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DNA Repair

DNA Repair. Drosophila BLM in Double-Strand Break Repair. Bloom Syndrome: Men & Flys. BLM is Bloom Syndrome Gene Predisposition to cancer BLM is a RecQ-type Helicase Involved in synthesis dependent DS-break repair

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DNA Repair

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  1. DNA Repair Drosophila BLM in Double-Strand Break Repair Karp/CELL & MOLECULAR BIOLOGY 3E

  2. Bloom Syndrome: Men & Flys • BLM is Bloom Syndrome Gene • Predisposition to cancer • BLM is a RecQ-type Helicase • Involved in synthesis dependent DS-break repair • Most double-strand break repair involves synthesis in flys (but not in plants, animals) Karp/CELL & MOLECULAR BIOLOGY 3E

  3. Synthesis Dep. Strand-annealing (SDSA) • DS-break, processed to 3’ ss-DNA end • Strand displacement and homolog annealing • Prime DNA synthesis • Gene conversion without crossing over • Unlike other forms of DS-break repair • Bloom Syndrome cells have increased cross-overs Karp/CELL & MOLECULAR BIOLOGY 3E

  4. Transposons make DS-breaks • Used P-elements and copia elements to generate DS-breaks • P-element engineered to contain white gene (red) • Copia insertion (orange) creates apricot allele • Yellow in single copy, apricot in two copies • Conversion to red “measures” SDSA • Single LTR in intron – near wild-type expression Karp/CELL & MOLECULAR BIOLOGY 3E

  5. Karp/CELL & MOLECULAR BIOLOGY 3E

  6. Interpreting Eye Colors... • Effect of BLM mutation • Causes fewer red eyes, fewer apricot, more yellow • Less SDSA (38 fold less), more aberrant repair • Why? Effect on synthesis during repair? • Little effect on fraction of extended ends, but... • Substantial effect on length of extended ends • Repair synthesis is significantly shorter (aborted?) Karp/CELL & MOLECULAR BIOLOGY 3E

  7. Karp/CELL & MOLECULAR BIOLOGY 3E

  8. Helicases and Repair • RecQ (E. coli) prevents association of microhomologies • Or, helicase could just facilitate synthesis • No effect on microhomology representation in repairs was observed • Some NHEJ after aborted synthesis? • Some templated additions as well • Likely multiple pathways for aberrant repair Karp/CELL & MOLECULAR BIOLOGY 3E

  9. Karp/CELL & MOLECULAR BIOLOGY 3E

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