title: Apoptosis and failure of checkpoint kinase 1 activation in human induced pluripotent stem cells under replication stress.
abstract: BACKGROUND
  Human induced pluripotent stem (hiPS) cells have the ability to undergo self-renewal and differentiation similarly to human embryonic stem (hES) cells. We have recently shown that hES cells under replication stress fail to activate checkpoint kinase 1 (CHK1). They instead commit to apoptosis, which appears to be a primary defense mechanism against genomic instability. It is not known whether the failure of CHK1 activation and activation of apoptosis under replication stress is solely a feature of hES cells, or if it is a feature that can be extended to hiPS cells.
  
  
  METHODS
  Here we generated integration-free hiPS cell lines by mRNA transfection, and characterised the cell lines. To investigate the mechanism of S phase checkpoint activation, we have induced replication stress by adding excess thymidine to the cell culture medium, and performed DNA content analysis, apoptosis assays and immunoblottings.
  
  
  RESULTS
  We are showing that hiPS cells similarly to hES cells, fail to activate CHK1 when exposed to DNA replication inhibitors and commit to apoptosis instead. Our findings also suggest the Ataxia Telangiectasia Mutated pathway might be responding to DNA replication stress, resulting in apoptosis.
  
  
  CONCLUSIONS
  Together, these data suggest that the apoptotic response was properly restored during reprogramming with mRNA, and that apoptosis is an important mechanism shared by hiPS and hES cells to maintain their genomic integrity when a replication stress occurs.
authors.affiliation: Centre for Stem Cell Biology, Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield, S10 2TN, UK. joelle.desmarais@mail.mcgill.ca.|Institute for Cancer Studies, Department of Oncology, School of Medicine, University of Sheffield, Beech Hill Road, Sheffield, S10 2RX, UK. joelle.desmarais@mail.mcgill.ca.
authors.forename: Joelle A
authors.initials: JA
authors.lastname: Desmarais
authors.affiliation: Centre for Stem Cell Biology, Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield, S10 2TN, UK. C.unger@sheffield.ac.uk.
authors.forename: Christian
authors.initials: C
authors.lastname: Unger
authors.affiliation: Department of Pathology, University of Kansas Hospital, 3901 Rainbow Blvd, Kansas City, KS, 66160, USA. idamjano@kumc.edu.
authors.forename: Ivan
authors.initials: I
authors.lastname: Damjanov
authors.affiliation: Institute for Cancer Studies, Department of Oncology, School of Medicine, University of Sheffield, Beech Hill Road, Sheffield, S10 2RX, UK. m.meuth@sheffield.ac.uk.
authors.forename: Mark
authors.initials: M
authors.lastname: Meuth
authors.affiliation: Centre for Stem Cell Biology, Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield, S10 2TN, UK. p.w.andrews@sheffield.ac.uk.
authors.forename: Peter
authors.initials: P
authors.lastname: Andrews
mesh_terms: D000818:Animals; D017209:Apoptosis; D000071877:Checkpoint Kinase 1; D004261:DNA Replication; D045325:HCT116 Cells; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000502:physiology*; D051379:Mice; D011494:Protein Kinases / Q000378:metabolism*
chemical_list: D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C000605869:Chek1 protein, mouse
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