title: DNA synthesis is required for reprogramming mediated by stem cell fusion.
abstract: Embryonic stem cells (ESCs) can instruct the conversion of differentiated cells toward pluripotency following cell-to-cell fusion by a mechanism that is rapid but poorly understood. Here, we used centrifugal elutriation to enrich for mouse ESCs at sequential stages of the cell cycle and showed that ESCs in S/G2 phases have an enhanced capacity to dominantly reprogram lymphocytes and fibroblasts in heterokaryon and hybrid assays. Reprogramming success was associated with an ability to induce precocious nucleotide incorporation within the somatic partner nuclei in heterokaryons. BrdU pulse-labeling experiments revealed that virtually all successfully reprogrammed somatic nuclei, identified on the basis of Oct4 re-expression, had undergone DNA synthesis within 24 hr of fusion with ESCs. This was essential for successful reprogramming because drugs that inhibited DNA polymerase activity effectively blocked pluripotent conversion. These data indicate that nucleotide incorporation is an early and critical event in the epigenetic reprogramming of somatic cells in experimental ESC-heterokaryons.
authors: Lymphocyte Development Group, MRC Clinical Sciences Centre, Imperial College London, Du Cane Road, London W12 0NN, UK.
authors: Tomomi
authors: T
authors: Tsubouchi
authors: Jorge
authors: J
authors: Soza-Ried
authors: Karen
authors: K
authors: Brown
authors: Francesco M
authors: FM
authors: Piccolo
authors: Irene
authors: I
authors: Cantone
authors: David
authors: D
authors: Landeira
authors: Hakan
authors: H
authors: Bagci
authors: Helfrid
authors: H
authors: Hochegger
authors: Matthias
authors: M
authors: Merkenschlager
authors: Amanda G
authors: AG
authors: Fisher
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology; D002459:Cell Fusion; D002467:Cell Nucleus / Q000378:metabolism; D065150:Cellular Reprogramming; D004261:DNA Replication; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism*; D051379:Mice; D009711:Nucleotides / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000378:metabolism
chemical_list: D009711:Nucleotides; D050814:Octamer Transcription Factor-3; C494174:Pou5f1 protein, mouse