Chromatin-modifying enzymes as modulators of reprogramming

Generation of induced pluripotent stem cells (iPSCs) by somatic cell reprogramming involves global epigenetic remodelling. Whereas several proteins are known to regulate chromatin marks associated with the distinct epigenetic states of cells before and after reprogramming, the role of specific chromatin-modifying enzymes in reprogramming remains to be determined. To address how chromatin-modifying proteins influence reprogramming, we used short hairpin RNAs (shRNAs) to target genes in DNA and histone methylation pathways, and identified positive and negative modulators of iPSC generation. Whereas inhibition of the core components of the polycomb repressive complex 1 and 2, including the histone 3 lysine 27 methyltransferase EZH2, reduced reprogramming efficiency, suppression of SUV39H1, YY1 and DOT1L enhanced reprogramming. Specifically, inhibition of the H3K79 histone methyltransferase DOT1L by shRNA or a small molecule accelerated reprogramming, significantly increased the yield of iPSC colonies, and substituted for KLF4 and c-Myc (also known as MYC). Inhibition of DOT1L early in the reprogramming process is associated with a marked increase in two alternative factors, NANOG and LIN28, which play essential functional roles in the enhancement of reprogramming. Genome-wide analysis of H3K79me2 distribution revealed that fibroblast-specific genes associated with the epithelial to mesenchymal transition lose H3K79me2 in the initial phases of reprogramming. DOT1L inhibition facilitates the loss of this mark from genes that are fated to be repressed in the pluripotent state. These findings implicate specific chromatin-modifying enzymes as barriers to or facilitators of reprogramming, and demonstrate how modulation of chromatin-modifying enzymes can be exploited to more efficiently generate iPSCs with fewer exogenous transcription factors. © 2012 Macmillan Publishers Limited. All rights reserved

The Pediatric Cell Atlas: defining the growth phase of human development at single-cell resolution

Single-cell gene expression analyses of mammalian tissues have uncovered profound stage-specific molecular regulatory phenomena that have changed the understanding of unique cell types and signaling pathways critical for lineage determination, morphogenesis, and growth. We discuss here the case for a Pediatric Cell Atlas as part of the Human Cell Atlas consortium to provide single-cell profiles and spatial characterization of gene expression across human tissues and organs. Such data will complement adult and developmentally focused HCA projects to provide a rich cytogenomic framework for understanding not only pediatric health and disease but also environmental and genetic impacts across the human lifespan

Radiometric Calibration Targets for the Mastcam-Z Camera on the Mars 2020 Rover Mission

The Mastcam-Z Camera is a stereoscopic, multispectral camera with zoom capability on NASA’s Mars-2020 Perseverance rover. The Mastcam-Z relies on a set of two deck-mounted radiometric calibration targets to validate camera performance and to provide an instantaneous estimate of local irradiance and allow conversion of image data to units of reflectance (R or I/F) on a tactical timescale. Here, we describe the heritage, design, and optical characterization of these targets and discuss their use during rover operations. The Mastcam-Z primary calibration target inherits features of camera calibration targets on the Mars Exploration Rovers, Phoenix and Mars Science Laboratory missions. This target will be regularly imaged during flight to accompany multispectral observations of the martian surface. The primary target consists of a gold-plated aluminum base, eight strong hollow-cylinder SmCo alloy permanent magnets mounted in the base, eight ceramic color and grayscale patches mounted over the magnets, four concentric, ceramic grayscale rings and a central aluminum shadow post (gnomon) painted with an IR-black paint. The magnets are expected to keep the central area of each patch relatively free of Martian aeolian dust. The Mastcam-Z secondary calibration target is a simple angled aluminum shelf carrying seven vertically mounted ceramic color and grayscale chips and seven identical, but horizontally mounted ceramic chips. The secondary target is intended to augment and validate the calibration-related information derived from the primary target. The Mastcam-Z radiometric calibration targets are critically important to achieving Mastcam-Z science objectives for spectroscopy and photometric properties.With funding from the Spanish government through the "María de Maeztu Unit of Excellence" accreditation (MDM-2017-0737