Nuclear and cytosolic fractions of SOX2 synergize as transcriptional and translational co-regulators of cell fate.

Stemness and pluripotency are mediated by transcriptional master regulators that promote self-renewal and repress cell differentiation, among which is the high-mobility group (HMG) box transcription factor SOX2. Dysregulated SOX2 expression, by contrast, leads to transcriptional aberrations relevant to oncogenic transformation, cancer progression, metastasis, therapy resistance, and relapse. Here, we report a post-transcriptional mechanism by which the cytosolic pool of SOX2 contributes to these events in an unsuspected manner.

Osteopontin drives neuroinflammation and cell loss in MAPT-N279K frontotemporal dementia patient neurons.

Frontotemporal dementia (FTD) is an incurable group of early-onset dementias that can be caused by the deposition of hyperphosphorylated tau in patient brains. However, the mechanisms leading to neurodegeneration remain largely unknown. Here, we combined single-cell analyses of FTD patient brains with a stem cell culture and transplantation model of FTD.

The homeobox transcription factor DUXBL controls exit from totipotency.

In mice, exit from the totipotent two-cell (2C) stage embryo requires silencing of the 2C-associated transcriptional program. However, the molecular mechanisms involved in this process remain poorly understood. Here we demonstrate that the 2C-specific transcription factor double homeobox protein (DUX) mediates an essential negative feedback loop by inducing the expression of DUXBL to promote this silencing.

Highly cooperative chimeric super-SOX induces naive pluripotency across species.

Our understanding of pluripotency remains limited: iPSC generation has only been established for a few model species, pluripotent stem cell lines exhibit inconsistent developmental potential, and germline transmission has only been demonstrated for mice and rats. By swapping structural elements between Sox2 and Sox17, we built a chimeric super-SOX factor, Sox2-17, that enhanced iPSC generation in five tested species: mouse, human, cynomolgus monkey, cow, and pig. A swap of alanine to valine at the interface between Sox2 and Oct4 delivered a gain of function by stabilizing Sox2/Oct4 dimerization on DNA, enabling generation of high-quality OSKM iPSCs capable of supporting the development of healthy all-iPSC mice.

Mechanisms, pathways and strategies for rejuvenation through epigenetic reprogramming.

Over the past decade, there has been a dramatic increase in efforts to ameliorate aging and the diseases it causes, with transient expression of nuclear reprogramming factors recently emerging as an intriguing approach. Expression of these factors, either systemically or in a tissue-specific manner, has been shown to combat age-related deterioration in mouse and human model systems at the cellular, tissue and organismal level. Here we discuss the current state of epigenetic rejuvenation strategies via partial reprogramming in both mouse and human models.

Human skeletal muscle organoids model fetal myogenesis and sustain uncommitted PAX7 myogenic progenitors.

In vitro culture systems that structurally model human myogenesis and promote PAX7 myogenic progenitor maturation have not been established. Here we report that human skeletal muscle organoids can be differentiated from induced pluripotent stem cell lines to contain paraxial mesoderm and neuromesodermal progenitors and develop into organized structures reassembling neural plate border and dermomyotome. Culture conditions instigate neural lineage arrest and promote fetal hypaxial myogenesis toward limb axial anatomical identity, with generation of sustainable uncommitted PAX7 myogenic progenitors and fibroadipogenic (PDGFRa+) progenitor populations equivalent to those from the second trimester of human gestation.

Evaluation of the determinants for improved pluripotency induction and maintenance by engineered SOX17.

An engineered SOX17 variant with point mutations within its DNA binding domain termed SOX17FNV is a more potent pluripotency inducer than SOX2, yet the underlying mechanism remains unclear. Although wild-type SOX17 was incapable of inducing pluripotency, SOX17FNV outperformed SOX2 in mouse and human pluripotency reprogramming. In embryonic stem cells, SOX17FNV could replace SOX2 to maintain pluripotency despite considerable sequence differences and upregulated genes expressed in cleavage-stage embryos.

Electrophysiological Properties of Tetraploid Cardiomyocytes Derived from Murine Pluripotent Stem Cells Generated by Fusion of Adult Somatic Cells with Embryonic Stem Cells.

Most cardiomyocytes (CMs) in the adult mammalian heart are either binucleated or contain a single polyploid nucleus. Recent studies have shown that polyploidy in CMs plays an important role as an adaptive response to physiological demands and environmental stress and correlates with poor cardiac regenerative ability after injury. However, knowledge about the functional properties of polyploid CMs is limited.

A mesh microelectrode array for non-invasive electrophysiology within neural organoids.

Organoids are emerging in vitro models of human physiology. Neural models require the evaluation of functional activity of single cells and networks, which is commonly measured by microelectrode arrays. The characteristics of organoids clash with existing in vitro or in vivo microelectrode arrays.

Homogeneity of XEN Cells Is Critical for Generation of Chemically Induced Pluripotent Stem Cells.

In induced pluripotent stem cells (iPSCs), pluripotency is induced artificially by introducing the transcription factors , , , and . When a transgene is introduced using a viral vector, the transgene may be integrated into the host genome and cause a mutation and cancer. No integration occurs when an episomal vector is used, but this method has a limitation in that remnants of the virus or vector remain in the cell, which limits the use of such iPSCs in therapeutic applications.

MAPT genotype-dependent mitochondrial aberration and ROS production trigger dysfunction and death in cortical neurons of patients with hereditary FTLD.

Tauopathies are a major type of proteinopathies underlying neurodegenerative diseases. Mutations in the tau-encoding MAPT-gene lead to hereditary cases of frontotemporal lobar degeneration (FTLD)-tau, which span a wide phenotypic and pathological spectrum. Some of these mutations, such as the N279K mutation, result in a shift of the physiological 3R/4R ratio towards the more aggregation prone 4R isoform.

Generation of Induced Pluripotent Stem Cells from Lymphoblastoid Cell Lines by Electroporation of Episomal Vectors.

Background And Objectives: Lymphoblastoid cell lines (LCLs) deposited from disease-affected individuals could be a valuable donor cell source for generating disease-specific induced pluripotent stem cells (iPSCs). However, generation of iPSCs from the LCLs is still challenging, as yet no effective gene delivery strategy has been developed.

Methods And Results: Here, we reveal an effective gene delivery method specifically for LCLs.

Early developmental plasticity enables the induction of an intermediate extraembryonic cell state.

Two fundamental elements of pre-implantation embryogenesis are cells' intrinsic self-organization program and their developmental plasticity, which allows embryos to compensate for alterations in cell position and number; yet, these elements are still poorly understood. To be able to decipher these features, we established culture conditions that enable the two fates of blastocysts' extraembryonic lineages-the primitive endoderm and the trophectoderm-to coexist. This plasticity emerges following the mechanisms of the first lineage segregation in the mouse embryo, and it manifests as an extended potential for extraembryonic chimerism during the pre-implantation embryogenesis.

Embryonic Stem Cells Lacking DNA Methyltransferases Differentiate into Neural Stem Cells that Are Defective in Self-Renewal.

Background And Objectives: DNA methyltransferases (Dnmts) play an important role in regulating DNA methylation during early developmental processes and cellular differentiation. In this study, we aimed to investigate the role of Dnmts in neural differentiation of embryonic stem cells (ESCs) and in maintenance of the resulting neural stem cells (NSCs).

Methods And Results: We used three types of Dnmt knockout (KO) ESCs, including Dnmt1 KO, Dnmt3a/3b double KO (Dnmt3 DKO), and Dnmt1/3a/3b triple KO (Dnmt TKO), to investigate the role of Dnmts in neural differentiation of ESCs.

Deterministic programming of human pluripotent stem cells into microglia facilitates studying their role in health and disease.

Microglia, the resident immune cells of the central nervous system (CNS), are derived from yolk-sac macrophages that populate the developing CNS during early embryonic development. Once established, the microglia population is self-maintained throughout life by local proliferation. As a scalable source of microglia-like cells (MGLs), we here present a forward programming protocol for their generation from human pluripotent stem cells (hPSCs).

OCT4 interprets and enhances nucleosome flexibility.

Pioneer transcription factors are proteins that induce cellular identity transitions by binding to inaccessible regions of DNA in nuclear chromatin. They contribute to chromatin opening and recruit other factors to regulatory DNA elements. The structural features and dynamics modulating their interaction with nucleosomes are still unresolved.

Induced Endothelial Cell-Integrated Liver Assembloids Promote Hepatic Maturation and Therapeutic Effect on Cholestatic Liver Fibrosis.

The transplantation of pluripotent stem cell (PSC)-derived liver organoids has been studied to solve the current donor shortage. However, the differentiation of unintended cell populations, difficulty in generating multi-lineage organoids, and tumorigenicity of PSC-derived organoids are challenges. However, direct conversion technology has allowed for the generation lineage-restricted induced stem cells from somatic cells bypassing the pluripotent state, thereby eliminating tumorigenic risks.

Generation of a human iPSC line (MPIi008-A) from a patient with Denys-Drash syndrome.

An induced pluripotent stem cell (hiPSC) line (MPIi008-A) was generated from fibroblasts of a 1-year-old male patient with Denys-Drash syndrome using lentiviral delivery of reprogramming factors OCT4, SOX2, KLF4 and c-MYC. The MPIi008-A iPSC line exhibited typical iPSC morphology and normal karyotype, expressed pluripotent stem cell markers, and showed developmental potential to differentiate into derivatives of all three germ layers in vivo. The hiPSC line harbours a heterozygous missense mutation (R394L) in exon 9 of the WT1 gene.

Oct4 differentially regulates chromatin opening and enhancer transcription in pluripotent stem cells.

The transcription factor Oct4 is essential for the maintenance and induction of stem cell pluripotency, but its functional roles are not fully understood. Here, we investigate the functions of Oct4 by depleting and subsequently recovering it in mouse embryonic stem cells (ESCs) and conducting a time-resolved multiomics analysis. Oct4 depletion leads to an immediate loss of its binding to enhancers, accompanied by a decrease in mRNA synthesis from its target genes that are part of the transcriptional network that maintains pluripotency.

Tissue clearing may alter emission and absorption properties of common fluorophores.

In recent years, 3D cell culture has been gaining a more widespread following across many fields of biology. Tissue clearing enables optical analysis of intact 3D samples and investigation of molecular and structural mechanisms by homogenizing the refractive indices of tissues to make them nearly transparent. Here, we describe and quantify that common clearing solutions including benzyl alcohol/benzyl benzoate (BABB), PEG-associated solvent system (PEGASOS), immunolabeling-enabled imaging of solvent-cleared organs (iDISCO), clear, unobstructed brain/body imaging cocktails and computational analysis (CUBIC), and ScaleS4 alter the emission spectra of Alexa Fluor fluorophores and fluorescent dyes.

SARS-CoV-2 infects and replicates in photoreceptor and retinal ganglion cells of human retinal organoids.

Several studies have pointed to retinal involvement in COVID-19, yet many questions remain regarding the ability of SARS-CoV-2 to infect and replicate in retinal cells and its effects on the retina. Here, we have used human pluripotent stem cell-derived retinal organoids to study retinal infection by SARS-CoV-2. Indeed, SARS-CoV-2 can infect and replicate in retinal organoids, as it is shown to infect different retinal lineages, such as retinal ganglion cells and photoreceptors.