Direct Quantitative Monitoring of Homology-Directed DNA Repair of Damaged Telomeres

Priyanka Verma, Robert L. Dilley, Melina T. Gyparaki, Roger A. Greenberg

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

5 Scopus citations

Abstract

Homology-directed DNA repair (HDR) is an evolutionary conserved mechanism that is required for genome integrity and organismal fitness across species. While a myriad of different factors and mechanisms are able to execute HDR, all forms necessitate common steps of DNA damage recognition, homology search and capture, and assembly of a DNA polymerase complex to conduct templated DNA synthesis. The central question of what determines HDR mechanism utilization in mammalian cells has been limited by an inability to directly monitor the DNA damage response and products of repair as they arise from a defined genomic lesion. In this chapter, we describe several methodologies to delineate major steps of HDR during alternative lengthening of telomeres in human cells. This includes procedures to visualize interchromosomal telomere homology searches in real time and quantitatively detect HDR synthesis of nascent telomeres emanating from synchronous activation of telomere DNA double-strand breaks. We highlight the critical details of these methods and their applicability to monitoring HDR at telomeres in a broad variety of mammalian cell types.

Original languageEnglish
Title of host publicationMethods in Enzymology
PublisherAcademic Press Inc.
Pages107-134
Number of pages28
DOIs
StatePublished - 2018

Publication series

NameMethods in Enzymology
Volume600
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988

Keywords

  • Alternative lengthening of telomeres
  • Cancer
  • DNA synthesis
  • Double-strand breaks
  • Homologous recombination
  • Telomeres

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