DNA Repair Flashcards
What are the main mechanisms for single strand repair?
Base excision repair, nucleotide excision repair and direct reversal of DNA damage.
What are the main mechanisms for double strand repair?
Homologous recombination and Nonhomologous end joining.
What is the process of Base Excision Repair?
Corrects most common types of DNA damage.
1) AP site (apurine or apyrimidine site indicates damage)
2) Glycosylase enzymes remove unwanted base
3) An endonuclease and phosphodiesterase cut the sugar-phosphate backbone and remove the sugar-phosphate residue.
4) Gap is filled by resynthesis with DNA polymerase I and nick sealed by ligase.
Backbone intact.
What is the process of Nucleotide Excision Repair and give an example of a disorder associated with defective NER?
Removes and resynthesizes a large patch around the damage, rather than just a single base as in BER. The sugar-phosphate backbone is cleaved at site of damage, exonucleases remove a large stretch of surrounding DNA and gap filled by resynthesis and sealed by DNA ligase.
mediated by UVRB/C/D complex (UVR C is endonuclease) and UVRD is helicase to separate bonds and lesion.
eg Xeroderma Pigemntosum unable to repair UV damage.
When is nucleotide excision repair used?
Removes thymine dimers caused by UV radiation.
When is Homologous Recombination used?
To repair complex DNA damage including DSB caused by exogenous sources eg radiation or cellular sources such as oxidative damage.
Describe the process of Homologous Recombination?
Two sub-pathways. Synthesis-dependent strand annealing (SDSA) and double-strand break repair. Both pathways share the same initial steps. Following DNA damage, DSB are recognised and processed to 3’OH ssDNA tails. The recombinase enzyme RAD51 is loaded onto the ssDNA tail (presynaptic filament) which then searches for a homologous chromosome to be used as a template for repair. The presynaptic filament invades and pairs with the homologous DNA which in turn initiates strand exchange (BRCA2 involved). This generates a D-loop which can be dissolved using either pathway.
SDSA - produces only noncrossover products. The D-loop is unwound and the freed invading ssDNA strand anneals with the complementary DNA strand at the other end of the DSB.
DSBR - proceeds by engaging the second end of the DSB producing an intermediate that harbours two Holliday Junctions. The resolution of HJ horizontally results in noncrossover products, vertically produces crossover products.
Gap filling DNA synthesis and ligation.
BRCA2 thought to regulate availability of RAD51. BRCA2 binds RAD51 through its BRC repeats and directs it to dsDNA-ssDNA junction of DSB.
Describe the process of nonhomologous End Joining?
Does not require a template. Large multiporptein complexes are assembled at broken ends of DNA molecules and DNA ligase joins the ends regardless of their sequence. Desperate measure likely to cause mutations or rearrangements but better than breaks.
When is Mismatch Repair used?
Corrects mismatches caused by replication error (1 in 10(6)). Cells defective in MMR have mutation rates 100-1000 fold normal with a tendency to replication slippage in homopolymeric runs.
Describe the process of MMR?
1) MutS recognises mismatch (MutS = MSH2/MSH6 dimer).
2) MutS translocates along the DNA and binds the MutL dimer (PMS2/MLH1) which binds to methylated strand (parent strand) and brings daughter strand in to position to make a cut.
4) MutH makes the cut.
5) MutH recruits uvrD helicase which cleaves the small section around the mismatch
6) Gap filled by new nucleotides by DNA pol I
6) Nick sealed by ligase
Provide examples of disease of inefficient DSB repair?
Fanconi Anaemia - at least 15 genes in FA pathway send proteins to area of damage. Defective intrastrand cross-links build up as DNA is not repaired which result in uncontrolled cell growth in fast dividing cells eg bone marrow. Increased risk of cancers, bone marrow failure.
Nijmegan Breakage Syndrome - short stature, immunodeficiency, increased risk of cancer, NBN gene involved in HR.
