RRC ID 68793
Author Nishimura K, Kato T, Chen C, Oinam L, Shiomitsu E, Ayakawa D, Ohtaka M, Fukuda A, Nakanishi M, Hisatake K.
Title Manipulation of KLF4 expression generates iPSCs paused at successive stages of reprogramming.
Journal Stem Cell Reports
Abstract The detailed mechanism of reprogramming somatic cells into induced pluripotent stem cells (iPSCs) remains largely unknown. Partially reprogrammed iPSCs are informative and useful for understanding the mechanism of reprogramming but remain technically difficult to generate in a predictable and reproducible manner. Using replication-defective and persistent Sendai virus (SeVdp) vectors, we analyzed the effect of decreasing the expression levels of OCT4, SOX2, KLF4, and c-MYC and found that low KLF4 expression reproducibly gives rise to a homogeneous population of partially reprogrammed iPSCs. Upregulation of KLF4 allows these cells to resume reprogramming, indicating that they are paused iPSCs that remain on the path toward pluripotency. Paused iPSCs with different KLF4 expression levels remain at distinct intermediate stages of reprogramming. This SeVdp-based stage-specific reprogramming system (3S reprogramming system) is applicable for both mouse and human somatic cells and will facilitate the mechanistic analysis of reprogramming.
Volume 3(5)
Pages 915-29
Published 2014-11-11
DOI 10.1016/j.stemcr.2014.08.014
PII S2213-6711(14)00268-9
PMID 25418733
PMC PMC4235142
MeSH Animals Blotting, Western Cells, Cultured Cellular Reprogramming / genetics* Cluster Analysis Embryo, Mammalian / cytology Fibroblasts / cytology Fibroblasts / metabolism Gene Expression Profiling* Green Fluorescent Proteins / genetics Green Fluorescent Proteins / metabolism Homeodomain Proteins / genetics Homeodomain Proteins / metabolism Humans Induced Pluripotent Stem Cells / metabolism* Kruppel-Like Transcription Factors / genetics* Kruppel-Like Transcription Factors / metabolism Mice Mice, Inbred C57BL Mice, Transgenic NIH 3T3 Cells Nanog Homeobox Protein Octamer Transcription Factor-3 / genetics Octamer Transcription Factor-3 / metabolism Oligonucleotide Array Sequence Analysis Proto-Oncogene Proteins c-myc / genetics Proto-Oncogene Proteins c-myc / metabolism Reverse Transcriptase Polymerase Chain Reaction SOXB1 Transcription Factors / genetics SOXB1 Transcription Factors / metabolism
Resource
Human and Animal Cells NB1RGB(RCB0222)