DNA Repair DNA repair is a system used to correct DNA damage caused by either: 1-Errors during DNA replication including incorrect base-pairing (mismatching) e.g. G may be paired with T not with C. 2- Chemicals: such as A- deaminating agents (e.g. nitrous acid): nitrous acid deaminates cytosine into uracil (base replacement) B- alkylating agents that cause base alteration 3- UV radiatiom which can join two adjacent pyrimidines (pyromidine dimer formation) 4- high-energy radiation (e.g. x-ray) and oxidative free radicals, which can cause double-strand breaks.

If the damage is not repaired, a permanent mutation may be introduced that can result in number of deleterious effects leading to cancer. Luckily, cells are remarkably efficient at repairing damage done to DNA. Most of repair systems involve: 1- recognition of the of damage on DNA, 2-removal or excision of the damage, 3-replacement or filling the produced gap with new DNA and 4-ligation

repair (Repair of incorrect base pairing) A- Mismatch ( nucleotide excision repair) Causes of mismach: replication errors Repair Mismatch repair is mediated by a group of proteins known as the Mut proteins. Analogous proteins are present in humans. Steps of repair: 1.Identification of the mismatched strand: When mismatch occurs, the Mut proteins that identify the mis-paired base (s) must be able to differentiate between the correct strand and the strand with the mismatch. For example: in mis-paired G-T, Mut proteins should recognize whether G or T is the false base added in the new strand.

Differentian is based on the presence of GATC sequence in which adenine base is methylated, this unique sequence is found approximately once every thousand nucleotides. Methylation of adenine is not done immediately after replication, so the newly synthesized DNA is hemi-methylated i.e. parent strand is methylated, but the daughter new strand is not. The methylated parent strand is assumed to be correct, and the daughter strand that will be repaired.

Identification of the mismatched strand

.Repair of mismatch: - The mismatch is identified by Mut proteins. - An endonuclease nicks (cuts) the mismatched strand and the mismatched nucleotide(s) is/are removed. Extra nucleotides at the 5' and 3' ends of the mismatch are also removed. -The gap left after removal of the nucleotides is filled by DNA synthesized by DNA polymerase I using the parent strand as a template. -The nick is sealed by ligase. NB: A defect in mismatch repair system in humans has been shown to cause hereditary nonpolyposis colon cancer (HNPCC), one of the most common inherited cancers.

Mismatch repair Note that in this type of repair not only the incorrect base is removed but also its sugar and phosphate ( i.e. all the nucleotide), in addition to extra adjacent nucleotides, so this repair system is called nucleotide excision repair

B- Repair of damage caused by ultraviolet (UV) light (another example on nucleotide excision repair) : Exposure of a cell to UV light can result in the covalent joining of two adjacent pyrimidines (usually thymines), producing a dimer. These thymine dimers prevent DNA replication.

The UV-damage repair includes: -Recognition and excision (removal) of dimers by UV-specific endonuclease (called also exinuclease): This enzyme recognizes the dimer and some nucleotides including the dimer is released leaving a gap. - The gap is then filled by DNA polymerase I and the nick is sealed by ligase. Since some nucleotides are removed so it is a second example on “nucleotide excision repair”

UV radiation and cancer: Pyrimidine dimers can be formed in the skin cells of humans chronically exposed to sunlight. In the rare genetic disease xeroderma pigmentosum, the cells can’t repair the damaged DNA due to genetic defects in exinuclease enzyme (required for UV- damage repair.) The result is accumulation of mutations and, consequently, skin cancer. Symptoms: -severe sun burns after short exposure to sun light. - Development of many freckles -Irregular dark spots on the skin -Thin and very dry skin.

C. Correction of base alteration (base excision repair): Causes: - Spontenous: - Deaminating agents or alkylating agents The bases of DNA can be altered for example, cytosine can be deaminated (loss of amino group) to form uracil either spontaneously or by the action of deaminating or alkylating agents. For example, nitrous acid which is formed by the cell is a potent compound that deaminates cytosine into uracil.

Repair of cytosine deamination: 1.Recognition and Removal of uracil by specific glycosylase: Abnormal base (uracil), is recognized by uracil N- glycosylases that remove them from its deoxyribose-phosphate. This leaves an apyrimidinic site (AP site) i.e. Site free from pyrimidine base. Note: uracil only is removed from UMP, leaving deoxyribose- phosphate. So this repair is called base-excision repair.

2. Recognition of an AP site: Specific AP-endonucleases recognize that a base is missing and make a cut at AP site. 3. A deoxyribose phosphate lyase removes the deoxyribose phosphate 4. DNA polymerase I and DNA ligase complete the repair process.

DamageCauseRecognition/ removal enzymes Filling of gap and sealing nicks Method of repair Mismatched pairs DNA replication errors Mut proteins (its deficiency lead to hereditary nonpolyposis colon cancer (HNCC) DNA polymerase I, ligase Nucleotide excision repair Thymine dimer UV radiationExinuclease (its deficiency lead to xeropderma pigmentosum) DNA polymerase I, ligase Nucleotide excision repair Cytosine deamination Spontaneous OR Chemicals such as nitrous acid -Uracil glycosylase - AP endonuclease DNA polymerase I, ligase Base excision repair

D- Repair of double strand break Causes: High energy radiation (x-ray) or oxidative free radicals can cause double-strand breaks in DNA which are potentially lethal to the cell.

Repair of DNA double strand break: - The main repair system in humans is called: Nonhomolgous end- Joining (NHEJ) repair. -In this system the ends of two DNA fragments are brought together by a group of proteins that affect their religation. -This system does not require that the two DNA sequences have any sequence homomlgy. - Defect in this repair system lead to cancer and immune deficiency diseases

Study question Choose the one correct answer: A ten-year-old girl is brought to the dermatologist by her parents. She has many freckles on her face, neck, arms, and hands and the parents report that she is usually sensitive to sunlight. Two basal cell carcinoma are identified in her face. Which of the following processes is most likely to be defective in this patient? a) Repair of double-strand breaks b) Removal of mismatched bases from the 3’-end of daughter strand c) Removal of thymine dimers from DNA d) Removal of uracil from DNA.