title: Proteomic analysis of early reprogramming events in murine somatic cells incubated with Xenopus laevis oocyte extracts demonstrates network associations with induced pluripotency markers.
abstract: The reprogramming of somatic cells into a pluripotent/embryonic-like state holds great potential for regenerative medicine, bypassing ethical issues associated with embryonic stem cells (ESCs). Numerous methods, including somatic cell nuclear transfer (SCNT), fusion to pluripotent cells, the use of cell extracts, and expression of transcription factors, have been used to reprogram cells into ES-like cells [termed induced pluripotent stem cells (iPSCs)]. This study investigated early events in the nuclei of permeabilized murine somatic cells incubated in cytoplasmic extract prepared from Xenopus laevis germinal vesicle-stage oocytes by identifying proteins that showed significant quantitative changes using proteomic techniques. A total of 69 protein spots from two-dimensional electrophoresis were identified as being significantly altered in expression after treatment, and 38 proteins were identified by tandem mass spectrometry. Network analysis was used to highlight pathway connections and interactions between these identified proteins, which were found to be involved in many functions--primarily nuclear structure and dynamics, transcription, and translation. The pluripotency markers Klf4, c-Myc, Nanog, and POU5F1 were highlighted by the interaction network analysis, as well as other compounds/proteins known to be repressed in pluripotent cells [e.g., protein kinase C (PRKC)] or enhanced during differentiation of ESCs (e.g., retinoic acid). The network analysis also indicated additional proteins and pathways potentially involved in early reprogramming events.
authors.affiliation: Division of Animal Sciences, School of Biosciences, University of Nottingham, Loughborough, LE12 5RD, UK. Alex.Rathbone@nottingham.ac.uk
authors.forename: Alex J
authors.initials: AJ
authors.lastname: Rathbone
authors.forename: Susan
authors.initials: S
authors.lastname: Liddell
authors.forename: Keith H S
authors.initials: KH
authors.lastname: Campbell
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry / Q000494:pharmacology*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000187:drug effects* / Q000502:physiology; D005260:Female; D057026:Induced Pluripotent Stem Cells / Q000187:drug effects* / Q000378:metabolism; D000090062:Kruppel-Like Factor 4; D053858:Metabolic Networks and Pathways / Q000187:drug effects*; D051379:Mice; D009865:Oocytes / Q000737:chemistry; D020543:Proteome / Q000032:analysis* / Q000187:drug effects; D040901:Proteomics; D013696:Temperature; D014982:Xenopus laevis
chemical_list: D015415:Biomarkers; D002457:Cell Extracts; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D020543:Proteome