Dual role of Ovol2 on the germ cell lineage segregation during gastrulation in mouse embryogenesis

In mammals, primordial germ cells (PGCs), the origin of the germ line, are specified from the epiblast at the posterior region where gastrulation simultaneously occurs, yet the functional relationship between PGC specification and gastrulation remains unclear. Here, we show that Ovol2, a transcription factor conserved across the animal kingdom, balances these major developmental processes by repressing the epithelial-to-mesenchymal transition (EMT) driving gastrulation and the upregulation of genes associated with PGC specification. Ovol2a, a splice variant encoding a repressor domain, directly regulates EMT-related genes and consequently induces re-acquisition of potential pluripotency during PGC specification, whereas Ovol2b, another splice variant missing the repressor domain, directly upregulates genes associated with PGC specification. Taken together, these results elucidate the molecular mechanism underlying allocation of the germ line among epiblast cells differentiating into somatic cells through gastrulation.

Cell lineage specification during development of the anterior lateral plate mesoderm and forelimb field

The lateral plate mesoderm (LPM) is a transient embryonic tissue that gives rise to a diverse range of mature cell types, including the cardiovascular system, the urogenital system, endoskeleton of the limbs, and mesenchyme of the gut. While the genetic processes that drive development of these tissues are well defined, the early cell fate choices underlying LPM development and specification are poorly understood. In this study, we utilize single-cell transcriptomics to define cell lineage specification during development of the anterior LPM and the forelimb field in the chicken embryo. We identify the molecular pathways directing differentiation of the aLPM towards a somatic or splanchnic cell fate, and subsequent emergence of the forelimb mesenchyme. We establish the first transcriptional atlas of progenitor, transitional and mature cell types throughout the early forelimb field and uncover the global signalling pathways which are active during LPM differentiation and forelimb initiation. Specification of the somatic and splanchnic LPM from undifferentiated mesoderm utilizes distinct signalling pathways and involves shared repression of early mesodermal markers, followed by activation of lineage-specific gene modules. We identify rapid activation of the transcription factor TWIST1 in the somatic LPM preceding activation of known limb initiation genes, such as TBX5, which plays a likely role in epithelial-to-mesenchyme transition of the limb bud mesenchyme. Furthermore, development of the somatic LPM and limb is dependent on ectodermal BMP signalling, where BMP antagonism reduces expression of key somatic LPM and limb genes to inhibit formation of the limb bud mesenchyme. Together, these findings provide new insights into molecular mechanisms that drive fate cell choices during specification of the aLPM and forelimb initiation.

Berberine alleviates LPS-induced apoptosis, oxidation, and skewed lineages during mouse preimplantation development

Female infertility is a heterogeneous disorder with a variety of complex causes, including inflammation and oxidative stress, which are also closely associated with the pathogenesis of Polycystic Ovary Syndrome (PCOS). As a new treatment for PCOS, berberine (BER), a natural compound from Berberis, has been clinically applied recently. However, the mechanisms underlying the association between BER and embryogenesis are still largely unknown. In this study, effects of BER on preimplantation development was evaluated by using both normal and inflammatory culture conditions induced by lipopolysaccharide (LPS) in the mouse. Our data first suggest that BER itself (25 nM) does not affect embryo quality or future developmental potency, moreover, it can effectively alleviate LPS-induced embryonic damage by mitigating apoptosis via ROS−/caspase-3-dependent pathways and by suppressing pro-inflammatory cytokines via inhibition of NF-κB signaling pathway during preimplantation embryo development. In addition, skewed cell lineage specification in inner cell mass (ICM) and primitive endoderm (PE) caused by LPS can also be successfully rescued with BER. In summary, these findings for the first time demonstrate the non-toxicity of low doses of BER and its anti-apoptotic and anti-oxidative properties on embryonic cells during mammalian preimplantation development.

MyoD is a 3D genome structure organizer for muscle cell identity

The genome exists as an organized, three-dimensional (3D) dynamic architecture, and each cell type has a unique 3D genome organization that determines its cell identity. An unresolved question is how cell type-specific 3D genome structures are established during development. Here, we analyzed 3D genome structures in muscle cells from mice lacking the muscle lineage transcription factor (TF), MyoD, versus wild-type mice. We show that MyoD functions as a “genome organizer” that specifies 3D genome architecture unique to muscle cell development, and that H3K27ac is insufficient for the establishment of MyoD-induced chromatin loops in muscle cells. Moreover, we present evidence that other cell lineage-specific TFs might also exert functional roles in orchestrating lineage-specific 3D genome organization during development.

SOX17 Is Not Required for the Derivation and Maintenance of Mouse Extraembryonic Endoderm Stem Cell Lines

Extraembryonic endoderm stem (XEN) cell lines can be derived and maintained in vitro and reflect the primitive endoderm cell lineage. SOX17 is thought to be required for the derivation and maintenance of mouse XEN cell lines. Here we have re-evaluated this requirement for SOX17. We derived multiple SOX17-deficient XEN cell lines from preimplantation embryos of a SOX17-Cre knockout strain and chemically converted multiple SOX17-deficient embryonic stem cell lines into XEN cell lines by transient culturing with retinoic acid and Activin A. We confirmed the XEN profile of SOX17-deficient cell lines by immunofluorescence with various markers, by NanoString gene expression analyses, and by their contribution to the extraembryonic endoderm of chimeric embryos produced by injecting these cells into blastocysts. Thus, SOX17 is not required for the derivation and maintenance of XEN cell lines.

Compartmentalization and Interaction of Pax5 and Pax6 in Brain of Mice

Many transcription factors play important roles to maintain the microenvironment, integrity of the blood-brain barrier, the neurons-glia interaction, activities of microglia, composition of cerebrospinal fluid, metabolic activities, concentration of neurotransmitters, presence of inflammatory and anti-inflammatory cytokines, ischemia, stress, aging, neurological disorders, and diseases. The Paired box transcription factors and multifunctional proteins, Pax6 and Pax5 are expressed in brain. They regulate several regulators from cell cycle to cell death. The Pax5, a B-cell lineage-specific activator protein (BSAP), is expressed in the cerebellum, cerebral cortex, hippocampus, olfactory bulb, third ventricles, and choroid plexus. The Pax5 has been observed down-regulated in autism, mental retardation, and Glioblastoma multiforme. The Pax6 affects genes of neurodegeneration, immunological surveillance, and energy homeostasis in brain of mice. The Pax5 and Pax6 recognize several similar DNA sequences and regulate the expression of genes in a tissue-specific manner. Therefore, it is presumed that Pax5 and Pax6, are compartmentalized in brain of mice. Results indicate interactions, cell and tissue-specific compartmentalization, and co-localization of Pax5 and Pax6 in the cerebral cortex, cerebellum, and hippocampus in brain of mice.

LIX1 controls digestive mesenchyme-derived cell fate decision by regulating cristae organization in mitochondria

Limb Expression 1 (LIX1) is a master regulator of digestive mesenchymal progenitor and GastroIntestinal Stromal Tumor (GIST) cell proliferation by controlling the expression of the Hippo effectors YAP1/TAZ and KIT. However, the underlying mechanisms of these LIX1- mediated regulations and tumor promotion remain to be elucidated. Here, we report that LIX1 is S-palmitoylated on cysteine 84 and localized in mitochondria. LIX1 knock-down affects the mitochondrial ultrastructure, resulting in decreased respiration and mitochondrial reactive oxygen species production. This is sufficient to downregulate YAP1/TAZ and reprogram KIT- positive GIST cells towards the smooth muscle cell lineage with reduced proliferative and invasive capacities. Mechanistically, LIX1 knock-down impairs the stability of the mitochondrial proteins PHB2 and OPA1 that are found in complexes with mitochondrial- specific phospholipids and are required for cristae organization. Supplementation with unsaturated fatty acids counteracts the effects of LIX1 knock-down on mitochondrial morphology and ultrastructure, restores YAP1/TAZ signaling, and consequently KIT levels. Altogether, our findings demonstrate that LIX1 contributes to GIST aggressive potential by modulating YAP1/TAZ and KIT levels, a process that depends on mitochondrial remodeling. Our work brings new insights into the mechanisms that could be targeted in tumors in which YAP1 and TAZ are implicated.

Hepatocyte generation in liver homeostasis, repair, and regeneration

The liver has remarkable capability to regenerate, employing mechanism to ensure the stable liver-to-bodyweight ratio for body homeostasis. The source of this regenerative capacity has received great attention over the past decade yet still remained controversial currently. Deciphering the sources for hepatocytes provides the basis for understanding tissue regeneration and repair, and also illustrates new potential therapeutic targets for treating liver diseases. In this review, we describe recent advances in genetic lineage tracing studies over liver stem cells, hepatocyte proliferation, and cell lineage conversions or cellular reprogramming. This review will also evaluate the technical strengths and limitations of methods used for studies on hepatocyte generation and cell fate plasticity in liver homeostasis, repair and regeneration.

From Cell States to Cell Fates: How Cell Proliferation and Neuronal Differentiation Are Coordinated During Embryonic Development

The central nervous system (CNS) exhibits an extraordinary diversity of neurons, with the right cell types and proportions at the appropriate sites. Thus, to produce brains with specific size and cell composition, the rates of proliferation and differentiation must be tightly coordinated and balanced during development. Early on, proliferation dominates; later on, the growth rate almost ceases as more cells differentiate and exit the cell cycle. Generation of cell diversity and morphogenesis takes place concomitantly. In the vertebrate brain, this results in dramatic changes in the position of progenitor cells and their neuronal derivatives, whereas in the spinal cord morphogenetic changes are not so important because the structure mainly grows by increasing its volume. Morphogenesis is under control of specific genetic programs that coordinately unfold over time; however, little is known about how they operate and impact in the pools of progenitor cells in the CNS. Thus, the spatiotemporal coordination of these processes is fundamental for generating functional neuronal networks. Some key aims in developmental neurobiology are to determine how cell diversity arises from pluripotent progenitor cells, and how the progenitor potential changes upon time. In this review, we will share our view on how the advance of new technologies provides novel data that challenge some of the current hypothesis. We will cover some of the latest studies on cell lineage tracing and clonal analyses addressing the role of distinct progenitor cell division modes in balancing the rate of proliferation and differentiation during brain morphogenesis. We will discuss different hypothesis proposed to explain how progenitor cell diversity is generated and how they challenged prevailing concepts and raised new questions.

Fluctuating methylation clocks for cell lineage tracing at high temporal resolution in human tissues

Molecular clocks that record cell ancestry mutate too slowly to measure the short-timescale dynamics of cell renewal in adult tissues. Here, we show that fluctuating DNA methylation marks can be used as clocks in cells where ongoing methylation and demethylation cause repeated ‘flip–flops’ between methylated and unmethylated states. We identify endogenous fluctuating CpG (fCpG) sites using standard methylation arrays and develop a mathematical model to quantitatively measure human adult stem cell dynamics from these data. Small intestinal crypts were inferred to contain slightly more stem cells than the colon, with slower stem cell replacement in the small intestine. Germline APC mutation increased the number of replacements per crypt. In blood, we measured rapid expansion of acute leukemia and slower growth of chronic disease. Thus, the patterns of human somatic cell birth and death are measurable with fluctuating methylation clocks (FMCs).