1. DNA R ECOMBINATION M ECHANISMS Fahareen Binta Mosharraf MNS 1

2. Genetic recombinationGenetic recombination the process by which genetic material is broken and joined to other genetic material Genetic exchange works constantly to blend and rearrange chromosome Most obvious during meiosis 2

3. Recombination ABCDEFGhijklmnoPQRSTUVWXYZ abcdefgHIJKLMNOpqrstuvwxyz ABCDEFGHIJKLMNOPQRSTUVWXYZ abcdefghijklmnopqrstuvwxyz 3

4. 4

5. Recombination between homologous DNA sites Recombination provides a means by which a genome can change to generate new combinations of genes Homologous recombination allows for the exchange of blocks of genes between homologous chromosomes and thereby is a mechanism for generating genetic diversity Recombination occurs randomly between two homologous sequences and the frequency of recombination between two sites is proportional to the distance between the sites 5

6. H OMOLOGOUS RECOMBINATION ( GENERAL RECOMBINATION ) Exchange of homologous regions between two DNA molecules. The exchange classically called crossing over Homologous recombination can be explained by Holliday model 6

7. T HE MAIN BIOLOGICAL ROLES OF HOMOLOGOUS RECOMBINATION (1) Inducing genetic variability through formation of recombinant DNA. During meiosis in eukaryotes, homologous recombination is needed for correct pairing of chromosomes ; it creates new combinations of genes which are transmitted to the descendants (2) Controlling DSB repair system because deleterious mutations would accumulate in each chromosome (3) Regulating gene expression by transferring genes between “dormant” or “active” sites in the genome 7

8. K EY STEPS OF HOMOLOGOUS RECOMBINATION 8

9. Holiday Junction for DNA recombination Exchange of the first single strand between two different DNA double helices is slow and difficult, then intermediate state Holiday Junction, then complete exchange

10. The Holliday model Two homologous duplexes are aligned Strand exchange leads to an intermediate with crossed strands This branch can move: Branch migration The branch is resolved by cleavage and sealing 10

11. HOMOLOGOUS RECOMBINATION PROCESS THROUGH DSB In reality, homologous recombination is often initiated by DSB. Meiotic crossing over is a result of this process 11

12. Double strand break model 12

13. Double strand break model 13

14. Double-strand breaks in DNA initiate recombination (cutting process) 14

15. Double-strand breaks in DNA initiate recombination (cutting process) 15

16. The cross-strand Holliday structure is an intermediate in recombination (SSB) 16

17. The cross-strand Holliday structure is an intermediate in recombination (SSB) 17

18. 18

19. Branch migration and resolution of Holliday structures depends on Ruv proteins 19

20. 20

21. DNA Branch Migration

22. N ON HOMOLOGOUS RECOMBINATION BY E ND J OINING 22

23. N ON HOMOLOGOUS END - JOINING (NHEJ) Broken ends are aligned and ends are trimmed or filled in, and their strands ligated. NHEJ in eukaryotes has several proteins: 1. Ku -heterodimer of Ku70 and Ku80. Ku binds to double-stranded breaks binding to DNA’s major and minor grooves. 2. DNA ligase IV and Xrcc4- Gaps filled in by DNA polymerases and sealed by DNA ligase IV and Xrcc4 3. Nucleotide trimming is carried out by ATP- dependent Mre11 complex.

24. End-joining repair of non homologous DNA 24