B1 SINE-binding ZFP266 impedes mouse iPSC generation through suppression of chromatin opening mediated by reprogramming factors

Induced pluripotent stem cell (iPSC) reprogramming is inefficient and understanding the molecular mechanisms underlying this inefficiency holds the key to successfully control cellular identity. Here, we report 24 reprogramming roadblock genes identified by CRISPR/Cas9-mediated genome-wide knockout (KO) screening. Of these, depletion of the predicted KRAB zinc finger protein (KRAB-ZFP) Zfp266 strongly and consistently enhances murine iPSC generation in several reprogramming settings, emerging as the most robust roadblock. We show that ZFP266 binds Short Interspersed Nuclear Elements (SINEs) adjacent to binding sites of pioneering factors, OCT4 (POU5F1), SOX2, and KLF4, and impedes chromatin opening. Replacing the KRAB co-suppressor with co-activator domains converts ZFP266 from an inhibitor to a potent facilitator of iPSC reprogramming. We propose that the SINE-KRAB-ZFP interaction is a critical regulator of chromatin accessibility at regulatory elements required for efficient cellular identity changes. In addition, this work serves as a resource to further illuminate molecular mechanisms hindering reprogramming.Induced pluripotent stem cell (iPSC) reprogramming is inherently inefficient. Here the authors identify 24 reprogramming roadblock genes through a CRISPR/Cas9-mediated genome-wide knockout screen including a KRAB-ZFP Zfp266, knockout of which consistently enhances murine iPSC generation.Peer reviewe

Constitutively active Smad2/3 are broad scope potentiators of transcription factor-mediated cellular reprogramming

Reprogramming of cellular identity using exogenous expression of transcription factors (TFs) is a powerful and exciting tool for tissue engineering, disease modeling, and regenerative medicine. However, generation of desired cell types using this approach is often plagued by inefficiency, slow conversion, and an inability to produce mature functional cells. Here, we show that expression of constitutively active SMAD2/3 significantly improves the efficiency of induced pluripotent stem cell (iPSC) generation by the Yamanaka factors. Mechanistically, SMAD3 interacts with reprogramming factors and co-activators and co-occupies OCT4 target loci during reprogramming. Unexpectedly, active SMAD2/3 also markedly enhances three other TF-mediated direct reprogramming conversions, from B cells to macrophages, myoblasts to adipocytes, and human fibroblasts to neurons, highlighting broad and general roles for SMAD2/3 as cell-reprogramming potentiators. Our results suggest that co-expression of active SMAD2/3 could enhance multiple types of TF-based cell identity conversion and therefore be a powerful tool for cellular engineering. Ruetz et al. show that constitutively active SMAD2/3 has a surprising ability to boost the efficiency of cell reprogramming both to iPSCs and across lineages and may therefore be a general factor that can enhance transcription-factor-mediated reprogramming in a variety of contexts

Copper( i )-photocatalyzed trifluoromethylation of alkenes

International audienc

Structure, thermal and mechanical properties of poly (ε-caprolactone)/organomodified clay bionanocomposites prepared in open air by in situ polymerization

The first example of the usefulness of titanium (IV) butoxide, as initiators for open air in-situ intercalative polymerization of ε-caprolactone (ε-CL) in the presence of organoclay is herein reported. The bionanocomposites based on poly(ε-caprolactone) (PCL) were prepared by in-situ Ring Opening Polymerization (ROP) of ε-caprolactone in the presence of different organomodified montmorillonite clay (ODA-MMT) loading (1, 3 and wt%). Structural, thermal and mechanical characterizations of the resulting bionanocomposites were investigated. The presence of the nanoclay increased PCL crystallinity, melting temperature and thermal stability, whereas some decrease in T was observed. TEM analyses confirmed the good dispersion of ODA-MMT with 1 and 3 wt% content into the PCL polymer as already asserted by XRD diffraction. Finally, the Young's modulus of the PCL nanocomposites was higher compared to the neat PCL, while a decrease of stress and strain at break for materials with different filler content was observed.Financial support for cooperative research came from the Hassan II Academy of Science and Technology of Morocco/CSIC-Spain (Project AH11STC-nano 2011 – 2012 and 2010MA003), the MESRSFC and CNRST of Morocco (PPR program), and the bilateral scientific committee programs of collaboration between Morocco and Tunisia (n 17/TM/20), and the Spanish Ministry of Science and Innovation (MICINN) through the project MAT2016-81138-R

Constitutively active SMAD2/3 are broad-scope potentiators of transcription-factor-mediated cellular reprogramming

Reprogramming of cellular identity using exogenous expression of transcription factors (TFs) is a powerful and exciting tool for tissue engineering, disease modeling, and regenerative medicine. However, generation of desired cell types using this approach is often plagued by inefficiency, slow conversion, and an inability to produce mature functional cells. Here, we show that expression of constitutively active SMAD2/3 significantly improves the efficiency of induced pluripotent stem cell (iPSC) generation by the Yamanaka factors. Mechanistically, SMAD3 interacts with reprogramming factors and co-activators and co-occupies OCT4 target loci during reprogramming. Unexpectedly, active SMAD2/3 also markedly enhances three other TF-mediated direct reprogramming conversions, from B cells to macrophages, myoblasts to adipocytes, and human fibroblasts to neurons, highlighting broad and general roles for SMAD2/3 as cell-reprogramming potentiators. Our results suggest that co-expression of active SMAD2/3 could enhance multiple types of TF-based cell identity conversion and therefore be a powerful tool for cellular engineering.This work was supported by the ERC (grants ROADTOIPS 261075 to K.K. and BRAINCELL 261063 to S.L. and iN-Brain 309712 to M.P.), the BBSRC (project grant BB/L023474/1 to K.K.), the Anne Rowling Regenerative Neurology Clinic (K.K.), the Swedish Research Council (grant STARGET to S.L. and grants 521-2012-5624 and 521-2013-3347 to M.P.), and the Wellcome Trust (grant WT098051 to K.Y.). T.R., L.T., J.A., and D.F.K. are funded by a Darwin Trust scholarship, a CMVM scholarship, and Principal's Career Development scholarship from the University of Edinburgh, respectively. D.F.K. is supported by the BBSRC (EASTBIO doctoral training partnership). T.V.T. is supported by a Juan de la Cierva postdoctoral fellowship (MINECO, FJCI-2014-22946). B.D.S. was supported by an EMBO long-term fellowship (ALTF 1143-2015). K.K. is an MRC senior non-clinical fellow (MR/N008715/1). M.P. is a New York Stem Cell Foundation Robertson Investigator

Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells

International audienceDifferentiated macrophages can self-renew in tissues and expand long term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network that controls self-renewal. Single-cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells