Cell-Cycle-Dependent ERK Signaling Dynamics Direct Fate Specification in the Mammalian Preimplantation Embryo

Pokrass, Michael; Ryan, Kathleen A.; Xin, Tianchi; Pielstick, Brittany; Timp, Winston; Greco, Valentina; Regot, Sergi

doi:10.1016/j.devcel.2020.09.013

Cited by 75 publications

(91 citation statements)

References 67 publications

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“…Although we observed low-level Spry4 H2BVenus expression within PGCs and PGCLCs, indicating that MAPK signaling may not be entirely shut down, this could also represent perdurance of the Venus reporter. Therefore, future studies using dynamic ERK biosensors [46, 47], combined with time-lapse imaging [48], will uncover precise signaling dynamics. We also observed that the MAPK response was elevated in PGCs within the hindgut endoderm, consistent with studies showing that FGF plays a role in germ cell migration [45, 49].…”

Section: Discussionmentioning

confidence: 99%

Quantitative analysis of signaling responses during mouse primordial germ cell specification

Morgani

Hadjantonakis

2021

Preprint

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During early mammalian development, the pluripotent cells of the embryo are exposed to a combination of signals that drive exit from pluripotency and germ layer differentiation. At the same time, a small population of pluripotent cells give rise to the primordial germ cells (PGCs), the precursors of the sperm and egg, which pass on heritable genetic information to the next generation. Despite the importance of PGCs, it remains unclear how they are first segregated from the soma, and if this involves distinct responses to their signaling environment. To investigate this question, we mapped BMP, MAPK and WNT signaling responses over time in PGCs and their surrounding niche in vitro and in vivo at single-cell resolution. We showed that, in the mouse embryo, early PGCs exhibit lower BMP and MAPK responses compared to neighboring extraembryonic mesoderm cells, suggesting the emergence of distinct signaling regulatory mechanisms in the germline versus soma. In contrast, PGCs and somatic cells responded comparably to WNT, indicating that this signal alone is not sufficient to promote somatic differentiation. Finally, we investigated the requirement of a BMP response for these cell fate decisions. We found that cell lines with a mutation in the BMP receptor (Bmpr1a−/−), which exhibit an impaired BMP signaling response, can efficiently generate PGC-like cells revealing that canonical BMP signaling is not cell autonomously required to direct PGC-like differentiation.

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Section: Discussionmentioning

confidence: 99%

Quantitative analysis of signaling responses during mouse primordial germ cell specification

Morgani

Hadjantonakis

2021

Preprint

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“…2020 JAX#035566 Mouse: ERK-KTR-LoxP Pokrass et al. 2020 N/A Software and algorithms ImageJ/Fiji NIH https://imagej.nih.gov/ij/index.html Deposited data Integrated time-lapse and end-point datasets of ERK KTR Embryos Pokrass et al. 2020 BioImage Archive, Accession S-BIAD28 Other Aspirator tube assemblies for calibrated microcapillary pipettes Sigma Aldrich A5177-5EA Disposable borosilicate glass pasteur pipets Fisher Scientific Cat# 13-678-20C New Brunswick Galaxy 170 R High Capacity CO 2 incubator Eppendorf Cat# 17334002 Nikon SMZ745 stereoscopic microscope Nikon Model: SMZ745 Tokai Hit ThermoPlate Tokai Hit Model: TPi-SMZSSX Nikon Eclipse Ti Microscope Nikon N/A Yokogawa CSU-W1 confocal scanner unit Yokogawa N/A Photometrics Prime 95B sCMOS camera Photometrics N/A Okolab humidified environmental control chamber Okolab Model: H301-K-FRAME …”

Section: Key Resources Tablementioning

confidence: 99%

“…By utilizing different biosensors and fixed readouts, this method allows interrogation of signaling dynamics that specify cell fates in developing embryos. For complete details on the use and execution of this protocol, please refer to Pokrass et al. (2020) .…”

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confidence: 99%

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3D time-lapse microscopy paired with endpoint lineage analysis in mouse blastocysts

Pokrass

Regot

2021

STAR Protocols

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Summary Determining how signaling dynamics relate to gene expression and cell fate is essential to understanding multicellular development. We present a unified live imaging and lineage analysis method that allows integrated analysis of both techniques in the same mouse embryos. This protocol describes the embryo isolation, confocal imaging, immunofluorescence, and in silico alignment required to connect time-lapse and endpoint measurements. By utilizing different biosensors and fixed readouts, this method allows interrogation of signaling dynamics that specify cell fates in developing embryos. For complete details on the use and execution of this protocol, please refer to Pokrass et al. (2020) .

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“…Examples include the signaling pulses observed from the tumor suppressor p53, the mitogen associated protein kinase (MAPK) Erk, and the immune signaling transcription factor NF-κB [1][2][3][4][5] . Pulses of Erk activity have been observed in vivo in the early mouse embryo 6,7 and in 53 tumors 8 , and self-organize into propagating waves from sites of epithelial injury in both mouse 9 54 and zebrafish 10 . The breadth of biological systems exhibiting signaling dynamics suggests that they may play important functional roles.…”

Section: Main Textmentioning

confidence: 99%

A synthetic gene circuit for imaging-free detection of dynamic cell signaling

Ravindran

McFann

Toettcher

2021

Preprint

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Cells employ intracellular signaling pathways to sense and respond to changes in their external environment. In recent years, live-cell biosensors have revealed complex pulsatile dynamics in many pathways, but studies of these signaling dynamics are limited by the necessity of live-cell imaging at high spatiotemporal resolution1. Here, we describe an approach to infer pulsatile signaling dynamics from just a single measurement in fixed cells using a pulse-detecting gene circuit. We computationally screened for circuit with pulse detecting capability, revealing an incoherent feedforward topology that robustly performs this computation. We then implemented the motif experimentally for the Erk signaling pathway using a single engineered transcription factor and fluorescent protein reporter. Our ‘recorder of Erk activity dynamics’ (READer) responds sensitively to both spontaneous and stimulus-driven Erk pulses. READer circuits thus open the door to permanently labeling transient, dynamic cell populations to elucidate the mechanistic underpinnings and biological consequences of signaling dynamics.

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Cell-Cycle-Dependent ERK Signaling Dynamics Direct Fate Specification in the Mammalian Preimplantation Embryo

Cited by 75 publications

References 67 publications

Quantitative analysis of signaling responses during mouse primordial germ cell specification

Quantitative analysis of signaling responses during mouse primordial germ cell specification

3D time-lapse microscopy paired with endpoint lineage analysis in mouse blastocysts

A synthetic gene circuit for imaging-free detection of dynamic cell signaling

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