleases cleaves intermediates
- exchange of displaced region only
- exhange or regions outside of repair region
*Also linked to DNA synthesis to restore loss of genetic info at a break in chromosome
Homologous recombination in stalled replication forks
DNA lesion
- Lesion on leading strand template causes arrest; lagging strand pol continues until it can no longer move forward since tethered → replisome arrest
- RecA causes the fork to regress; parental strands reanneal and new strands regress on each other
- DNA pol I continues replication using new strand as template; passes lesion
- reverse branch migration; origin-independent replication start
Homologous recombination in stalled replication forks
DNA nick caused by nuclease
- Pol encounters nick and dissociates from template; ds break
- Exonuclease eats away at 5’ end → ss 3’end exposed
- RecA (rad51) binds SS DNA initiates search for homology in duplex DNA (3’OH end invades)→ D loop → forms replication fork structure → replication continues
* tumor suppressor p53 may play role in controlling homologous recombination promoted by rad51 - Branch migration forms Holliday structure behind replication fork
- RuvAB, RuvC resolves Holliday jxn and conserves genetic material
Translesion DNA synthesis/ Error-prone DNA polymerase
- Active site of pol are flexible, prone to making mistakes b/c low fidelity and lack of 3’-5’ proofreading activity
- Allows pol to go straight through the T-T dimer and accurately adds adenosine residues
- Replaced by high fidelity pol afterwards; method to avoid homologous recombination method
Role of BRCA2 in ds break repair
- BRCA2 assists loading rad51 onto SS DNA
- Forms filament proteins coating DNA
- Forms D loop to pair with sister chromatid (template)
- Disengage and pair
*occurs during G2/S phases before M phase
Mutations in tumor suppressor genes BRCA1 and BRCA2 lead to increased risk of breast cancer
Nonhomologous end joining
Occurs in somatic cells during G1 phase
- Ku binds to ends of DNA w ds break
- ku bridges ends
- processing enzymes fill in gap; ligase
*doesn’t occur frequently b/c cell is not in replication phase
*low fidelity
*no template sequence so information is lost
*if there is a deficiency in BCRA genes can’t do homologous recombindation and this pathway is used
Transposons
- DNA segments that can move from one position to another
- Parasitic: cell to cell, vertically and horizontally
- Mutagenic: inactivation, gene amplication, deletions, genome rearrangement
- bacterial elements can encode antibiotic resistance
- Transposases and integrases catalyze the integration of the transposable elements into new site
Conservative transposition
“cut and paste”
direct
- Transposases identify ends of transposons and cut out
- Transposases bound promote 3’Oh ends to insert itself into new strands and integrate into target
- DNA pol and ligase fill gap
Replicative transposition
&
Retrotransposition
- Nick is created, no ds break→ SS integration into new strand → fork like structure at left and right end and can initiate txn replication
- RNA intermediate is copied by reverse transcriptase to produce a DNA copy to be integrated into chromosome
- AIDS retrovirus
- retroposons
V(D)J Recombination
- Noncontiguous segments are brought together by recombination
- RAG proteins introduce ds breaks at sites to be joined recognize recombination ss (RSS) at ends of segments
- 4 J segments and 300 V segments
- first make a nick
- intramolecular transesterification → remove region → circularizes
- hairpin loop structures form at J and V ends
- During B cell differentiation, V region selected at random and one J region brought together (imprecise)
- Newly formed Ig gene is transcribed. RNA sequence btwn selected J region and C region is spliced out
- translated to become light chain in Ig
*these regions exist in hypermutable zones of the genome to increase variability
