ZNF143 is a transcriptional regulator of nuclear-encoded mitochondrial genes that acts independently of looping and CTCF

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Март 12, 2024

Summary Gene expression is orchestrated by transcription factors, which function within the context of a three-dimensional genome. Zinc finger protein 143 (ZNF143/ZFP143) factor that has been implicated in both gene activation and chromatin looping. To study direct consequences ZNF143/ZFP143 loss, we generated degron line. Our results show depletion no effect on Systematic analysis occupancy data revealed commonly used antibody cross-reacts with CTCF, leading to its incorrect association loops. Nevertheless, specifically activates nuclear-encoded mitochondrial genes loss leads severe dysfunction. Using an vitro embryo model, find essential regulator organismal development. establish as conserved transcriptional cell proliferation differentiation safeguarding activity. Highlights Acute degradation ZFP143 rapid specific transcription. Molecular are inconsistent role proteins. regulation homeostasis critical for multicellular Graphical Abstract

Язык: Английский

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Spatially resolved genome-wide joint profiling of epigenome and transcriptome with spatial-ATAC-RNA-seq and spatial-CUT&Tag-RNA-seq

Nature Protocols, Год журнала: 2025, Номер unknown

Опубликована: Март 21, 2025

Язык: Английский

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Stem cell culture conditions affect in vitro differentiation potential and mouse gastruloid formation

PLoS ONE, Год журнала: 2025, Номер 20(3), С. e0317309 - e0317309

Опубликована: Март 26, 2025

Aggregating low numbers of mouse embryonic stem cells (mESCs) and inducing Wnt signalling generates ‘gastruloids’, self-organising complex structures that display an anteroposterior organisation cell types derived from all three germ layers. Current gastruloid protocols considerable heterogeneity between experiments in terms morphology, elongation efficiency, type composition. We therefore investigated whether altering the mESC pluripotency state would provide more consistent results. By growing lines two different genetic backgrounds intervals ESLIF 2i medium was modulated, culture as well resulting gastruloids were analysed. Microscopic analysis showed a pre-culture-specific effect on formation, aspect ratio reproducibility. RNA-seq start population confirmed short-term pulses modulate state, result cellular states. Since multiple epigenetic regulators detected among top differentially expressed genes, we further analysed genome-wide DNA methylation H3K27me3 distributions. observed differences conditions, most dominantly promoter regions developmental regulators. Lastly, when composition grown these pre-cultures, mESCs subjected to 2i-ESLIF preceding aggregation generated consistently, including mesodermal contributions compared ESLIF-only control. These results indicate optimisation allows modulation differentiation during formation.

Язык: Английский

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Marangoni-like tissue flows enhance symmetry breaking of embryonic organoids

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Сен. 23, 2023

During early development of multi-cellular animals, cells self-organize to set up the body axes, such as primary head-to-tail axis, based on which later plan is defined. Several signaling pathways are known control axis formation. Here, we show, however, that also tissue mechanics plays an important role during this process. We focus emergence a in initially spherical aggregates mouse embryonic stem cells, mirrors events embryo. These break rotational symmetry establish axial organization with domains different expression profiles, e.g. transcription factor T/Bra and adhesion molecule E-cadherin. Combining quantitative microscopy physical modeling, identify large-scale flows recirculation component demonstrate they significantly contribute breaking. show recirculating explained by difference surface tension across domains, akin Marangoni flows, further confirm aggregate fusion experiments. Our work highlights formation not only driven biochemical processes, but it can be amplified flows. expect type amplification may operate many other organoid in-vivo systems.

Язык: Английский

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Control of gastruloid patterning and morphogenesis by the Erk and Akt signaling pathways

Development, Год журнала: 2023, Номер 150(16)

Опубликована: Авг. 15, 2023

ABSTRACT Many developmental processes rely on the localized activation of receptor tyrosine kinases and their canonical downstream effectors Erk Akt, yet specific roles played by each these signals is still poorly understood. Gastruloids, 3D cell culture models mammalian gastrulation axial elongation, enable quantitative dissection signaling patterns responses in a simplified, experimentally accessible context. We find that mouse gastruloids contain posterior-to-anterior gradients Akt phosphorylation induced distinct kinases, with features pattern expression its target Snail exhibiting hallmarks size-invariant scaling. Both contribute to proliferation, whereas also sufficient induce precipitate profound tissue shape changes. further uncover convert entire gastruloid one two mesodermal fates depending position along anteroposterior axis. In all, data demonstrate functional for core development suggest how modules might be harnessed engineer user-defined tissues predictable shapes fates.

Язык: Английский

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Deciphering lineage specification during early embryogenesis in mouse gastruloids using multilayered proteomics

Cell stem cell, Год журнала: 2024, Номер 31(7), С. 1072 - 1090.e8

Опубликована: Май 15, 2024

Gastrulation is a critical stage in embryonic development during which the germ layers are established. Advances sequencing technologies led to identification of gene regulatory programs that control emergence and their derivatives. However, proteome-based studies early mammalian scarce. To overcome this, we utilized gastruloids multilayered mass spectrometry-based proteomics approach investigate global dynamics (phospho) protein expression gastruloid differentiation. Our findings revealed many proteins with temporal unique profiles for each layer, also validated using single-cell technology. Additionally, profiled enhancer interaction landscapes P300 proximity labeling, numerous gastruloid-specific transcription factors chromatin remodelers. Subsequent degron-based perturbations combined RNA (scRNA-seq) identified role ZEB2 mouse human somitogenesis. Overall, this study provides rich resource developmental synthetic biology communities endeavoring understand embryogenesis.

Язык: Английский

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Early autonomous patterning of the anteroposterior axis in gastruloids

Development, Год журнала: 2024, Номер 151(22)

Опубликована: Ноя. 15, 2024

Minimal in vitro systems composed of embryonic stem cells (ESCs) have been shown to recapitulate the establishment anteroposterior (AP) axis. In contrast native embryo, ESC aggregates - such as gastruloids can break symmetry, which is demarcated by polarization mesodermal marker T, autonomously without any localized external cues. However, associated earliest patterning events, spatial restriction cell fates and concomitant transcriptional changes, remain poorly understood. Here, we dissect dynamics AP axis mouse gastruloids, particularly before Wnt stimulation. Through single-cell RNA sequencing, identify key state transitions molecular signatures T+ T- populations underpinning polarization. We also show that this process robust modifications aggregate size. Finally, transcriptomic comparison with embryo indicates develop similar mesendodermal types, despite initial differences their primed pluripotent populations, adopt a more mesenchymal lieu an epiblast-like transcriptome. Hence, our findings suggest possibility alternate states vivo converge onto fates.

Язык: Английский

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Recording morphogen signals reveals origins of gastruloid symmetry breaking

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Июнь 5, 2023

Abstract When cultured in three dimensional spheroids, mammalian stem cells can reproducibly self-organize a single anterior-posterior axis and sequentially differentiate into structures resembling the primitive streak tailbud. Whereas embryo’s body axes are instructed by spatially patterned extra-embryonic cues, it is unknown how these cell gastruloids break symmetry to define (A-P) axis. Here, we use synthetic gene circuits trace early intracellular signals predict cells’ future position gastruloid. We show that Wnt signaling evolves from homogeneous state polarized state, identify critical 6-hour time period when single-cell activity predicts cellular position, prior appearance of patterns or morphology. Single-cell RNA sequencing live-imaging reveal Wnt-high Wnt-low contribute distinct types suggest axial breaking driven sorting rearrangements involving differential adhesion. further extend our approach other canonical embryonic pathways, revealing even earlier heterogeneity TGFβ A-P modulates during period. Our study reveals sequence dynamic processes transform uniform aggregate structure demonstrates morphological emerge out movements absence exogenous patterning cues. Highlights Symmetry-breaking gastruloid protocol where high posterior domain. Synthetic record Wnt, Nodal BMP with resolution. Heterogeneity at 96 h positions types. differences influenced Nodal/BMP activity.

Язык: Английский

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Gastruloid-derived primordial germ cell-like cells develop dynamically within integrated tissues

Development, Год журнала: 2023, Номер 150(17)

Опубликована: Авг. 1, 2023

ABSTRACT Primordial germ cells (PGCs) are the early embryonic precursors of gametes – sperm and egg cells. PGC-like (PGCLCs) can currently be derived in vitro from pluripotent exposed to signalling cocktails aggregated into large bodies, but these do not recapitulate native environment during PGC formation. Here, we show that mouse gastruloids, a three-dimensional model gastrulation, contain population gastruloid-derived PGCLCs (Gld-PGCLCs) resemble PGCs vivo. Importantly, conserved organisation gastruloids leads coordinated spatial temporal localisation Gld-PGCLCs relative surrounding somatic cells, even absence specific exogenous PGC-specific or extra-embryonic tissues. In self-organised interactions between tissues, including endodermal epithelium, enables specification subsequent maturation pool Gld-PGCLCs. As such, represent new source and, owing their inherent co-development, serve as novel study dynamics development within integrated tissue environments.

Язык: Английский

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Coordination between endoderm progression and mouse gastruloid elongation controls endodermal morphotype choice

Developmental Cell, Год журнала: 2024, Номер 59(17), С. 2364 - 2374.e4

Опубликована: Июнь 5, 2024

Язык: Английский

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