TitleMicroRNA discovery and profiling in human embryonic stem cells by deep sequencing of small RNA libraries.
Publication TypeJournal Article
Year of Publication2008
AuthorsBar M, Wyman SK, Fritz BR, Qi J, Garg KS, Parkin RK, Kroh EM, Bendoraite A, Mitchell PS, Nelson AM, Ruzzo WL, Ware C, Radich JP, Gentleman R, Ruohola-Baker H, Tewari M
JournalStem cells (Dayton, Ohio)
Date or Month Published2008 Oct
KeywordsBase Sequence, Cell Differentiation, Cell Line, Databases, Genetic, Embryonic Stem Cells, Expressed Sequence Tags, Gene Expression Profiling, Gene Expression Regulation, Developmental, Gene Library, Humans, MicroRNAs, Molecular Sequence Data, Multipotent Stem Cells, Nucleic Acid Conformation, Pluripotent Stem Cells, Reproducibility of Results, Reverse Transcriptase Polymerase Chain Reaction, Ribonuclease III, RNA, Small Interfering, Sequence Analysis, RNA, Transcription Factors
AbstractWe used massively parallel pyrosequencing to discover and characterize microRNAs (miRNAs) expressed in human embryonic stem cells (hESC). Sequencing of small RNA cDNA libraries derived from undifferentiated hESC and from isogenic differentiating cultures yielded a total of 425,505 high-quality sequence reads. A custom data analysis pipeline delineated expression profiles for 191 previously annotated miRNAs, 13 novel miRNAs, and 56 candidate miRNAs. Further characterization of a subset of the novel miRNAs in Dicer-knockdown hESC demonstrated Dicer-dependent expression, providing additional validation of our results. A set of 14 miRNAs (9 known and 5 novel) was noted to be expressed in undifferentiated hESC and then strongly downregulated with differentiation. Functional annotation analysis of predicted targets of these miRNAs and comparison with a null model using non-hESC-expressed miRNAs identified statistically enriched functional categories, including chromatin remodeling and lineage-specific differentiation annotations. Finally, integration of our data with genome-wide chromatin immunoprecipitation data on OCT4, SOX2, and NANOG binding sites implicates these transcription factors in the regulation of nine of the novel/candidate miRNAs identified here. Comparison of our results with those of recent deep sequencing studies in mouse and human ESC shows that most of the novel/candidate miRNAs found here were not identified in the other studies. The data indicate that hESC express a larger complement of miRNAs than previously appreciated, and they provide a resource for additional studies of miRNA regulation of hESC physiology. Disclosure of potential conflicts of interest is found at the end of this article.
Downloadshttp://www.ncbi.nlm.nih.gov/pubmed/18583537?dopt=Abstract Abstract
Alternate JournalStem Cells
Citation Key1869
PubMed ID18583537