Heterochromatin protein ERH represses alternative cell fates during early mammalian differentiation.

During development, H3K9me3 heterochromatin is dynamically rearranged, silencing repeat elements and protein coding genes to restrict cell identity. Enhancer of Rudimentary Homolog (ERH) is an evolutionarily conserved protein originally characterized in fission yeast and recently shown to be required for H3K9me3 maintenance in human fibroblasts, but its function during development remains unknown. Here, we show that ERH is required for proper segregation of the inner cell mass and trophectoderm cell lineages during mouse development by repressing totipotent and alternative lineage programs.

Pretreatment with IL-15 and IL-18 rescues natural killer cells from granzyme B-mediated apoptosis after cryopreservation.

Human natural killer (NK) cell-based therapies are under assessment for treating various cancers, but cryopreservation reduces both the recovery and function of NK cells, thereby limiting their therapeutic feasibility. Using cryopreservation protocols optimized for T cells, here we find that ~75% of NK cells die within 24 h post-thaw, with the remaining cells displaying reduced cytotoxicity. Using CRISPR-Cas9 gene editing and confocal microscopy, we find that cryopreserved NK cells largely die via apoptosis initiated by leakage of granzyme B from cytotoxic vesicles.

Human embryo live imaging reveals nuclear DNA shedding during blastocyst expansion and biopsy.

Proper preimplantation development is essential to assemble a blastocyst capable of implantation. Live imaging has uncovered major events driving early development in mouse embryos; yet, studies in humans have been limited by restrictions on genetic manipulation and lack of imaging approaches. We have overcome this barrier by combining fluorescent dyes with live imaging to reveal the dynamics of chromosome segregation, compaction, polarization, blastocyst formation, and hatching in the human embryo.

Shedding light on stem cells: Optogenetics uncover the role of ERK dynamics in pluripotency.

In this issue of Developmental Cell, Arekatla et al. use optogenetic technologies to dissect the roles of ERK and AKT dynamics in pluripotency. They show how mouse embryonic stem cells can retain memory of signaling events controlling their fate.

The nuclear lamina couples mechanical forces to cell fate in the preimplantation embryo via actin organization.

During preimplantation development, contractile forces generated at the apical cortex segregate cells into inner and outer positions of the embryo, establishing the inner cell mass (ICM) and trophectoderm. To which extent these forces influence ICM-trophectoderm fate remains unresolved. Here, we found that the nuclear lamina is coupled to the cortex via an F-actin meshwork in mouse and human embryos.

Neurulation of the cynomolgus monkey embryo achieved from 3D blastocyst culture.

Neural tube (NT) defects arise from abnormal neurulation and result in the most common birth defects worldwide. Yet, mechanisms of primate neurulation remain largely unknown due to prohibitions on human embryo research and limitations of available model systems. Here, we establish a three-dimensional (3D) prolonged in vitro culture (pIVC) system supporting cynomolgus monkey embryo development from 7 to 25 days post-fertilization.

Updates on preimplantation embryo research.

Preimplantation development is the only stage of human development that can be studied outside the body in real time, as human embryos can be produced by in vitro fertilization and cultured in the laboratory as self-contained structures until the blastocyst stage. Here, we focus some of the key cellular and morphogenetic processes by which the 1-cell embryo is transformed gradually into a blastocyst ready for implantation. Although most of our knowledge about the dynamic series of events patterning preimplantation human development derives from work in mouse embryos, we discuss key differences that could exist with humans.

Sequential changes in cellular properties accompanying amniote somite formation.

Somites are transient structures derived from the pre-somitic mesoderm (PSM), involving mesenchyme-to-epithelial transition (MET) where the cells change their shape and polarize. Using Scanning electron microscopy (SEM), immunocytochemistry and confocal microscopy, we study the progression of these events along the tail-to-head axis of the embryo, which mirrors the progression of somitogenesis (younger cells located more caudally). SEM revealed that PSM epithelialization is a gradual process, which begins much earlier than previously thought, starting with the dorsalmost cells, then the medial ones, and then, simultaneously, the ventral and lateral cells, before a somite fully separates from the PSM.

The trophectoderm acts as a niche for the inner cell mass through C/EBPα-regulated IL-6 signaling.

IL-6 has been shown to be required for somatic cell reprogramming into induced pluripotent stem cells (iPSCs). However, how Il6 expression is regulated and whether it plays a role during embryo development remains unknown. Here, we describe that IL-6 is necessary for C/EBPα-enhanced reprogramming of B cells into iPSCs but not for B cell to macrophage transdifferentiation.

A monoastral mitotic spindle determines lineage fate and position in the mouse embryo.

During mammalian development, the first asymmetric cell divisions segregate cells into inner and outer positions of the embryo to establish the pluripotent and trophectoderm lineages. Typically, polarity components differentially regulate the mitotic spindle via astral microtubule arrays to trigger asymmetric division patterns. However, early mouse embryos lack centrosomes, the microtubule-organizing centres (MTOCs) that usually generate microtubule asters.

The embryonic node behaves as an instructive stem cell niche for axial elongation.

In warm-blooded vertebrate embryos (mammals and birds), the axial tissues of the body form from a growth zone at the tail end, Hensen's node, which generates neural, mesodermal, and endodermal structures along the midline. While most cells only pass through this region, the node has been suggested to contain a small population of resident stem cells. However, it is unknown whether the rest of the node constitutes an instructive niche that specifies this self-renewal behavior.

Cytoskeletal control of early mammalian development.

The cytoskeleton - comprising actin filaments, microtubules and intermediate filaments - serves instructive roles in regulating cell function and behaviour during development. However, a key challenge in cell and developmental biology is to dissect how these different structures function and interact in vivo to build complex tissues, with the ultimate aim to understand these processes in a mammalian organism. The preimplantation mouse embryo has emerged as a primary model system for tackling this challenge.

Keratins are asymmetrically inherited fate determinants in the mammalian embryo.

To implant in the uterus, the mammalian embryo first specifies two cell lineages: the pluripotent inner cell mass that forms the fetus, and the outer trophectoderm layer that forms the placenta. In many organisms, asymmetrically inherited fate determinants drive lineage specification, but this is not thought to be the case during early mammalian development. Here we show that intermediate filaments assembled by keratins function as asymmetrically inherited fate determinants in the mammalian embryo.

Novel approaches to link apicobasal polarity to cell fate specification.

Understanding the development of apicobasal polarity (ABP) is a long-standing problem in biology. The molecular components involved in the development and maintenance of APB have been largely identified and are known to have ubiquitous roles across organisms. Our knowledge of the functional consequences of ABP establishment and maintenance is far less comprehensive.

Specification of the First Mammalian Cell Lineages In Vivo and In Vitro.

Our understanding of how the first mammalian cell lineages arise has been shaped largely by studies of the preimplantation mouse embryo. Painstaking work over many decades has begun to reveal how a single totipotent cell is transformed into a multilayered structure representing the foundations of the body plan. Here, we review how the first lineage decision is initiated by epigenetic regulation but consolidated by the integration of morphological features and transcription factor activity.

Instructions for Assembling the Early Mammalian Embryo.

The preimplantation mouse embryo is a simple self-contained system, making it an excellent model to discover how mammalian cells function in real time and in vivo. Work over the last decade has revealed some key morphogenetic mechanisms that drive early development, yielding rudimentary instructions for the generation of a mammalian embryo. Here, we review the instructions revealed thus far, and then discuss remaining challenges to discover upstream factors controlling cell fate determination, test the role of mechanisms based on biological noise, and take advantage of recent technological developments to advance the spatial and temporal resolution of our current understanding.

Dynamic Fluctuations in Subcellular Localization of the Hippo Pathway Effector Yorkie In Vivo.

The Hippo pathway is an evolutionarily conserved signaling network that integrates diverse cues to control organ size and cell fate. The central downstream pathway protein in Drosophila is the transcriptional co-activator Yorkie (YAP and TAZ in humans), which regulates gene expression with the Scalloped/TEA domain family member (TEAD) transcription factors [1-8]. A central regulatory step in the Hippo pathway is phosphorylation of Yorkie by the NDR family kinase Warts, which promotes Yorkie cytoplasmic localization by stimulating association with 14-3-3 proteins [9-12].

Expanding Actin Rings Zipper the Mouse Embryo for Blastocyst Formation.

Transformation from morula to blastocyst is a defining event of preimplantation embryo development. During this transition, the embryo must establish a paracellular permeability barrier to enable expansion of the blastocyst cavity. Here, using live imaging of mouse embryos, we reveal an actin-zippering mechanism driving this embryo sealing.

Cell Fate Decisions During Preimplantation Mammalian Development.

The early mouse embryo offers a phenomenal system to dissect how changes in the mechanisms controlling cell fate are integrated with morphogenetic events at the single-cell level. New technologies based on live imaging have enabled the discovery of dynamic changes in the regulation of single genes, transcription factors, and epigenetic mechanisms directing early cell fate decision in the early embryo. Here, we review recent progress in linking molecular dynamic events occurring at the level of the single cell in vivo, to some of the key morphogenetic changes regulating early mouse development.

In Vivo Imaging of Single Mammalian Cells in Development and Disease.

Live imaging has transformed biomedical sciences by enabling visualization and analysis of dynamic cellular processes as they occur in their native contexts. Here, we review key recent efforts applying in vivo optical imaging with single-cell resolution to mammalian systems ranging from embryos to adult tissues and organs. We highlight insights into active processes regulating cell fate and morphogenesis during embryonic development, how neuronal circuitry and non-neuronal cell types contribute to neurological functions, and how novel imaging-based approaches enable the dissection of neurological disorders and cancer with high spatio-temporal resolution.

A microtubule-organizing center directing intracellular transport in the early mouse embryo.

The centrosome is the primary microtubule-organizing center (MTOC) of most animal cells; however, this organelle is absent during early mammalian development. Therefore, the mechanism by which the mammalian embryo organizes its microtubules (MTs) is unclear. We visualize MT bridges connecting pairs of cells and show that the cytokinetic bridge does not undergo stereotypical abscission after cell division.

Quantifying transcription factor-DNA binding in single cells in vivo with photoactivatable fluorescence correlation spectroscopy.

Probing transcription factor (TF)-DNA interactions remains challenging in complex in vivo systems such as mammalian embryos, especially when TF copy numbers and fluorescence background are high. To address this difficulty, fluorescence correlation spectroscopy (FCS) can be combined with the use of photoactivatable fluorescent proteins to achieve selective photoactivation of a subset of tagged TF molecules. This approach, termed paFCS, enables FCS measurements within single cell nuclei inside live embryos, and obtains autocorrelation data of a quality previously only attainable in simpler in vitro cell culture systems.