DNA repair

DNA repair

1. DNA Repair Mohammed Fathy Bayomy, MSc, MD Lecturer Clinical Oncology & Nuclear Medicine Faculty of Medicine Zagazig University

2. Mutation Definition: any permanent & heritable change in DNA base sequence Classification * Germline mutation: mutation occurs in gamete cells, mutation will be passed on to all cells of body * Somatic mutation: mutation affects somatic cell it will be more restricted & passes to only descendents of that cell lineage in particular organ 1) According to cell type affected 2) According to structural changes * Point mutation: involves single base that substitute one amino acid for another * Deletions or insertions: remove or add one or more bases * Inversion mutation: inverts segment of DNA sequence

3. * Missense mutation: substitutes one amino acid for another thus resulting in abnormal protein * Nonsense mutation: substitutes a stop codon for amino acid codon leading to incomplete or truncated protein * Frameshift mutation: adds or deletes base with shift of base sequence, hence, changing reading frame, resulting in mistranslation of all amino acid sequence beyond mutation * Splicing mutation: will act on splice junction usually causing major changes in m-RNA & encoded protein 3) According to consequence of mutation on encoded protein

4. Pathogenesis * Mispairing of bases ( A-C pair instead of normal A-T pair) during DNA synthesis ( replication error ) or recombination * Slippage of newly synthesized DNA strand during/replication with loop formation is another replication error resulting in copying of bases twice * Loss of purine bases ( depurination ) occurs as a result of effect of body heat and cellular metabolism * De-amination of cytosine leading to C-T substitution 1) Spontaneous mutations

5. * Ultraviolet radiation produce pyrimidine dimers in which adjacent thymine residues in same DNA strand become covalently attached * Exposure to ionizing radiation may produce various lesions such as : base damage (e.g. conversion of thymine to thymine glycol or change of guanine to hydroxy guanine), single or double strand breaks in DNA phosphate-sugar backbone, and cross-links between the two strands of DNA or between DNA & proteins * Chemotherapeutic-agents may cause base damage (alkylating agents), strand breaks (bleomycin) or various adducts or cross-links (cisplatin) 2) Induced mutations

6. DNA Repair Cells have evolved an elaborate array of enzymatic systems to maintain integrity of their genetic material in the face of numerous agents that alter DNA structure or base sequence Cellular repair of macromolecule is known to occur only for DNA, vital molecule for cell survival Tumor suppressor gene p5 3 plays key role in this regard, hence it is called guardian of genome. Thus, with DNA injury, p53 causes arrest of cell cycle to give time for DNA repair enzymes. If DNA lesions are severe & irrepairable, p5 3 eliminates mutant cells by stimulating apoptosis

7. DNA Repair Systems Damaged DNA is reverted to its original state without replacement of constituents Example 1) Damage Reversal * Repair of base damage, by alkyl transferase enzyme , removing methyl & larger alkylating groups from DNA bases * Rejoining single strand break by ligase 2) Mismatch repair (MMR) Four types of mismatch repair genes : hMSH-2 (2p), hMCH-1 (3p), hPMS-1, hPMS-2 During replication, mismatch protein gene products act as "spell checkers" and if there is replication error (erroneous pairing of G-T instead of A-T), this system corrects defect

8. Mismatched lesion is recognized by hMSH-2 protein, or hMSH-2: GTBP/P160 protein heterodimer Removal of mismatch, followed by resynthesis & ligation completes repair process 3) Excision repair A) Base Excision Repair (BER) Limited to small lesions (de-aminated cytosine, single strand breaks Repair patches consist usually of only one or two nucleotides (short patch repair) Base damage is recognized & excised by glycosylase , removal of sugar residue by endonuclease & exonuclease , followed by resynthesis & ligation

9. b) Nucleotide Excision Repair (NER) Involves replacement of long patches of DNA of order of thirty nucleotides (long patch repair) There are 5 steps in NER process, namely: recognition of DNA lesion, incision of damaged strand on each side of lesion, removal of damaged nucleotides, synthesis of new nucleotides, & its ligation 4) Recombination repair Double strand breaks are most difficult to repair due to absence of template strand Unrepaired or misrepaired double strand breaks may lead to serious consequences: end-to-end joining of nonhomologous chromosomes, or formation of new telomeres generating chromosomal rearrangements deletions, use of nonhomologous DNA strands as template results in formation of abnormal base sequence (mutation)

10. Double strand breaks, in view of difficulty of repair, is most common cause of cell lethality following genotoxic agents Repair requires recruitment of homologous or heterologous DNA strands with formation of Holliday junction caused by crossing over of single strands from two adjacent DNA duplexes This junction requires resolution, by enzymatic cutting of crossing point, before repair & duplex separation are complete