A Cut above the Rest: Targeted Genome Editing Technologies in Human Pluripotent Stem Cells

Targeting DNA With Fingers and TALENs.

COSMID: A Web-based Tool for Identifying and Validating CRISPR/Cas Off-target Sites.

Gene Correction by Homologous Recombination With Zinc Finger Nucleases in Primary Cells From a Mouse Model of a Generic Recessive Genetic Disease

Expanding or Restricting the Target Site Repertoire of Zinc-finger Nucleases: The Inter-domain Linker as a Major Determinant of Target Site Selectivity

Highly Efficient and Marker-free Genome Editing of Human Pluripotent Stem Cells by CRISPR-Cas9 RNP and AAV6 Donor-Mediated Homologous Recombination.

Manipulating piggyBac Transposon Chromosomal Integration Site Selection in Human Cells

Therapeutic Translation of iPSCs for Treating Neurological Disease

Generation of an HIV Resistant T-cell Line by Targeted “Stacking” of Restriction Factors

Zfp206 (recently renamed Zscan10) encodes a zinc finger transcription factor specifically expressed in human and mouse embryonic stem cells (ESC). It has been shown that Zfp206 is required to maintain ESC in an undifferentiated, pluripotent state. Presented here are data showing that Zfp206 works together with two other transcription factors, Oct4 and Sox2, which are also essential regulators of ESC pluripotency. We show that Zfp206 binds to the Oct4 promoter and directly regulates Oct4 expression. Genome-wide mapping of Zfp206-binding sites in ESC identifies more than 3000 target genes, many of which encode transcription factors that are also targeted for regulation by Oct4 and Sox2. In addition, we show that Zfp206 physically interacts with both Oct4 and Sox2. These data demonstrate that Zfp206 is a key component of the core transcriptional regulatory network and together with Oct4 and Sox2 regulates differentiation of ESC.

Modeling human hematopoietic cell development from pluripotent stem cells

Pluripotent cells have the potential to differentiate into all of the cell types of an animal. This unique cell state is governed by an interconnected network of transcription factors. Among these, Oct4 plays an essential role both in the development of pluripotent cells in the embryo and in the self-renewal of its in vitro counterpart, embryonic stem (ES) cells. Furthermore, Oct4 is one of the four Yamanaka factors and its overexpression alone can generate induced pluripotent stem (iPS) cells. Recent reports underscore Oct4 as an essential regulator of opposing cell state transitions, such as pluripotency establishment and differentiation into embryonic germ lineages. Here we discuss these recent studies and the potential mechanisms underlying these contrasting functions of Oct4.

Embryonic stem cells are pluripotent and capable of unlimited self-renewal. Elucidation of the underlying molecular mechanism may contribute to the advancement of cell-based regenerative medicine. In the present work, we performed a large scale analysis of the phosphoproteome in mouse embryonic stem (mES) cells. Using multiplex strategies, we detected 4581 proteins and 3970 high confidence distinct phosphosites in 1642 phosphoproteins. Notably, 22 prominent phosphorylated stem cell marker proteins with 39 novel phosphosites were identified for the first time by mass spectrometry, including phosphorylation sites in NANOG (Ser-65) and RE1 silencing transcription factor (Ser-950 and Thr-953). Quantitative profiles of NANOG peptides obtained during the differentiation of mES cells revealed that the abundance of phosphopeptides and non-phosphopeptides decreased with different trends. To our knowledge, this study presents the largest global characterization of phosphorylation in mES cells. Compared with a study of ultimately differentiated tissue cells, a bioinformatics analysis of the phosphorylation data set revealed a consistent phosphorylation motif in human and mouse ES cells. Moreover, investigations into phosphorylation conservation suggested that phosphoproteins were more conserved in the undifferentiated ES cell state than in the ultimately differentiated tissue cell state. However, the opposite conclusion was drawn from this conservation comparison with phosphosites. Overall, this work provides an overview of phosphorylation in mES cells and is a valuable resource for the future understanding of basic biology in mES cells.

CRISPR/Cas9-Mediated Genome Editing Corrects Dystrophin Mutation in Skeletal Muscle Stem Cells in a Mouse Model of Muscle Dystrophy.

Engineering of Human Pluripotent Stem Cells by AAV-mediated Gene Targeting

A CRISPR Approach to Gene Targeting