5'-Cytosine-phosphoguanine (CpG) methylation impacts the activity of natural and engineered meganucleases

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

  • Julien Valton
  • Fayza Daboussi
  • Sophie Leduc
  • Rafael Molina
  • Pilar Redondo
  • Rachel Macmaster
  • Montoya, Guillermo
  • Philippe Duchateau

In this study, we asked whether CpG methylation could influence the DNA binding affinity and activity of meganucleases used for genome engineering applications. A combination of biochemical and structural approaches enabled us to demonstrate that CpG methylation decreases I-CreI DNA binding affinity and inhibits its endonuclease activity in vitro. This inhibition depends on the position of the methylated cytosine within the DNA target and was almost total when it is located inside the central tetrabase. Crystal structures of I-CreI bound to methylated cognate target DNA suggested a molecular basis for such inhibition, although the precise mechanism still has to be specified. Finally, we demonstrated that the efficacy of engineered meganucleases can be diminished by CpG methylation of the targeted endogenous site, and we proposed a rational design of the meganuclease DNA binding domain to alleviate such an effect. We conclude that although activity and sequence specificity of engineered meganucleases are crucial parameters, target DNA epigenetic modifications need to be considered for successful gene editions.

OriginalsprogEngelsk
TidsskriftThe Journal of Biological Chemistry
Vol/bind287
Udgave nummer36
Sider (fra-til)30139-50
Antal sider12
ISSN0021-9258
DOI
StatusUdgivet - 31 aug. 2012
Eksternt udgivetJa

ID: 203018819