Harpers

336 / CHAPTER 36Table 36–9. Mechanism of DNA repairMechanism Problem SolutionMismatch Copying errors (single Methyl-directedrepair base or two- to five- strand cutting, exobaseunpaired loops) nuclease digestion,and replacementBase Spontaneous, chem- Base removal by N-excision- ical, or radiation dam- glycosylase, abasicrepair age to a single base sugar removal, replacementNucleotide Spontaneous, chem- Removal of an apexcision-ical, or radiation dam- proximately 30-repair age to a DNA segment nucleotide oligomerand replacementDouble- Ionizing radiation, Synapsis, unwindstrandchemotherapy, ing, alignment,break repair oxidative free ligationradicals3′5′3′5′3′5′CH 3CH 3CH 3 CH 3CH 3 CH 35′3′SINGLE-SITE STRAND CUTBY GATC ENDONUCLEASE5′3′DEFECT REMOVEDBY EXONUCLEASE5′3′DEFECT REPAIREDBY POLYMERASECH 3 CH 33′ 5′Mismatch RepairMismatch repair corrects errors made when DNA iscopied. For example, a C could be inserted opposite anA, or the polymerase could slip or stutter and insert twoto five extra unpaired bases. Specific proteins scan thenewly synthesized DNA, using adenine methylationwithin a GATC sequence as the point of reference (Figure36–22). The template strand is methylated, and thenewly synthesized strand is not. This difference allowsthe repair enzymes to identify the strand that containsthe errant nucleotide which requires replacement. If amismatch or small loop is found, a GATC endonucleasecuts the strand bearing the mutation at a site correspondingto the GATC. An exonuclease then digeststhis strand from the GATC through the mutation, thusremoving the faulty DNA. This can occur from eitherend if the defect is bracketed by two GATC sites. Thisdefect is then filled in by normal cellular enzymes accordingto base pairing rules. In E coli, three proteins(Mut S, Mut C, and Mut H) are required for recognitionof the mutation and nicking of the strand. Othercellular enzymes, including ligase, polymerase, andSSBs, remove and replace the strand. The process issomewhat more complicated in mammalian cells, asabout six proteins are involved in the first steps.Faulty mismatch repair has been linked to hereditarynonpolyposis colon cancer (HNPCC), one of themost common inherited cancers. Genetic studies linkedHNPCC in some families to a region of chromosome2. The gene located, designated hMSH2, was subsequentlyshown to encode the human analog of the3′5′CH 3 CH 3RELIGATEDBY LIGASEFigure 36–22. Mismatch repair of DNA. This mechanismcorrects a single mismatch base pair (eg, C to Arather than T to A) or a short region of unpaired DNA.The defective region is recognized by an endonucleasethat makes a single-strand cut at an adjacent methylatedGATC sequence. The DNA strand is removedthrough the mutation, replaced, and religated.E coli MutS protein that is involved in mismatch repair(see above). Mutations of hMSH2 account for 50–60%of HNPCC cases. Another gene, hMLH1, is associatedwith most of the other cases. hMLH1 is the human analogof the bacterial mismatch repair gene MutL. Howdoes faulty mismatch repair result in colon cancer? Thehuman genes were localized because microsatellite instabilitywas detected. That is, the cancer cells had a microsatelliteof a length different from that found in thenormal cells of the individual. It appears that the affectedcells, which harbor a mutated hMSH2 orhMLH1 mismatch repair enzyme, are unable to removesmall loops of unpaired DNA, and the microsatellitethus increases in size. Ultimately, microsatellite DNAexpansion must affect either the expression or the functionof a protein critical in surveillance of the cell cyclein these colon cells.5′3′